AU778585B2 - Secreted and transmembrane polypeptides and nucleic acids encoding the same - Google Patents

Description

AUSTRALIA

Patents Act 1990 COMPLETE SPECIFICATION STANDARD PATENT Applicant(s): GENENTECH, INC.

Invention Title: SECRETED AND TRANSMEMBRANE POLYPEPTIDES AND NUCLEIC ACIDS ENCODING THE SAME The following statement is a full description of this invention, including the best method of performing it known to me/us: la SECRETED AND TRANSMEMBRANE POLYPEPTIDES AND NUCLEIC ACIDS ENCODING THE SAME FIELD OF THE INVENTION The present invention relates generally to the identification and isolation of novel DNA and to the recombinant production of novel polypeptides encoded by that DNA.

BACKGROUND OF THE INVENTION All references, including any patents or patent applications, cited in this specification are hereby incorporated by reference. No admission is made that any reference constitutes prior art. The discussion of the references states what their authors assert, and the applicants 15 reserve the right to challenge the accuracy and pertinency of Sthe cited documents. It will be clearly understood that, although a number of prior art publications are referred to herein, this reference does not constitute an admission that any of these documents forms part of the common general knowledge in the art, in Australia or in any other country.

Extracellular and membrane-bound proteins play o important roles in the formation, differentiation and maintenance of multicellular organisms. The fate of many individual cells, proliferation, migration, 25 differentiation, or interaction with other cells, is typically governed by information received from other cells and/or the immediate environment. This information is often transmitted by secreted polypeptides (for instance, mitogenic factors, survival factors, cytotoxic factors, differentiation factors, neuropeptides, and hormones) which are, in turn, received and interpreted by diverse cell receptors or membrane-bound proteins. These secreted polypeptides or signaling molecules normally pass through the cellular secretory pathway to reach their site of action in the extracellular environment, usually at a membrane-bound receptor protein.

Secreted proteins have various industrial applications, including use as pharmaceuticals, diagnostics, biosensors and bioreactors. In fact, most protein drugs available at present, such as thrombolytic agents, interferons, Ib interleukins, erythropoietins, colony stimulating factors, and various other cytokines, are secretory proteins. Their receptors, which are membrane-bound proteins, also have potential as therapeutic or diagnostic agents. Receptor immunoadhesins, for instance, can be employed as therapeutic agents to block receptor-ligand interaction. Membrane-bound proteins can also be employed for screening of potential peptide or small molecule inhibitors of the relevant receptor/ligand interaction. Such membrane-bound proteins and cell receptors include, but are not limited to, cytokine receptors, receptor kinases, receptor phosphatases, receptors involved in cell-cell interactions, and cellular adhesin molecules like selectins and integrins. Transduction of signals that regulate.cell growth S. and differentiation is regulated in party by phosphorylation of 15 various cellular proteins. Protein tyrosine kinases, enzymes that catalyze that process, can also act as growth factor receptors. Examples include fibroblast growth factor receptor and nerve growth factor receptor.

Efforts are being undertaken by both industry and academia to identify new, native secreted and membrane-bound receptor proteins. Many efforts are focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel secreted and membrane-bound receptor proteins. Examples of screening methods and techniques are 25 described in the literature [see, for example, Klein et al., Proc. Natl. Acad. Sci., 93:7108-7113 (1996); U.S. Patent No.

5,536,637].

We herein describe the identification and characterization of novel secreted and transmembrane polypeptides and novel nucleic acids encoding those polypeptides.

The entire disclosure in the complete specification of our Australian Patent Application No.

93178/98 is by this cross-reference incorporated into the present specification.

I. PROII I and PRO217 Epidermal growt1h factor (EGF) is a orwczuional mizogenic factor that stimulates the proliferation of various types of cells indhidir epithielial cells and fibroblasts. EGF binds to a&d activates dhe EGF receptor (EUPR), which initiates intracellular signaling and subsequent effects. The EGFR is expressed in neurons of the ccrtbral cortex.

crbelum, .Lnd hippocazupus in addition to other regions of the central nervous system (CNS). In addition, EGF is also expressed in various regions of the CNS. Therfore, EGF acts not oily on rmitotic cells, but also on postroitotic neurons. In fact, many studies have indicated tat EGF has reurotrophic or neuromodulatory effects on various types of neurons in the CNS. For example. EGF acts directly on cultured cerebral cortical and cerebellar neurons, enhancing neurite outgrowth and survival. On the oter hand. EGF also acts on other cell types. including septa4 eholiriergic arnd -tesneephalic dopaminergic neutrons, indirctly through glial cells. Evidence of the effects of EGF on neurons in the CNS is accumulating, but the meachanisms of action remain essentially unknown. EGF-induced signalin in mitotic cells is betterunderstood than in postriniod neurons. Stuidies of doed pheochromocytona PC12 cells anid cultured cerebral cortical neurons have suggested that the EGP-induced neurotiuphic actions are mediated by sustained activation of the EGFR and snitogen-activated protein kinase (MAPK) in response to EGF. The Mutied intracellular signalin correlates with the decreased rate of EGFR down-regulation, which maight determine 15 the response of neuronal cells to EGF. It is likely that EGE is a multi-potent growthi factor that acts upon various types ofcells including mitotic cells and postanitotic neurons.

EGF is produced by the salivary and Brunner's glands of the gastrointestinal system. kidney, pancreas, thyroid gland, pituitary gland, and the nervous system, and is found in body fluids such as saliva, blood, crebrospinal fluid (CSP), urine, amniotic fluid, prostatic fluid, pancereatic juice, and breast milk, Plata-Salasnan.

20 Peptides .12: 653-663 (1991).

EGF is mediated by its menrtanm specific receptor, which contains an intrinsic tyrosine kinase. Stoscheck er at. J. Cell Biochem. Ul: 135-152 (1986). EGF is believed to function by binding to the,extracellular portion of its receptor which induces a transinembratm signal that activates the intrinsic tyrosine kinase.

Purification and sequence analysis of the EGF-like domain has revealed die presece of six conserved 25 cysrene residues which cross-bind to create three peptide loops, Savage et J. W. Client. 241: 7669-7672 (1979).

It is now generally known that several other peptides can react with the EGF receptor which share the same generalized motif X CC,,CX 10 CXCXSGXCX,,, where X represents any nora-cysteine amino acid, arnd n is a variable repeat mabr. Non isolated peptides having this motif include TGF-a. amphiregulin, schwartnoma-derived growth factor (SDGF),. heparin-binding EGF-like growth factors and certain virally encoded peptddes Vaccinia virus, Reisner, Nature 2n3: 801-803 (19815), Shope fibroma virus, Clung et al., MoI Cell Biol. 7: 535-540 (1987).

Molluscum contagisum, Porter and Ardiard, Gen. Viral. ~fi 673-68 (1987). and Myxoana virus, Upton a1 al., f. Virol. fil: 1271-1275 (1987), Prigent and Lesnoine, Prog. Growj Factor Res. 1-24 (1992).

EGF-like domains arc not confined to growth factors but have been observed in a variety of cell-surface: and extracellilar proteins which have in tng properties in cell adhesion, protein-protein interaction apad development, Lmarerme arid Gusterson, Tumor Biol. 11: 229-261 (1990). These proteins include blood coagulation factors (factors VI. Ex. X. XII. protein C, protein S. protein Z. tissue plasminogen activator, urokinase), extracellular matrix components (lamninin, cytotactin. entactin), cell surface receptors (LDL receptor, thrombomodulin receptor) and immunity-related proteins (complement CMr. uromnodulin).

Even more interesting. the gcneral structure pattern of EGP-like precurors is Preserved through lower organisins as well as in mnammalian cells. A number of genes with developmental signficanCe have been identified in invertebrates with EUF-Like repeats. For example. the notch gene of Drosophila encodes 36 tandemly arranged amino acid repeats which show homology to EGF. Wharton ei aL. Cell 557-581 (1985). Hydropathy plots indicate a putative membrane spanning domain, with the EGF-rdated sequences being located on die extracellular side of the membrane. Other homeotic genes with EGF-like repeats include Delta, 95F and SZD which were identified using probes based on Notch, and the nemnatode gene Lin-)2 which encodes a putative receptor for a developmental signal transmitted between two specified cells.

Specifically. EGF has been shown to have potential in the preservation and maintenance of gastrointestinal anicosa and the repair of acute and chronic mucosal lesions, Konturek et at.. Eur. J. Gas troente rot Repaid. 72(10).

933-37 (1995), including the treatment of necrotizing eniteracoids, Zollinger-Ellison syndrome, gastrointestinal *.*ulceration gastrointestinal ulcerations and congenital microvillus atrophy, Guglienta and Sullivan. Eur. 1.

Gwsromerul Heparol, 2(10), 945-50 (1995). Additionally, EGF has been implicated in hair follicle difftrentiation; %:du Cros, J. Invest. Dermato. IU~ (U Suppl.), 106S-1 13S (1993), Hillier, GIn. Endocrinol. 427-28 (1990); kidney function, Harner al.. Semins. Nqphrol. 13 109-15 (1993), Harris, Am. J. Kidney Duj. 11(6):,627-30 (1991); tear fluid, van Setten et at., mI. Ophrhalmol 359-62 (1991); vitamin K mediated blood coagulation, Steflo a aL. Blood7.fl(7: 1637-51 (1991). EGF is also implicated various skin disease characterized by Abnormal keratinocyte differentiation, psoriasis, epithlal cancers such as squamous cell Carcinomas Of the lung, epidcrnid carcinoma of the vulva and gliouas. King ar al., Am. J. Med. Sri. 226: 154-158 (1988).

.:20 Of great interest is mounting evidence that genetic alterations in growth factors signaling pathways are closely linked to developmental abnormalities and to chronic diseases including Cancer. Aaronson, Science ZM: 1146-1153 (1991). For example, c-erb-2 (also known as HER-2), a protO-oncogene with close structural similarity :to EGFecoepto: protein. is ov~mesod in human breast cancer. King ar al., Sciatce 22: 974-976 (1985); Gullick, Hormnerandther atios, ook etat.. eds, Amsterdam,. Elsevier. pp 349-360 (1986).

We herein describe the identification and characterization of novel polypeptides having homology to EGF, wherein those polypeptides are herein designated PRO211I and PRO2 17.

PRO230 Nephrids is a condition characterized by inflammation of the kidney affecting the structure and normal function of the kidney. This condition can be chronic or acute and is geneally caused by infection, degenerative process or vascular disease. In all case, early detection is desirable so that the patient with nephids can begin treatment of the condition.

An approach to detecting nephritis is to determine the antigens associated with nephritis and antibodies thereto. In rabbit, a tubulointerstitial nephzitis antigen (TIN-ag has been reported in Nelson, T. et al.. 1. Biol.

akm, 270c27):im26570 (July 1995) (GENBANK1J24270). This stud reports that the rabbit TIN-ag is a basement membrane glycoprotein having a preicted amnino, acid sequenc which has a carboxylkermiial region exhibiting homology with human preprocathepsin B, a member of the cystein proteinase family of proteins. it is also reported that the rabbit TIN-ag has a domain in the amino4erminal region containing an epidermal growth factor-like motif that shares bomology with laminin A and S chains, alpha 1 chain of type I collagen, von Willebrand's factor and mucin, indicating structural and functional similarities. Studies have also been conducted in mice. However, it is desirable to identify tubulointerstitial nephritis antigens in humans to aid in the development of early detection methods and treatment of nephritis.

Proteins which have homology to ubulonterstiial nephritis antigens are of particular interest to the medical and industrial communities. Often, proteins having homology to each other have similar function. It is also of inerest when proteins having homology do not have similar functions, indicating that certain structural motifs identify information other than function, such as locality of function. We herein describe the identification and characterization of a novel polypeptide, designated hgcrcin as PR0230, which has homology to tubulointerstitial nephritis antigens.

3. PRO232 Stem cells are undifferentiated cells capable of proliferation, self maintenance, the production of a large nmnber of differentiated functional progeny, regeneration of tissue after injury and/or a flexibility in the use of these options. Stem cells often express cell surface antigens which are capable of serving as cell specific markrs that can be exploited to identify stem cells, thereby providing a means for identifying and isolating specific stem cell populations.

Having possession of differnt stem cell populations will allow for a number of important applications. For example, possessing a specific stem cell population will allow for the identification of growth factors and other S. 20 proteins which arc involved in their proliferation and differentiation. In addition, there may be as yet undiscovered proteins which are associated with the early steps of dedication of the stem cell to a particular lineage, (2) prevention of such dedication, and negative control of stem cell proliferation, all of which may be identified if one has possession of the stem cell population. Moreover, stem cells are important and ideal targets for gene therapy where the inserted genes promote the health of the individual into whm the stem cells are transplanted. Finally, stem cells may play important roles in transplantation of organs or tissues, for example liver regeneration and skin grafting.

Given the importance of stem cells in various different applications efforts are currently being undertaken S by both industry and academia to identify new, native stem cell antigen proteins so as to provide specific cell surface markers for identifying stem cell populations as well as for providing insight into the functional roles played by stem cell antigens in cell proliferation and differentiation. We herein describe the identification and characterization of novel polypeptides having homology to a stem cell antigen, wherein those polypcptides are herein designated as PR0232 polypeptides..

4. PROQ87 Growth factors are molecular signals or mediators that enhance cell growth or proliferation, alone or in concert, by binding to specific cell surface receptors. However, there are other cellular reactions than only growth upon expression to growth factors. As a result, growth factors are better characterized as multifunctional and potent cellular reguLitors. Their biological effects include proliferation, c-emoaxis and stimulation of extracellular matrix production. Growth factors can have both stiunulatozy and inhibitory effects. For example, transforming growth factor (rGFI)) is highly pleiatrapic and can stimulate proliferation in 3oMe cells, especially connective tissue, while being a potent inhibitor of' proliferation in others, such as lymphocytes and epithelial cells.

T*Re physiological effect of growth stimulation or inhibition by growth factors depends upon the state of developmntt and differentiation of the targt tissue. The mechanism of local cellular regulation by classical endocrine miolecules involves comprehends autocrine (same cell), juxtacrine (neighbor cell), and paracrinc (adjacent cells) pathways. Pepdec growth factors ame elements of a complex biological language, providing the basis for intercellular Tacy permit cells to convey information between each other, mediate interaction between cells ard change gene expression. The effect of these multil'unctional and phiripotent factors is dependent on the presetce or absence of other peptides.

FGF-8 is a member of the fibroblast growth factors (FGFz) which are a family of heparin-binding, potent rntogens for both normal diploid fibroblasts and established cell lines. Gospodarowicz ar at. (1984). Proc. Nat!.

Acad. Sd. MS 11:6W6. The FGF family comprises acidic FGF (FGF-l), basic FGF (FPGF-2). INT-2 (FGF-3). K- FGFIHST (FGF.4), FGF-5. FGF-6. KGF (FGF-7). AIGF (FGF48) among others. All FGFs have two conserved cysteine residues and share 30-50% sequence homology at the amino acid level. These factors are mitogenic for a wide variety of normal diploid mesoderm-derived and neural crest-derived cells, including granulosa cells, adrenal cortical cells, chondrocytses, inyoblasts, cornea] and vascular endothelial cells (bovine or human), vascular smooth rmiscle cells, lens, retina and prostatic epithelial cells. oligodendrocytes, astrocytes, chrondocytes, myoblasts and osteoblasts.

.20 Fibroblast growth factors can also stinuiflate a large number of cell types in a non-mitogenic manner. These activities includ promotion of cell migration into wounds ame (chemotaxis), initiation of new blood vessel fominulation (argiogeirsis). modulation of nerve regeneration and survival (neurotrophism), modulation of endocrine functions, :and stimulation or suppression of specific cellular protein expression, extracell ular mnatrix production and cell survival. Baird Bohlen, Handbook of Exp. Pharmacol. 95(l): 369-418. Springer. (1990). These properties provide a basis for using fibroblast growth factors in therapeutic approaches to accelerate wound healing, nerve repair.

collateral blood vessel formation, and the like. For example, fibroblast growth factors have been suggested to mimiz ocn~ard4him damage in heart disease and su...y 4,378,347).

FGF-8, also kno~wn as androgen-induced growth factor (AIGF), is a 2 15 amino acid protein which shares 30-40% sequence homology with the other members of the FGF family. FGF-8 has been proposed to be under androgenic regulation and induction in the mouse mammary carcaamna cell line SCI. Tanaka ar aL. Proc. Nat!.

Aca. Sd. USA Z2: 8928-8932 (199); Sato ea J. Steroid Biochwm. Molec. BioL 42: 91-98 (199). As a result, FGF-8 may have a local role in the prostate, which is kinown to be an androgen-responsive organ. FGF-8 can also be oncogenic, as it displays tansforming activity when transfected into NIH-373 fibroblasts. Kouhara ar 02scogwn 2 455-462 (1994). While FGF-8 has been detected in heart brain, lung. kidney, testis. prostate and ovary, expression was also detected in the absence of exogenous: androgens. Schmitt a at.. J. Steroid Blochem. MoL Biol.

.U 173-78 (1996).

FGF-8 shares ie property with several other FG~s of being expressed at a variety of stages of murine embryogeneis. which supports the theory that the various FGFs have multiple and perhaps coordinated role& in differentiation and cmbryogenesis. Moreover, FGP-8 has also been identified as a protoncgcnc that cooperates with Wr±.l in the process of mmary tuzmrigencsis (Shackleford et at.. Proc. NaI. A cad. Sci. USA 2Q, 740-744 (1993); Heikinheimo el aL. Mech. Dev. 129-138 (1994)).

In contrast to the other FOFs, FGF-B exists as three protein isoforms, as a result of alternative splicing of thc primary transcript. Tanaka et al.. sup ra. Normal adult expression of FGF-8 is weak and confined to gonadal tissue, however northern blot analysis has indicated that FOP-8 niRNA is present from day 10 through day 12 or snurie gestation, which suggests that FGF-8 is important to normal development. Heiltinheirno et al.. Mich Dev.

4B(2): 129-38 (1994). Further in .lu hybridization assays between day 8 and 16 of gestation indicated initial expression in the surface etodenn of the first bronchiial arrhes the frontnasal process. the forebrain and the midbrairt-hindbmain junction, At days 10-12, FGF-8 was expressed in the surface ectoderm of the forelimb and hindlinib buds. the nasal its and nasopharynx. the infuzaiibulurn and in the telencphalon, diencphalon and ce halon. Expression contimies in dhe developing hindlimba through day 13 of gestation, but is undetectable dirtafter. The results suggest that FGF-8 has a unique temporal and spatial pattern in embryogenesis and suggests a role for this growth factor in multiple regions of ectodernial differentiation in the post-gastrulation embryo.

We herein describe the identification of novel poypeptides having homology to FGP-8, wherein those polypeptides are heein designated PRO 187 polypeptdecs.

S. PR26 :20 Protein-protein interactions include rceptor and antigen complexes and signaling mechanisms. As more is known about the structural and functional mechanisms underlying protein-protein interactions, protein-protein interactions can be morm easily manipulated to regulate the particular result of the protein-protein interaction. Thus.

the underlying mechanisms of protein-protein interactions are of interest to tie scientific and medical communnity.

SAll proteins containing leucane-ric repeats are thought to be involved in protein-protein interactions.

Lcucmne-rich repeats are short sequence motits present in a mnber of proteins with diverse functions and cellular locations. The crystal structure of ribonucase inhibitor protein has revealed that leucine-rich repeats correspond to beta-alpha structural units. These units are arranged so that they form a parallecl beta-sheet with one surface exposed to solvent, so that the protein acquires an unusual, nonglubular shape. These two features have been indicated as responsible for the protein-binding functions of proteins containing leucine-ricli repeats. See, Kobe and Deisenhofer, Trends iochem- Sci., 19(10):415-421 (Oct. 1994).

A study has been reported on leucirt-rich proteoglycans which serve as tissue organizers. orienting and ordering collagen fibrils during ontogeny and are involved in pathological processes such as wound healing, tissue repair, anid tumor stroma formation. lozzo, R. Crit. Rev. Bochem. Mol. iol., 32(2):141-174 (19917). Others studies implicating leucine rich proteins in wound healing and tissue repair are Do LA Salle, et al.. XgI. Rey Er. HeEatol. (Germany). 37(4):215-2 (1995), reporting mautations in the leucine rich motif in a complex associated with the bleeding disorder Bemnard-Soulier syndromne and Chlemetson, K. Ib~b arot (Germany), 74<1): 111-116 (July 1995). reporting that platelets have leuciac rich repeats. Another protein of particular interest which has been reported to have Iecane-rich repeats is the SlIT protein which has been reported to be useful in treating ne~gencrative diseases such as Alzheimer's disease, nerve damage such as in Parkinson's disease, and fordianois f ancrsee, Artavanistsakonas. S. and Rotberg. J. W09210518-Al by Yale University. Other studics reporting on the biological funetons of proteins having leucine-rich repeats include: Tayar. et al.. MQI CcI ndocrnoI., (Ireland). 125(1-2):65-70 (Dec. 1996) (gonadoiropin receptor involvement); Miura, et al., Vji22aiMliW (Japan). 54(7):1784-1789 (July 1996) (apoptosis brvolvement); Harris. P. et al., J.A .Sc NcibI., 6(4):1125-1133 (Oct. 199.5) (kidney disease involvement); and Ruosaixi, E. et al., W091 10727-A by LA Jolla Can= Rsearch Founidation (decorin binding to transforming growth factor-n involvement for treatment for canc=, twvwi healing and scarring). Also of particular interest is fibromodulin and its use to prevent or reduce dermal scarring. A study of fibromodulin is found in U.S. Patent No. 5,654.270 to Ruoslabri. et al.

Efforts are therefore being undertake by both industry and academia to identify new proteins having leucine rich repeats to better undlerstand protein-protein interactions. Of particular interest are those proteins having leucine rich repeats and homology to knmwn proteins having leucine rich repeats such as fibrornodulin, the SUTr protein and platelet glycoprotein V. Many efforts are focused on the screening of mamnmalian recmbinant DNA libraries to identify dhe coding sequences for novel secreted and membrane-bound proteins having leucine rich repeats. We herein describe the identification and chamacterization of novel polypeptides having homology to fibronrodulin, herein designated as PR0265 polypeptides.

6. PRO.219 Human matrilin-2 polypeptide is a member of the von Willebrand factor type A-like module superfamily.

.20 von Wiliebrand factor is a protein wich plays an important role in the maintentence of hem stasis. More specifically, von Willebrand factor is a protein which is known to participate in phtelet-vessd wall interactions at the rite of vascular Injury via its ability to ulerac and form a complex with Factor VIII. The absence of von Willebrand factor in the blood causes an abnormality with the blood platelets that prevents platelet adhesion to die vascular wall at the site of the vascular injury. The result is the propensity for brusing, nose bleeds, intestinal bleeding, and the like comprising von Willebrand's disease.

Given the physiological importanc of the blood clotting factors, efforts are currently being undertaken by both indlustry and academia to identify new, native proteins which may be involved in the coagulation process. We herein desce the identification of a novel MIl-length polypeptrde which possesses homology to the human matuilin-2 precursor polypeptide.

7- PRO246 The cell surae protein HCAR is a memnbrane-bound protein that acts as a receptor for subgroup C of the adenoviruses and subgroup B of the coxsackievinuses. Thius, HCAR may provide a means for mediating viral infection of cells in that the presence of the HCAR receptor on the cellular surface provides a binding site for viral particles, thereby facilitating viral infection.

In light of die physiological importance of membrane-bound proteins and specficially those which serve a cell surface receptor for viruses, efforts are currently being underaken by both industry and academia to identify new, native membrane-bourd receptor proteins. Many of these efforts are focused an the screning of mammalian recombian DNA libraries to identify die coding sequences for novel receptor proteins. We herein describe a novel mem rnemunl polypcpdde (designated herein as PROM4) having homology to the cell surface protein HCAR and to vanous anor antigens itichning A'33 and caxeiremhryonic antigen, wherein this polypepdde may be a novel cell surfce vinui receptor or timior antigen.

S. PRQ228 There are a number of knoiwn seven trnssmtubrar proeais and within this family is a group which includes CD97 and EMRI. CD97 is a seveni-spani transmnobnrane receptor which has a cellular ligand, CDS5. DAR.

Hamann, et al., Mc4. 184C3):1189 (1996). Additionally, CD97 has been reported as being a dediffeaunton marker in humn thyroid carcinomas and! as associated with inflammation. Aust, et al., CeLe~s.

57(9):1798 (1997); Gray. et al., J. Inimunol. 157(12):5438 (1996). CD97 has also been reported as ~being related to the sectmn receptor superfamily, but unlik known members of that family, CD97 and EMR1 have extended extr-Acellular regions that possess several EGF domains at the N-teximzs. Hamann. cc al., Gemmicnh.

32(l):144 (1996); Harrnann. et al..,J Llzmnal.. 155(4):1942 (1995). EMR1 is further described in Lin. et al., Ggimis 41(3):301 (1997) and Baud, ct al.. Gckwmics. 26(2):334 (1995). While CD97 and EMRI appear to be related to the secretim receptors. a known member of the secretin family of G protein-coupled receptors includes the alpalatroxin receptor. latrophilin, which hias been described as calcium independent and abundant among neuronal tissues. Lelianova, et al.. 1. Biol. Cbam.. 272(4). 21504 (1997); Davletov, et al., J. Biolgkm. 271(38):23239 (1996). Both members of the secretin receptor superfamily and non-members which are related to the secretin receptor superfamily, or CRF and calcitonin receptors are of interest. In particular, new members of these families, identified by their homology to known proteins, are of interest.

Efforts are being undertaken by both industry and academia to identify new membrane-bound receptor :proteins, particularly trammembrane proteins with EGF repeats and large N-terminuses which may belong to the family of seven-transrnembrane proteins of which CD97 and EMRI are members. We herein describe the identification and charactization of novel polypeptides having homology to CD97 and EMRI. designated herein as PR0228 polypeptides.

P.RS33 Growth factors are molecular signals or mediators that enhance cell growth or proliferation, alone or in conert by binding to specific cell surface receptors. however, there are other cellular reactions than only growth upon expression to growth factors. As a result, growth factiors are better characterized as multfunctional and potent cellular regulators. Their biological effects include proliferation, chemootaxis and stimulation of extracellular matrix production. Growth factors can have both stirnilatory, and inhibitory effects. For example, transforming growth factors (rGF-3) is highly plertropic and can mbnuilate proliferation in some ceils especially connective tissues, whilc being a potent inhibitor of proliferation in others, such as lymphocytes and epithelial cells.

The physiological effect of growth stimulation or inhibition by growth factors depends upon the state of development and differentiation of the target tisue. The mechanism of local cellular regulation by classical endocrine WO "/14328 WO 9914328PCr/US9811 9330 moleculcs comprehends autocrnne (same cell). jwdtacrin (neighbor cell), and paracrne (adjacent cell) pathways.

Peptide growth factors arce lements of a complex biological Ianguage- providing the basis for intercellular coniunucation. They permit cells to convey information between each other, mediae interaction betwecti cells and change gene expression. the effect of these rnultifunctionial and plwipotent factors is dependent on the prescsice or absence of Q&hCr peptides.

Fibroblast growth factors (FGFs) are a family of heparin-binding. potent mitogens for both normal diploid fibroblasts and established cell lines Godpodarowicz., D. ec al. (1984). Proc. Nial. Acad. Sci. USA 8 1: 6983. the FOF family catnprises acidic FOF (FOF- basic FGF (FGF-2). INT-2 (FGF-3). K-FOF/HS (FGF-4). FGI'-S, FGF-6. KGF (FGF-7). AIGF (FGF-8) among others. All FGFs have two cnserved cysteime residues and share 30-500/ sequence hanology at the amino acid level. These factors are mitogenic for a wide variety of normal diploid mesoderm-deiived and neural crest-derived cells, inducing granulosa cells, adrenal cortical cells. chrondocytes, myoblasts, corneal and vascular eadothelial cells (bovine or human). vascular smooth muscle cells. lens, retina and prostatic epithelial cells, :00 e oligodendrocytes, astrocytes, chrondocytes, myoblasts and osteoblasts.

Fibroblast growth factors cani also stimulate a large number of cell types in a non-niitogenic wanner. These' aclvities include promotion ofcll ngration into a wound area (chemnotaxis), initiation of new blood vessel formulation (argiogees), modulation of ncrve regeneration and survival (netrtrophisin), modulation of endocrine functions, and stimulation or suppression of specific cellular protein expression, octracellular matrix production and cell survival.' A. Bohlen, Handbook of~xp. PhrmacoL 91(l1): 369-418 (1990). These properties provide a basis for using fibroblast growth factors in theapeutic approaches to acclerate wound healing, nerve repair, collateral blood vessel formation, and thc like. For example, fibroblast growth factors, have been suggested to minimize myocardium .20 damage in heart disease and surgery 4,378,437).

We herein describe the identification and characterization of novel polypeptides having homology to FGF, herein designated PR0533 polypeptides.

PR

Some of the most important proteins involved in the above described regulation and modulation of cellular processes ame the enzymes which regulate levels of protein phosphorylation in the cell. For example, it is known that goo":dte transduction of signals that regulate cell growth and diffcrentiaton is regulated a least in pan by phosphorylation dephosphorylation of various cellular proteins. The enzymes that catalyze these processes include the protein kinases. which function to phosphorylate various cellular proteins, and the protein phosphatases, which function to remove phosphate residues from various cellular proteins. The balance of the level of protein phosphorylation in the cell is thus mediated by the relative activities of these two types of enzymes.

Although many protemin kinsenzymes have been identified, the physiological role played by many of these catalytic proteins has yet to be elucidated. It is well knwn, however, that a mnber of the known protein kinases fiincion to phosphorylate tyrosinie residues in proteins, thereby leading t o a variety of different effects. Perhaps most importantly, there has been a great deal of interest in the protein tyrosine kina=seaince the discovery that many oncogcne products and growth factors possess intrinsic protein tyrosine kinase activity. There is, therefore, a desire to identify new members of the protein tyrosine kinase family.

Given the Phys1ologcal importance of the protin kinases, efforts are being undertaken by both industry and academia to identify new. native kinuec proteins. Many of these efforts arm focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel kinase proteins. We herein describe the identification and characterization of novel polypeptides having homology to tyrosine kinase protins, designated herein as P*R0245 polypcptides.

IL. PROZZO. PR0221 and PRO227 Protein-protein interactions include receptor and antigen complexes and signaling mechanisms. As more is knwn about the structual and functional tnechanisms underlying prote-in-protein interactions, protein-protein interactions can be mome easily manipulated to regulate the particular result of the proteinl-protein interaCtion. Thus, the underlying mechanisms of protein-protein interactions ame of intereu to dhe scientific and medical community.

All proteins containing leucine-ridi repeats are thought to be involved in proteinw-proei interactions.

Leucine..rlch repeats are short sequece motifs present in a imiber of proteins with diverse functions and cellular locations. The crystal structure of uibonuclease inhibitor protein has revealed that leucine-rich repeats correspond to beta-alpha structural units. These units are arranged so that they form a parallel beta-sheet with one surface exposed to solvent, so that the protein acquires an unusual, nonglubular shape. These two features have been indicated as responsible for the protein-binding functions of proteins containing leucine-rich repeats. See, Kobe and Deisenhofer, Trends Riochem. Sci,. 19(10):415-421 (Oct. 1994).

A study has been reported on leucine-rich proteoglycans which serve as tissue organizers, orienting and :ordering collagen fibriLs during ontogeny and are involved in pathological processes such as wound healing, tissue repair, andtUMrlor troma formation. lo~zo. R. Crit. Rev. Riochem. Mot, Biol., 32(2):141-174 (1997). Others studies implicating leucine rich proteins in wound healing and tissue repair are De IA Salle, C. et aL. Vouv. Rev- Fi eratl. (Germany), 37(4):215-22 (1995), reporting mutations in the leucine rich motif in a complex associated with the bleeding disorder Bcrnard-Soulier syndrome and Cblenietson, K. Thom. Hemos. (Germany), 74(l):111-116 (July 1995), reporting that platelets have leucine rich repeats. Anthe protein of particular interest which has been reported to have leucine-rich repeats is dhe SUIT protein which has been reported to be useful in treating neuro-degenerative diseases such as AzIeimer's disease. nerve damage such as in Parkinson's disease, and for diagnosis of cancer, see, Artavanistsakonas, S. and Rotibcrg. J. W09210518-Al by Yale University. Other stuie reporting on the biological functions of proteins having leucine-rich repeats include: Tayar. et al., MtiL CeJl n4djizin1.. (Ireland), 125(1-2):65-70 (Dec. 1996) (gonadotropin receptor involvement); Miura, et al..

NtC2n insho (Japan), 54(7):1784-1789 (July 1996) (apoptosis involvement); Harris, P. et al., 1.Am. Soc.

ftbiul., 6(4):1125-1 133 (Oct. 1995) (kidney disease involvement)-. and Ruoslahji E. L. et al., W09110727-A by la Jolla Caree Research Foundation (decorin binding to transforming growth factorfl involvement for treatment for cancer, wound healing and scarring).

Efforts are therefore being undertakien by both industry and academia to identify new proteins having leucine rich repeats to better understand protein-protein interactions. Of particular interest arc those protein having leucine: rich repeats and homology to Iniown proteins having leucine rich repeats such as the SLIT protein and platelet glyroproteus V.

12. EBDZM8 Immutnoglobulins are artibiody molecules, the proteins that function both as receptors (or antigen on the 13cell membrane and as the secreted products of the plasma cell. Like all antibody molecules. inimunoglobulins perform two major functions: they bind specifically to an antigen and they participate in a limited mnmber of biological effector fiimtions. Therefore, new members of the Ig superfamily are always of interest. Molecules which act as receptors by various vinuses and those which act to regulate imnrme (unction are of particular interest. Also of particular interest are those molecules which have homology to known Ig family members which act as virus receptors or regulate inmmune function. Thus, molecules having homology to poliovirus receptors, CRTAM and CD166 (a ligand for lymphocyte antigen CD6) are of particular bntrest.

Extracellular and Iemrane-bound proteins play important roles in tie formatioa. differentiation and maintenance: of niulticelhilar organisms. The fate of many individual cells, proliferation, migration, differentiation, or interaction with other cells, is typically governed by information received from other cells and/or the immediat envirnment. This information is often transmitted by secreted pobypptdc (for ins tance. mitogenic factors, survival factors, cytotoxic: factors, differentiation factors, neuropeptides, and hormones) which are, in turn, received and interpreted by diverse cell receptors or membrane-bound proteins. These secreted polypeptides or signaling molecules normally pass through the celllar secretory pathway to reach their site of action in the extraccllular environment, usually at a membrane.bound receptor protein.

We herein describe the idetiaion and! chaacrization of novel polypeptides having homology to CRTAM, designated herein as PRO2S8 polypeptides.

13. PRO~Z Protein-protein interactions include receptor and antigen complexes and signaling mechanisms. As more is known abouit the smricuiral and functional mnechanisms underlying protein-protein interactions, protein-protein interactions can be more easily manipulated to regulate dhe particular result of the protein-protein interaction. Thus, the underlying mechanisms of protein-protein interactions are of interest to the scientific and medical community.

All proteins containing leucine-rich repeats are thought -to be involved in protein-protein interactions.

Leucine-rich repeats are short sequence motifs present in a nmber of proteins with diverse functions and cellular locations. The crystal structre of ribonuclcase inhibitor protein has revealed that leucine-rich repeats correspond to beta-alpha stnzcauaj units. These units are arranged so that they form a parallel beta-sheet with one surface exposed to solvent, so that the protein acquires an unusual, nonglobular shape. These two features have been indicated as responsble for the protein-binding functions of proteins containing leucinenrch repeats. See, Kobe and Deisenhofr, Trends iochem- Sc., 19(10):415-421 (Oct. 1994).

A study has been reported on leucine-ridi proteoglycans which serve as tissue organizers, orienting and ordering collagen fibrils during ontogeny and are involved in pathological processes such as wound healing, tissue repair, and tumor strana formnation. lorzo, R. Grit. Rev. icem -Mol 32(2): 141-174 (1997). Others studs implicating leucine rich proteins in wound healing and tissue repair are Dc LA Salle. et Vouv.e Fr. H=WI. (Germany). 37(4)215-2 (1995). reporting imitations in the leucine rich motif in a complex associated with the bleeding disorder Bernard-Soulier syndrome and Clemetsos, K. 7flmb Hamnu. (Germany), 74(l):!l11-116 (July 1995). reporting that platelets have leucinenrch repeats. Anotherprotein of particular interest which has been reported to have leucine-rich repeats is the SUT protein which has been reported to be useful in treating rtamo-degenerative diseases such as Alzheimier's disease. nerve damage such as in Parkinson's disease, arnd for diagnosis of cancer. we,. Artavanistsakonas. S. and Rothberg, J. W09210518-Al by Yale University. Other saidies repogng on the biological functions of proteins having leucine-rich repeats include: Tayar, et al., finaL CelF~ocio. (Irelandl). 125(1-2):65-70 (Dec. 1996) (gonadolropini receptor involvement); Miura, et al..

NipponRimlza (Japan). 54(7):1784-1789 (July 1996) (apoptosis involvement); H~arris, P. cc al.. L.Am~. NcbM. 6(4):1125-1 133 (Oct. 1995) (kidney disease involvement); and Ruoslahti, E. L. et al., W09110727-A by La Jolla Cane Research Foundation (decorin binding to transforming growth factouil involvement for treatmient for cancer. wound healing and scarring).

Efforts are therfor being undertaken by both industry and academia to identifyr new proteins having leucine rich repeats to better understand protein-protein interactions. neuronal development and adhesin molecules. Of particular interest are dose proteins having leucine rich repeats and homology to Imown proteins having leucine rich repeats such as the SIIF protein. We herein describe novel polypeptides having homology to SUTr, designated herein as PR0266 polypeptides.

14. PRO26 Thrombomodulin binds to and regulates the activity of thrombin. It is important in the control of blood coagulation. Thrombomodulin functions as a naual anticoagulant by accelerating the activation of protein C by *thronmbin. Soluble thrombomodulin may have therapeutic use as an antithrombotic agent with reduced risk for hemorrhage as compared with hepauin. Thrombomodulin is a cell surface trans-membrane glycoprotein, present on aidofrlial cells and platelets. A smaller, functionally active form of throxnbomodulin circulates in the plasm and is also fouWx in urine. (in Haeberli, Humant Protein Data. VCH Oub., 199). Peptades having homology to thrombomodulin arc particularly desirable.

We herein describe the identification and characterization of novel polypetides having homology to thrombomodulin. designated herein as PR0269 polypeptides.

PRO28 Procollagen C-proteinase enhancer protein binds to and enhances the activity of bone morphogenic protein 'BMPI */prooolagen C-proteinase (PCP). It plays a role in extracellular matrix deposition. BMP I proteins may be used to induce bone and/or cartilage formation and in wound healing and tisse repair. Therefore, procollagen Cprotesnase -4-no protein. BMP I and proteins having homology thereto, arc of interest to the.-scntific and medical communities.

We herein describe the identification and characterization of novel polypeptides having homology to procollagen C-pro[Cinae =bhan= protein pracwsor and proco~lagen C-proteinase enhancer protein, designated herein as PR0287 polypeptides.

16. PR.Q214 Growth factors art molecular signals or mediators that enhance& cell growth or proliferntion, &lone or in concert. by binding to specific CClU surface receptors. However, there are other cellular reactions than Only growth upon explression to growth fators. As a result, growth factors are better characterized as muitifur~tional and potent Cellular regulators. Their biological effects include proliferation, chemotaxis and stimulation of extraccllular mnatrix production. Growth factors can have both stinmulatory and inhibitory effects. For example, transforming growth factor P is highly plekotpic and can stim-late proliferation in some cells, especially connective tissue. while being a potent inhibitor of proliferation in others, such as lymophocytes and epithelial cells.

The physiological effect of growth stirmulation or inhibition by growth factors depends upon the state of development and diferanidation of the target tissue. fT mechanism of local cellular regulation by classical endocrine molecules involves comprehends autocrme (sam cell), juarine (neighbor cell), and paracrine (adjacent cells) pathways. Peptid growth factors ame elements of a complex biological language. providing the basis for intercellular comnication. They permit cells to convey information between each other, mediate interaction between cells and chnge gene expression. The effect of these uudhmcdoua and pluripotemi fatr is dependent on th presence or absence of other peptides.

Eipidermal growth factor (EGP) is a conventional mitogenic factor that stimulates the proliferation of various types of cells including epithelial cells and fibroblasts. EOF binds to and activates the EGF receptor (EGFR), which initiates intracllular signaling and subsequent effects. The EGFR is expressed in neurons of the cerebral cortex, cerebeihan, and bippocanipus in addition to other regions of the central nervous system (CNS). In addition, EGF is also expressed in various regions of the CNS. Therefore, EUF acts not only on mitotic cells, but also on postinitotic nmuron. In fact. many studies have indicated that EGF has neurotrophic or neurmodulatory effects on various types of neurons in the CNS. For example, EGF acts directly on cultured cerebral cortical and cerebellar neurons, enhancing nmuite outgrowth and survival. On the other hand, EGF also acts on otr cell types, including septa] inewrgi and snesencephalic dopaminergic neurons, indirectly through glial cells. Evidence of the effects of EGF on neurons in the CNS is accuraulating, but the mechanisms; of action remain essentially unbwwn. EGF-inctuced si=gnagin mitotir-cells is beteruonrtood thanin posnitotic neurons. Studies of coned pbeochromocytom PC 12 cells and culatd cerebral cortical neurons have suggested that the EGF-induced neurotrophic actions axe mediated by sustained activation of the EGFR and mitogen-activated protein kinase (MAPK) in response to EGF. The sustained iniacllular signaling correlates with the decreased rate of EGFR down-egulation, which miighit determine the response of neuronal cells to -EGF. It is likely that EGF is a multi-potent growth factor that acts upon various types of cells including mitotic cells and postiiotic neurons.

EGF is produced by the salivary and Bnurnnes glands of the gastrointestinal system, kidney, pancreas.

thyroid gland, pituitary gland. and the nervous system, and is found in body fluids such as saliva, blood.

cerbrospinal fluid (CSF). urine, aniotic fluid, prostatic: fluid, pancreatic juice, and breast milk Plata-Salaman, CR Peptdesj12: 653-663 (1991).

EGF is -mdaed by its metraz specific receptor, which contains an intrinsic tyrosine kinase. Stoscheck CM et aL, J. Cell Biodem. 135-152 (1986). EGF is believed to function by binding to the extracelular portion of its receptor which induces a transtuembrane signal that activates the intrinsic tyrosine kinase.

Plurification and sequence analysis of the EGF-like domain has revealed the presne~ of six conserved cysteine residues which cross-bind to create three peptdde loops. Savage CR ar at.. I. Mot. Chem. M: 7669-7672 (1979). It is now generally known dnat several other peptides can react with the EOF receptor whiich share the sanme generalized motif XCXCX.,CX,,CXCXGXCX 5 whtere X represents any non-cysteine ammacid. and n is a variable repeat nmber. Non isolated pcptides having this mocif include TGF-a, amaphiregulin. schwannorna-derived growth factor (SDGF). bcparin-bizxlitg EGF-like growth factors and certain vitaly ecoded peptdecs Vacclnia vinis. Itzisner AM. Nacure 313: 8014903 (1985). Shope fibroma virus, Chang et al., Mat CCUi Biol. 7: 535-54 (1987), Molluscuin conagiosum. Porter CD Archard J. Gen. Vito!. 673-692 (1987), anid Myxoma virus.

Uipton C at al.. J. Vio!. 1271-1275 (1987). Puigent SA Lemoine Prog. Growfh Factor Res. 4: 1-24 (1992).

EGF-lik domairts ame not conlimed to growth (actors; but have been observed in a variety of cell-surface and extrcllar proteins which have interestinig properties in ccli adhesion, protein-protein interaction and development, Laurence DIR Gusterson BA. Tumor Biol. U: 229-261 (1990). These proteins include blood coagulation factors (facors VI. IX. X. XlU. protein C. protein S. protein 7, tissue plasminogen activator, urokinase), extracelular matrix components (laminin. cytotactin, entactin). cell surface receptors (LDL receptor, thrombomodulin receptor) and immurilnity-related proteins (complement CMr. uroinodulin).

Even more interesting, the general structure pattern of EGF-like precursors is preserved through lower organisms as well as in mammalian cells. A number of genes with developmental significance have been identified in invertebrates with EGF-Iike repeats. For example, the notch gene of Drosophila encodes 36 tandemly arranged 20 40 amino acid repeals which show homology to EGF, Wharton W et al, Cell 11: 557-581 (1985). Hydropathy plots indicate a putative membrane spanning domain, with the EGF-related sequence being located on the extracelhilar side of dhe membrane. Other homeotic genes with EGF-Iike repeats include Delta. 95F and SZD which were identified using probes based on Notch. aWt the nemiatode gene Lin-12 which encodes a putative receptor for a developmental signal transmitted between two specified cells.

Specifically, BOF has been shown to have potential in the preservation and maintenance of gastrointestinal mucosa. and the repair of acute and chronic ncosal lesions, Konturek, PC et Eur. J. Gastroemterol Hepatol.

2 933-37 (1995). including the treatmnt of necrotizing enterocolitis, Zollinger-Ellison syndrome.

gastroitestinl ulcerationi gastroitestinal ulcerations anid congenital nicrovllus atropy. A. Gughenaa PB Sullivan, Eur. Gastroenterol Hepatol. 10). 945-50 (1995). Additially, EGF has been implicated in hair follicle difterentiation; C.L. dum Cms. J. Invest. Dennol (I Suppl.), 106S-1 13S (1993), SG Hillier, Clin. Endocrinol.

33(4),.427-28 (1990); kIdney function, L.L. Hamm at al.. Semin. Nephrol. 113(1): 109-15 (1993), RC Harris. Am.

J. Miey Dis. 12(6): 627-30 (1991); tear fluid, GB van Setten ei al., Int. Ophthalmol 15(6); 359-62 (1991): vitamin K mediated blood coagulation, 1. Stenflo ar al., Blood 2j(7): 1637-51 (1991). EGF is also implicated various sin disease characterized by abnormal keratinocyte differentiation psoriasis, eithelial cancers such as sqilancos cell caxcinomas of the lung, epidamoid carcinoma of dhe vulva and gliomas. King. LE ai al.. Am. J. Merl. Sci. 226: 154-158 (1988).

Of great interest is mounting evidence that genetic alterations in growth factors signaling pathways are closely linked so developmental abnormalities and to chronic diseases including cancer. Aaronson SA, Science 1146-1153 (1991). For example, c-crb-2 (also known as HER.2). a proto-oWncm with close structural similarity to EGF receptor protein. is overex~ptessed in human breast cancer. King ef al, Science 222: 974-976 (1905); Gullick, WI, Hormones and their actions. Cooke BA er al.. eds. Amsterdam. ELsevier, pp 349-360 (1986).

17. PRO317J.

MWe TGF"f suPergesie family, Or simply TGELP superfamily, a group of secreted proteins, includes a large number of related growth and differentiation factors expressed in virtly all phyla. Superfamily members bind to specific cell surface receptors that activate signal transduction mechanisms to elicit their multifunctional cytokim effects. Koxtejczyk and Hall, Biochem- Cell -Bil. 7A- 299-314 (1996); Attisan and Wran.ya, ineGMlb Factor 7: 327-339 (1996). and Hill. Cellular Sialing. 8: 533-544 (1996).

Members of this family inichzl five distinct fortns of TGFP (Sporn and Roberts. in Peprde Growth Factors and lIbeir Recentors, Sporn and Roberts, e&s. (Springer-Verlag: Berlin, 1990) pp. 419-472). as well -as the ~differentiation factors vglI (Weeks and Meltn Q&~l U: 861-867 (1987)) and DPP-C polypeptide (Padgett ei al., :Nature, W: 81-84 (1987)), the hormnes activmn and inliibin (Mason et Nanirr. Ml: 659-663 (1985); Mason et Grwth Factr, 77-88 (1987)). the Mullerian-inhibiting substance (MIS) (Cate et aL. C£niI. -45: 685-698 (1986)). the bone morphogeectic, proteins (BM[Ps) (Womcy el al., Scec 2A: 1528-1534 (1988); PCT WO 88100205 published January 14. 1988; U.S. 4,877864 issued October 31. 1989). the developmentally regulated proteins Vgr-I (Lyons et al., Proc- Nati_ Acad. Sci USA. 4554-4558 (1989)) and Vgr-2 (Jones et al., Moinc.

Endocrinol. 1961-1968 (1992)). the mouse growth differentiation factor (GDF), such as GDF-3 and GDF-9 (Kingsley, Gee e- 1: 133-146 (1994); McPherron and Lce.I HiolL Chem .2.-21 3444-3449 (1993)), the mouse ~20 lefty/Stral (Meno et al., Nature, In1: 151-155 (1996); Bouillet et Bil. M: 420-433 (1995)). gtial cell lime-denived ncurotrophic factor (GDNF) (Lin er al., Scim. 2W: 1130-1132 (1993), neurturin (Kotzbauer er al., Nature, JI: 467-470 (1996)), and endotiutial bleeding-associated factor (EBAF) (Kothapalli ei al.. J. Cin. Inest S22: 2342-2350 (1997)). The subset BMP-2A and BMP-2B is approximately 75% homologous in sequence to DPP-C and may represent the mammnalian equivalent of that protein.

The proteins of the TGF-P superfamily are disulfide-linked homno- or heterodiniers encoded by larger precursor polypeptide chains containing a hydrophobic signal sequence, a long and relatively poorly conserved Nterminal pro region of several hundred amino acids, a cleavage site (usually polybasic), and a shorter and more highly conserved C-terminal region. This C-terminal region corresponds to the processed mature protein and contains approximatly 100 amino acids wni a chaacteristic cysteine motif, ie., the conservation of seven of the nine cystelne tesidues of TGF-3 amng all known family members. Although the position of the cleavage site between the mature and pro regions varies among the family members, the C-tertimz of all or the proteins is in the identical position.

ending in the sequence Cys-X-Cys-X, but differing in every case from the TGF-3 consensus C-terminus of Cys-Lys- Cys-Ser. Sporn and Roberts, 1990, supra.

There are at least five forms of TOGF- aurra*l identfied, TGF-PI1. TGF-P2. TGF-33, TGF-P4. and TGF- P5S. The activated form of TGF-flI is a lxxnodimer formed by dimerization of the caxboxy-terniinal 112 amino acids of a 390 amino acid precuror. Recombinant TGF-P I has been cloned (Dcrynck etal.. Niature, M:701-705 (1985)) and expressed in Chinese hamster ovary cellu (Gentry at al.. MoIgl.l. iol 7: 3418-3427 (1987)). Additionally, recombinant human TGF.P2 (deMartin er EMBO 9: 3673 (1987)). as well as human and porcine TGF-P3 (Deryuck ar aL. EIABOIL. 2: 3737-3743 (19M.) ten Dijim ea aL. Proc. Nall. Acad. Sc. UISA, 15: 4715 (1988)) have been cloned. TGF-02 has a precursor form of 414 amino acids and is also processed to a bomodimer from the carboxy-terminal 112 amino acids that shares approximately 70% homology with the active form of TGF-fl (Marquardt er at.J 4j L Cb.m 2: 12127 (1987). See also EP 200.341; 169,016; 268,561; and 267,463; U.S.

Pat. No. 4,774.322; Cheifctz ar aL, COl. 4A: 409-415 (1987); Jakowlew eta!., Molecular Endocrnt, 2: 747-755 .(1988); Deryock et at..J LRiol. Ote.. 2U1: 4377-4379 (1986); Sharples et at.. DkLA. fi 239-244 (1987) Derynck etd4., Nul AGs es,1: 3188-3189 (1987); Derynck ar al.. Nuci. Acids- Res-, 15: 3187 (1987); Seyedin ci a..

LBiol Chem. ZUI: 5693-5695 (1986); Madisen et al., 2: 1-8 (1988); and Hanks al Prc al cd Sci, 1j: 79-82 (1988).

TGF-P4 and TGF-P5 were cloned from a chicken chorulrocyte cDNA library (Jakowlew at MOC EndorinaL-. 2: 1186-1195 (1988)) and from a frog cocyte cDNA library, respectively.

The pro region of TG"- associates non-covalendly with the mature TGOP dimer (Wakefield era., L.Biol.

ChM 20: 7646-7654 (1988); Wakefield ea., Grwcth Factors, 1: 203-218 (1989)), and the pro regions are found to be necessary for proper folding and secretion of the active mature dimers of both TGF-Il and activin (Gray and mas MI.d 1328-1330 (1990)). The association between the mature and pro regions of TGF-P masks the biological activity of the mature dimer, resulting in formation of an inactive latent form. Latency is not a constant of dte TGF-O superfamily, since the presence of the pro region has no effect on activin or inhibin biological activity.

A unifying feature of the biology of the proteins from the TOFP superfamily is their ability to regulate developmental processes. TGFP has been shown to have numerous regulatory actions on a wide variety of' both normal and neoplastic cells. TOFP is nmltiliznctonal, as it can either stimulate or inhibit cell proliferation, differentiation. and other critical processes in cell function (Sporn and Roberts, stpra).

One member of the TGF-f3 superfiamily, EBAF. is expressed in-endornetriun only in the late secretory phase and during abnormal endomnetrial bleeding. Kothapalli ar al., L.Clin vet- 22: 2342-2350 (1997). Human erulometrum is unique in that it is the only tisse in die body that bleeds at regular intervals. In addition, abnormal endonietrial bleeding is one of the most common manifestations of gynecological diseases, and is a prime indication for hysterectomy. In sifu hybridization showed that th e mRNA of EBAF was expressed in the stronia without any significant tuRNA expression in the endomesrial glands or endothelial cells.

The predicted protein sequence of EBlAF showed a strong homology to the protein encoded by mouse L-fry/sra3 of the TG"p superfamnily. A motif search revealed that the predicted EBAF protein contains moss of the cysteine residues which are conserved among the TGF- -related proteins and which are necssary for the formation of the cysteim kno structure. The EBAF sequence contains an additional cysteine residue, 12 amino acids upstream from the first conserved cyseine residue. The only other family members known to contain an additional cysteirie residue are TGF-os, inhibins. and GDF-3. EBAF. similar to LEFT7Y, GDF-3lVgr2, and GDF-9, lacks the cysteine residue that is known to form the intermolecular disulfide bond. Therefore, EBAF appears to -be an additional member of the TOFP superfamily with an uwpaired cysseine residue that may not exist as a diner. However, hydrophobic contacts between the two monomer subunits may promote dirner formation. Fluorescence in situ hybridization showed that the ebaf gene is located on human chromosome I at band q42. 1.

Additional membr of t TGF-P superfamily, such as those related to BAF, are being searched for by industry and academics. We herein describe the identification and characterization of novel polypeptides having homology to EBAF. designated herein as PR0317 polypeptides.

i8. The widespread occurrne of cancer has prompted the devotion of considerable resources and discovering new treatments of treatment. One particular method involves the creation of tumor or cancer specific monoclonal antibodies (mAbs) which are specific to tumor antigens. Such snAbs, which can distinguish between normal and cancerous cells are useful in the diagnosis, prognosis and treatruent of the disease. Particular antigens arm known to be associated with neoplastic diseases, such as colorectal cancer.

One particular antigen, die A33 antigen is expressed in more than 90% of prinary or mietastatic colon cancers as well as normal colon epithetiurn. Since colon cancer is a widespread disease, early diagnosis and ucatmemt is an importan- medial goal. Digoi and tra of colon canerca be implemented using monoclonal antibodies (mAbs) specific therefore having fluorescent, nuclear magnetic or radioactive tags.

Raoactive genie, toxins andl/or drug taggcd mAhe can be used for treatment in Sm with mininai patient description.

=iAbs can also be used to diagnose during the diagnosis and treatment of colon cancers. For example, when the serun leveb of the A33 antigen are elevated in a patient, a drop of the evels after surgery would indicate the tumor resection was successful. On the other hand, a subsequent risc in serum A33 antigen levels after surgery would indicate that mnete of the original tumor may have fortned or chat new primary tumors may have appeared. Such monoclonal antibodies can be used in lieu of. or in conjunction with surgery and/or other chemotherapies. For example, U.S.P. 4,579,827 and U.S.S.N. 424,991 199.141) are directed to therapeutic administration of monoclonal antibodies, the latter of which relates to the application of anti-A33 niAb.

Many cancers of ecial origin have adenovirus receptors. In fact, adenovirus-derived vectors have been proposed as a means of inserting antisense nucleic acids into tumors 5.518,885j. TIbs, the association of viral receptors with neoplastic tumors is not unexpected.

We herein describe the idenitification and characteriation of novel polypeptides having homology to certain cancer-associated antigens, designated herein as PRO301 polypeptides.

19. PRO224 Cholesterol uptake can have serious implications on one's health. Cholesterol uptake provides cells with most of the cholesterol they require for membrae synthesis. If this uptake is blocked, cholesterol accumnulates in the blood and can contribute to the formation of atherosclerotic plaques in blood vessel walls. Most cholesterol is transported in the blood bound to protein in the form of complexes known as low-density lipoproteins (LDLs). LDUs are endocytosed into cells via LDL receptor proteins. Therefore. LDL receptor proteins, and proteins having homology thereto, are of interest to the scientific and medical communities. Membrane-bound proteins and receptors can play an importan role in die formation differentiation and maintenanc of multiceliular organisms.

7be LDL receptors are an example of membranec-bound proteins which are involved in the synthesis and formation of cell membranes, wherein the health of an individual is affected directly and indirectly by its function. Many rnenbrne-bownd proteins act as receptors such as die LDL receptor. These receptors can husction t0 endocytose substrates or they can function as a receptor for a channel. Other niemrane-bond proteins (unction as signals or antigens.

Mcmbrane-bound proteins and receptor molecules have various inustrial applications. including as pharmaceutcal and diagnostic agents. *The inenran-boutil proteins can also be employed for screening of potential peptide or small molecule regulators of the relevant receptor/ligand interaction. In the case of the LDL receptor, it is desirable to fin olecilks wich enhance endocytosis so as to lower blood cholesterol levels and plaque formation.

It is also desirable to identif molecules whi inhibit ezziocytosis so that these molecules can be avoided or regulated by individuals having high blood cholesterol. Polypeptides; which ame homologous to lipoprotcin receptors but which do not function as lipoprotein receptors ame also of interest in the determination of the (unction of the fragments which show homnology.

The following studies report on previously imown low density lipoprotean receptors and related proteins i~hzingapoipoprotemns: Sawamoura, a al. Nippon Cheiniphar Co, Japan patent application i09098787; Novak. S..

c~t al.,1 Biol. Chm.. 271:CZ)1 173246(1996); fllaas, 1. iroI., 69(11)7244-7 (Nov. 1995); Scott, .nbcrii.

Metak...Dis. 9/Supp. 1 (3-16) (1986); Yamamoto. ct al., f",i 39:27-38 (1984); Rebece, ct al.. Neurbiol Aging 15:5l117 (1994); Novak, et al., J.Biol. Chemitr. 271:11732-11736 (1996); and Sestavel and Fruchart.

CelMol.Biol.. 40(4):461-81 (June 1994). These publications and others published prior to the filing of this application provide further background to peptides alredy known in the art.

Efforts are being undertaken by both industry and academia to identify new, native membrane-bound receptor proteins. particularly thos having homology to lipoprotein receptors. We herein describe the identific-ation and characterization of novel polypeptides having homology to lipoprotein receptors, designated herein as PRO224 polypeptides.

PRO222 Complement is a group of proteins found in the blood that are important in hurnoral immunity and inflammation. Complement proteins are sequenially activated by antigen-antibody coDmplexes or by proteolytic enzymes. When activated, complement proteins; kill bacteria and other microorganisms, affect vascular permeability.

release histamrin and attact witf blood cells. Complement also enhances phagocytosis when bound to target cells.

In order to prevent harm to auologous cells. the complement activation pathway is tightly regulated.

Deficiencies in the regulation of complement activation or in the complement proteins themselves may lead to immune-complex diseases, such as systemic lupus erythematosus, and may result in increased susceptibility to bacterial infection. In all cases, early detection of complement deficiency is desirable so that the patient can begin treatmez Thus, research efforts are airrently directed toward identification of soluble and membrane proteins that regulate complement activation.

Proteins bIown to be imiportant in regulating complement activation in humans include Factor H and Complement receptor type I (CR1). Factor H is a 150 kD soluble serumn protein that interacts with complement protein C3b to accelerate the decy of C3 convertase and acts as a cofactor for Factor [-mediated cleavage of comiplemient protein COb. Complement receptor type I is a 19G-280 kD membrane bound protein found in mast cells and moest blood cells. CR1 intracts with complement proteins 0~b. C0., and iC3b to accelerate dissociation of C3 convertases, acts as a cofactor for Factor I-mediated cleavage of C 3b and C4b. and binds immune complexes and promotes their dissolution and phagocytosis.

Proteins which have homology to compl-i proteins ame of particular interest to the medical and industrial commnities. Often, proteins having homology to each other have similar fuanction. ft is also of interest when proteins having homology do no have similar functions, indicating that certain structural motifs identify information other than function, such as locality of function.

Efforts are being undertaken by both indlustry and academia to identif new, native secreted and membrane-bound proteins, particularly those having homology to known proteins involved in .the complemeitt pathway. Proteins involved in the complement pathway were reviewed in Birmingham DJ (1995), Criicl eiew kiMn gjg 15(2):133-154 and in Abbas AK, ct al. (1994) Cellular and Molecular Immunology, 2nd Ed. W.B Saunders Company, Philadelphia, pp 295-315.

We herein describe the identification and characterization of novel polypeptides having homology to omplement receptors, deuignatcd herein as PR0222 polypeptides.

e 21. PO3 The successful function of many system within mlidcetlular organism is dependent on CCU-CeCU interactions. Such interactions are affected by the alignment of particular ligands; with particular receptors in a manner which allows for ligand-receptor binding and thus a celk-ell adhesion. While protein-protein interactions in cell recognition have been recognized for some time, only recently has the role of carbohydrates in physiologically relevant recognition been widely considered (see B.K. Brandlcy et al., Leuk. Biot. nfl: 97 (1986) and N. Sharon et Sdence 24ti: 227 (1989). Oligosaccbarides are well positioned to act as recognition novel lectins due to their Cell surfce location and structural diversity. Many oligosacchaide structures can be created through the differential activities of a smaller number of glycosyltranferascs. The diverse structures of oligosaccharides can be generated by tanscription of relatively few gene products, which suggests that the oligosaccharides are a plausible mechanism by which is directed a wide range of cell-cell interactions. Examples of differential expression of ccli surface carbohydrates and putative carbohydrate binding proteins (lectins) on interacting cells have been described Dodd T.M. Jessel, J. Neurosc. 3278 (1985). LJ. Regan et Proc. Nail. Acad. Sci. USA Ul: 2248 (1986); M.

Constantene-Paton et Nature VA4: 459 (1986); and M. Tiemeycr et at.. J. Biol. Chem. ZaI: 1671 (1989). One interesting member of the lectin family are selectins.

The migration of leukocytes; to sites of acmt or chronic inflammation involves adhesive interactions between these cells and the endothelitum. This specific adhiesion is die initial event in the cascade that is initiated by inflammatory insults, and it is. therefore, of paramonoit importace to the regulated defense of the organism.

The types of cell adhesion molecules that are involved in the interaction between leukocytes and the erilothelium during an inflammatory response currently stands at four. selecms (Carbohydrate and glycoprotein) ligands for slctiris; integrins; and integrin ligands, which are members of the immunoglobulin gene superfamily.

Mrt selectins arm cell adhesion umolecules that are unified both stuurally and functionally. Structurally, selectins ame eharactered by the inclution of a domain with homology to a calcium-ependent lectin (C-lectins), an epidermal growth factor (egf)-like domain and several complement binding-like domains, Bevilacqua. M.P. et al., Science 2&3: 1160-1165 (1989); Johinston Cell 5: 1033-1044 (1989); Lasky ercl. Celi: 1045-1055 (1989); Siegalman, M. et at.. Science W4: 1165-1172 (1989); Stoolman, LM.. Cell U: 907-910 (1989). Functionally.

seletins share the common property of their ability to mediate ell binding through interactions between their lectin domains and! cell surface carbohydrate ligands (Braricy. B. ei aL. Cell 63. 861-863 (1990); Springer. T. and Lasky, Nature M,2 19-1917 (1991); Bevilacqua, M.P. and Nelson. A. Clin. Invest. 21 379-387 (1993) and Tedder er al, J. Exp. Med. flQ: 123-133 (1989).

7h=c aretre snenimcbers; identified so far in the selectin family of cell adhesion molecules: L-selectin (also alld pripera lyph od hoingrecpto (pHR) LE-CM-!. LAM-I, gp90 1 *EL. gpl00IL. gpl 101. MEL- 14 antigen, Leu-8 antigen. TQ-I antigen, DREG antigen), E-selectmn (LEC-CAM-2, LECAM-2, ELAM-1) and Pselectin (LEC-CANI-3. LECAM-3. GMP-140. PADGEM).

ibe identification of the C-lectin domain Wa led to an intense effort to define carbohydrate binding ligands for proteins containing such domains. E-selectin is believed to recognize the carbohydrate sequence NeuNAca2- 3GalP3-4F=cI -3)GlcNAc (sialyt-Lewis x. or sic') and related oligosaocbarides. Berg eta!.. A. Biol. aem. Z.U: 14869-1497n (199 Lowe eaaL. Cell f3: 475484 (1990); Phillps at al.. Sdence 2M: 1130.1132 (1990); Ti1emeyer et al.. Proc. Natl. Acad. So. USA H: 1138-1142 (1991).

L-seclectin. which comprises a lectin domain, performs its adhesive function by recognizing carbohydrate- 20 containing ligands on endothelial cells. L.-sclcctin is exprecssed on the surface of leukocytes. such as lymphocytes, nemzrqlils, monocytes and cosinophils, and is involved with the trafficking of lymphocytes to peripheral lympboid tissues (Gallauin et aL, Nature 3Q1: 30-34 (1983)) and with acute neutrophil-medicated inflammatory responses (Watson. Nature M42:164-167 (1991)). The amino acid sequence of L-seleetin and die encoding nucleic acid *.:sequence are, for example. disclosed in U.S. patent No. 5,098,833 issued 24 March 1992.

L-selectin (LECAM-1) is particularly interesting; because of its ability to block neutrophil influx (Watson et al., Nature 3A2: 164-167 (1991). It is expressed in chronic lymphocytic leukcemia cells which bind to HEV (Spertini cia.Nature 34: 691-694(1991). It is also believed that HEV Structures at sites of chronic inflammation arc associated with the symptoms of diseases such as rheumatoid arthritis, psoriasis and multiple sclerosis.

E-scetin (El-AM-I). is particularly interesting because of its transient expression on endothelial ceils in response to ILI or ThF. Bevilacqua et al., Science W4: 1160 (1989). The time course of this induced expression C2-8 h) suggests a role for this receptor in initial neutrophil induced extravasnuori in response to infection and injury.

It has further been reported that and-ELAM- I antibody blocks ie influx of neutrophils in a primate asthma model and th= is beneficial for preventing airway obstruction resulting from the inflammatory response. Gundel ei al., J.

Glin. Invest. f: 1407 (1991).

The adhesion of circulating neutrophils to stimulated vascular endothelium is a primary event of the inflamnitooy response. P-selectin has been reported to recognize die Lewis x structure (Galp1-4(Fucal-3) G~cNAc), Larsen doal.. Cell 467-474(1990). Others report that an additional terminal linked sialic acid is required for high alfmity binding. Moore et al., 1. Cell. Not. 12: 491-499 (1991). P-selectin has been shown to be significant in acute lung &#jwy. Aii-P-selertin aztiody has been shown to have strong protective effects in a rodent lung injury model.

M.S. Mulligan et al., I. Clin. Invesit. 29: 1600 (1991).

We herein describe the identification and characterization of novel polypeptides having homology to lectin proteins. herein designated as PR0234 polypeptides.

22. PR23.1 It Sonme of the mocst importan proteins involved in the above described regulation and modulation of cellular processes ame the enzymes which regulate levels of protein phosphorylation in the cell. For example, it is known that the tansduction of signals that regulate cell growth and differentiation is regulated at least in part by pliosphorylation and dephosphorylation of various cellular proteins. The enzymes that catalyze these processes include the protein ldmes which function to phosphorylate various cellular proteins, and the protein phosphatases, which fuinction to remove phocsphate residues firomn various cellular proteins. The balance of the level of protein phosphorylation in the cell is thus mediated by the relative activities of these two types of enzymes.

Protein phosphatases represent a growing family of enzymes that are found in many diverse forms, including both membrane-bound and soluble forms. While many protein phosphatases have been described, the fimedtons of 15 onl a very few are beginning to be understood (ranks. Semrn. Cell Biol. 4:373-453 (1993) and Dixon, Recent Prog.

Honn. Res. 51.405414 (1996)). However, in general, it appears (hat many of the protein phosphatases function to modulate the positive or negative signals induced by various protein kinases. Therefore, it is likely that protein phosphatases play critical roles in numerous and diverse cellular processes.

Given the physiological importance of the protein phosphatases, efforts are being undertaken by both industry and academia to identify new, native phosphatase proteins. Many of these efforts are focused on the screening of mammalia recomnbinant DNA libraries to identify the coding sequences for novel phosphatase proteins.

Examples of screening methods and techniques arc described in the literature (see, for example, Klein et mr .:Natl, Acad. Sci., 91:7109-7113 (1996); U.S. Patent No. 5.536,637)).

We herein describe the identification and characterization of novel polypeptides having homology to acid pho3phatases, designated herein as PR0231 polypeptides.

5*S23. PR-2 Scavenger receptors are known to protect lgG mnolecules from catabolic degradation. Riechmann and Hlnger, NtrBiecnlg, 15:617 (1997). In particular, studies of the CH2 and CH3 domains have shown that specific sequences of these domains are important in determining the half-lives of antibodies: Ellerson. et al., L.hImtml.. 116: 510 (1976). Yasmeen, et al., L ulmnl. 116: 518 (1976; Pollock. et al., Er- LJmmuna 2021 (1990). Scavenger receptor proteins and antibodies thereto are further reported in U.S. patent No. 5,510,466 to Krieger. et al. Due to the ability of scavenger receptors to increase the half-life of polypeptides and their involvement in immune function, molecules having homology to scavenger receptors are of importance to the scienitific and medical commutnity.

Efforts wre being undlertaken by both industry and academia to identify new, native secreted and membranebound receptor proteins, particularly those having homology to scavenger receptors. Many efforts are focused on the screening of mammalian recombinant DNA libraries to identify the coding sequemme for novel secreted and membrane-bound receptor proteins. Examples of screening methods and techniques are described in the literature [sec. for examnple, Klein et ad.. Proc. Natt Acad. Sci., 21:710&-7113 (1996); U.S. Patent No. 5,536,637)).

We herein describe the identification and characterization of novel polypeptides having homology to scavenger receptors. designated herein as PR0229 polypeptides.

s 24. P~RO238 Oxygen free radicals and antioxidants appear toplay art importarst role in the centra] nervous system after cerebral ischaznia and reperfusian. Moreover, cardiac injuty, related to ischaemnia and reperfusion has been reported to be caused by dhe action of free radicals. Additionally. studies have reported that the redox state of the ccll is a pivotal determinant of the fate of die cells. Furthermore, reactive oxygen spedies have been reported to be cytotoxic, causing inflammatory disease, including tissue necrosis, organ. failure, atherosclerosis, infertility, birth defects, prcatnrc aging, utatians and malignancy. Thus, the control of oxidation and reduction is important for a nrumber See .of reasons including for control and prevention of strokes, heart attacks, oxidative stress and hypertension. In this regard, reductases, and particularly, oxidoreductases, are of interest. Publications futrther describing this subject matte r Incle Kelsey. et al., Br a 76Q):852-4 (1997); Friedrich and Weiss, J.her B~2iol., 197(4):529-40 (1997) and Piculle, et al.. J.Bactedol. 179(18):56U4-92 (1997).

Efforts are being undertaken by both industry and academia to identify new, native secreted and membranebound receptor proteins, Particularly secreted proteins which have homology to reductase. Many efforts are focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel secreted and membrane-bound receptor proteins. Examples of screening methods and techniques are described in the literature too@ [sec. for example, Klein e( al.. Proc. NaI. Acad. Sci., 22:7108-7113 (1996); U.S. Patent No. 5,536,637)].

We herein describe the identification arnd characterization of novel polypeptides having homology to 0* reductase, designated herein as PR0238 polypeptides.

25. rRQ233 Studies have reported that the redox. state of the ccli is an important determinant of the fate of the cell.

Furthermore, reactive oxygen species have been reported to be cytotoxic. causing inflammatory disease, including tissue necrosis, organ failre, atherosclerosis. infertility, birth defects, premature aging, mnutations and malignancy.

Thius, the control of oxidation and reduction is importa= for a number of reasons, including the control and prevention of strokes, heart attacks, oxidative stress and hypertension. Oxygen free radicals and antioxidants appear to pa an important role in the central nervous system after cerebral ischernia and reperfusion. Moreover, cardiac injury, related to ischaernia an4 reperfusion has been reported to be caused by the action of free radicals. I this regard, reductases, and particularly, oxidoreductases are of interest. In addition, the trascription factors, NF-kappa B and AP-l1. arc known to be regulated by redox state and to affect the expression of a large variety of genes thought to be involved in the pathogenesis of AIDS, cancer, atherosclerosis and diabetic comiplications. Publications further describing this subject matter include Kelsey, et al.. Br. L.C 76(7):852-4 (1997); Friedrich and Weiss, jL 7beor- i 187(4):529-40 (1997) and Picuil, et al., J.Bactriol., 179(18):5684-92 (19917). Given the physiological urnportamc of redox reactions in vivo. efforts arm currently being wider taken to identify new, native proteins which ame involved in redox reactions. We describe herein the identification of novel polypeptides which have homology to reductase. designated herein as PR0233 polypeptides.

26, PR23 The carboxypeptidase family of exopeptidases constitutes a diverse group of enzymes that hydrolyze carboxyl-tersoinal amide bonds in polypeptides, wherein a large mnber of mammalian tissues produce these enzymes. Many of the carboxypeptidase enzymes that have been identified to date exhibit rather strong cleavage specificities for certain amino acids in polypeptdecs. For example. caulioxypepudase enzymes have been identified which prefer lysine, arginine, serine or amino acids with either aromatic or branched aliphatic side chains as substrates at the carboxyl icruiin of the polypeptide.

With regard to the scrinc carboxypeptidases, such amino acid specific enzymes have been identified from a variety of diffautt -Inian and non-mammalian organisms. The maimmalian senine carboxypeptidase enzymes play important roles in many different biological processes including, for examle, protein digestion, activation, inactivation. or modulation of peptide hormone activity, and alteration of the physical properties of proteins and enzymes.

In light of the physiological irnportance of the seizie caftoxypeptidases, efforts are being undertaken by both industry and acadania to identify new, native secreted and memnbrane-bound receptor proteins and specifically novel carboxypeptidases. Many of these efforts are focused on the screening of mammixalian recombinant DNA libraries to identify the codi sequtaxncs for novel secreted and memnbrane-bound receptor proteins. We describe herein novel 20 polypeptides having homology to one or more serine carboxypeptidase polypeptides, designated herein as PR0223 polypeptides.

*27. PRO235 Plexin was first identifid in Xenopus tadpole nervous systemi as a membrane glycoprotein which was shown to mediate cell adhesion via a homophilic binding mechanism in the presence of calcium ions. Strong evolutionary conservation between Xenopus. mouse and human homologs of plexin has been observed. [Kaneyama et al., Biocliem. And Biophys. Res. Commn. 226: 524-529 (1996)]. Given the physiological importanc of cell adheion mechanisms in Wvo. efforts are currently being under taken to identify new, native proteins which are involved in cell adheion. We describe herein the identification of a novel polypeptide which has homnology to plexin. designated herein as PR0235.

28. PRO236 and PR262 P-galactosidasc is a well known enzymatic protein which functions to hydrolyze jP-galactoside molecules.

P-galactosidase has been employed for a variety of different applications. both in vitro and in vivo and has proven to be an extremely useful research tool. As such. there is an interest in obtaining novel polypeptides which exhibit homology to the P-galactosidasc polypeptide.

Given the wtong interest in obtaining novel potypeptides having homology to P-galactosidase, efforts arc currently being undertaken by both industry and academia to identify new. native k-a~ar-tosidaze homolog proteins.

Many of these efforts are focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel P-gahactosidase-like proteins. Examples of screening methods and techniques arm described in the Iiteracurc (see, for example. Klein et al., Proc. Nad. Ara.u Sci_ 2:710&-7113 (1996); U.S. Patent No.

5,536.637)]. We herein describe novel poylpeptides having siginificant homology to the jl-galabtosidase enzyme, designated herein as PR0236 and PR0262 polypeptides.

29. PROD2 Densin is a glycoprotein which has been isolated from the brain which has all the hallmarks of an adhesion molecule. It is highl concentrated at synaptc sites in die brain and is expressed prominently in dendritic processes in developing nettrons. Densin has been characterized as a member of the 0-linked sialogiycoproteins. Densin has relevance to medically iipesunt processes such as regeneration. Given the physiological importance of synaptic processes and cell adhesion mechanisms in vivo, efforts are currently being under taken to identify new, native proteins which are involved in synaptic machinery and cell adhesion. We describe herein the identification of novel 15 polypeptides which have homology to dewnm designated herein as PR0239 polypeptidecs.

PRO257 Ebrierin Is a cell surface protein associated with von Ebner glands in mammals. Efforts arc being undertaken by both industry and academia to identify new, native cell surface receptor proteins and specifically those which 20 possess sequence homology to cell surface proteins such as cbnerin. Many of these efforts are focused on the screening ofmammualian recombinant DAlibraries to identify tecoding sequences frnovel receptor proteins.

We herein describe the identification of novel polypeptides having significant homology to the von Ebner's gland- :associated protein ebnerin, designatcd herein as PR0257 polypeptudes.

31. PRO260 Fucosidases are enzymes that remove fucose residues from facose containing proteoglycans. In some pathlogical conditions, such as cancer. rheumatoid arhritis, and diabetes, there is an abnormal fucosylation of serum proteins. Thierefore, fucosidases. and proteins having homology to fucosidase, are of importance to the study and abrogation of these conditions. In particular, proteins having homology to dhe alpba-l-fucosidase precursor are of interest. Fucosjdases and fucosidase inhibitors are further described in U.S. Patent Nos. 5.637,490. 5.382.709, 5.240.707, 5,153,325, 5,100,797, 5,096.909 and 5,017,704. Studies arc also reported in Valk. et al.. L. imrl..

71(9):6796 (1997), Aktogu. et al.. Monaldi. Arch Cest Dis. (Italy). 52(2)-.118 (1997) and Focarelli, et al., Ripchem.

Bionhvs. Rcs. Commun. 234(1)54 (1997).

Efforts are being undertaken by both indlustry and academia to identify new, native secreted and membranebound receptor proteins. Of particular interest are proteins having homology to dhe alpha-l-fucosidase precursor.

Many efforts; are focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel secreted and memnbrane-bound receptor proteins. Examples of screening methods and techniques are describd in dieterature (see. for example. Klein et Proc Nail. Acad, Sc., 2:7108-7113 (1996); U.S. Patent No. 5.536.637)].

We herein describe the identification and characterization of novel polypeptidcs having honiology to fucosidases. designated herein as PR0260 polypeptides.

32. PR.Q263 CD44 is a ccli surface adhesion molecule involved in cell-ell and cell-matrix interactions. Hyaluronic acid.

a component of the extracelfular matrix is a major ligand. Other ligands include collagen. fibronectin, laminin.

chrondroitin sulfate. mucosal addrcssin. serglycin anid osteoponin. CD44 is also important in regulating cell traffic, lymph node hombWng transmission of growth signals. and presentation of chemnokincs and growth factors to traveling cells. CD44 surface proteins are associated with mnetatic urmors and CD44 has been used as a marker for HIV infction. Certain splice variants are associated with metastasis arnd poor prognosis of can=e patients. Therefore, molecules having homsology with CD44 are of particular interest. as their homology indicates that they may have functions rlate to those functions of CD44. CD44 is further described in U.S. Patent Nos. 5.506.119. 5,504,194 and 5.108.904; Gerberick, et al., Toxicol. A221l Puarmacol., l46(1):1 (1997), Wittig, et al., Immnl.letters (Netherlads), 57(l-3):217 (1997); and Oliveira and Odell, OralQi~l. (England), 33(4):260 (1997).

Efort are being unidert.aken by both imiwtay and academia to identify new, native secreted and membranebound receptor proteins, particularly transmembrane proteins with homology to CD44 antigen. Many efforts are focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel secreted and membrane-bound receptor proteins. Examples of screening methods and techniques are described in the literature [see, for example, Klein ct al.. Proc. Nall. Acad. Sci-, 9:7108-7113 (1996); U.S. Patent No.

5,536.637)].

We herein describe the identification and characterization of novel polypeptides having homnology to CD44 antigen, designated herein as PR0263 polypeptdecs.

33. PRQZ270 Thioredoxins effect reduction-wcidation (redox) gte. Many diseases are potentially related to redox state and reactive oxygen species may play a role in many important biological processes. Te transcription factors, NF-kappa B and AP- 1, are rcgulated by redox state and are known to affect the expession of a large variety of genes thought to be involved in thc pathogenesis of AIDS, cancer, atherosclerosis and diabetic complications. Such proteins mhay also playa role in cellular antioxidant defense, and in p athological conditions involving oxidative stress such as stroke and inflammnation in addition to having a role in apoptosis. Therefore, thoredoxins, and proteins having homnology thereto.

are of interest to the scientific and medical communities.

We herein describe the identification and characterization of novel polypeptides having homology to tioredoxin. designated herein as PR0270 polypeptides.

34. PRO271 Tke procoglycan link protein is a protein which is intimately associated with various extracelular matrix proteins and more specifically with proteins sucb as collagen. For example. one primary component of' collagen is a large proteoglycan called aggrecan. This molecule is retained by binding to the glYcwsaminoglycan hyaluronan through the amino terminal GI globular domain of the core protein. T1his binding is stabilized by the proteoglycan link protein which is a protein that is also associated with other tissues containing hyaluronan binding proteoglycans such as versican.

Link protein has been identified as a potential target for antoirmmune antibodies in individuals who suffer fzomjuvmile rheimlitid arthritis (see Guerassinov et al.. J. Rhewnarology 24(5):959-964 (1997)). As such, there is strong intrest in identifying novel protens having homology to link protein. We herein describe the identification and characterization of novel polypeptides having such homology. designated herein as PR0271 polypepades.

*35. PRQ2 Retictilocalbin is an endoplasmic retacular protein which may be involved in protein tanspout and himinal protein processing. Reticulocalbin resides in the lumen of the endopladsmic reraculurn, is Imown to bind calcium.

and may be involved in a luniinal retention mechanism of the endoplasmic reticulun. It contains six donains of the EF-band motif associated with high affinity calcium binding. We describe hmrin the identification and characterization of a novel polypeptide which has homology to the reticulocalbin protein, designated herein as PROM7.

36. PRQ24 Collagen, a naturally occurring protein, finds wide application in industry. Chemically hydrolyzed natural collagen can be denatured and renatured by heating and cooling to produce gelatin, which is used in photographic :and medical, among other applications. Collagen has important properties such as the ability to form interthain aggregates having a conformation designated as a triple. helix. We herein describe die identification and characterization of a novel polypeptide which has homology to portions of the collagen molecule, designated herein as PROM9.

37. The integrins comprise a supergene family of cell-surface glycoprotein receptors that promnote cellular adhesion. Each cell has mimu receptors that define ks cell adhesive capabilities. lntegin are involved in a wide variety of interaction between cells and other cells or matrix components. The integrins are of particular importance in regulating movement and function of immune system cells The platelet [lblIA integrin complex is of particular inporta=e in regulating platelet aggregation. A member of the integria family, integrn P-6, is expressed on epithelial cells and modulates epithelial inflammation. Another integrin, leucocyte-associated antigen-l (LFAl) is important in the -theion of lymphocytes during an immune response. The integrins are expressed as hcterodixnrs of noncovalently associated alpha and beta subunits. Given the physiological importanc of cell adhesion mechanisms in Wvo efforts ame currently being under taken to identify new, native proteins which are involved in cell adhesion. We describe herein the identification and characteration of a novel polypeptide which has homology to inzcgrin.

designated herein as PR0295.

38. PRO293 Protein-protein interactions include receptor and antigen complexes and signaling mechanisms. As more is known about the structural and funcional mechanisis; underlying protein-protein interaerioni. protein-protein interactions can be morm easily manipulated to regulate tbe particular result of the protein-protein interaction. Thus.

the underlying mechanisms of protein-protein interactions arc of interest to the scientific and medical community.

All proteins containin leucine-rich repeats are thought to be involved in protein-protein interactions.

Lcucine-rich repeats are short sequence motifs present in a number of proteins with diverse functions and cellular locations. Ile crystal structure of riborrucleasec inhibitor protein has revealed that leucine-rich repeats correspond to bera-alpba structural units. These units are arranged so that they form a parallel beta-sheet with one surface :9 exposed to solvent. so that the protein acquires in unursual. nonglubular shape. These two features; have been iicedas responsible for the protein-binding funictions of proteins containing leucine-ricli repeats. SeKobe and Deisenhofer. Trends Biocbem. Sci., 19(10):415A421 (Oct. 1994).

15 A study has been reported on leucine-rich proteoglycans which serve as tissue organizers, orienting and ordering collagen fibrils during ontogeny and are involved in pathological proceses such as wound healing, tissue repair, and tumor stronra formation. lozzo. R. Critr Rev- Biochemn Mot DiOW., 32(2):141-174 (1997). Others studies implicating leucine rich proteins in wound healing and tissue repair arc De IA Salle, et Vouv. Rev.

Fr ernhatm. (Germany). 37(4)215-22 (199:5), reporting inutations in the leucine rich motif in a complex associated 20 with the bleeding disorder Bcrnard-Soulier syndrome and Chlemetson, K. Ibwmb. Hamosj. (Germany), 74(1):1 11-116 (July 1995). reporting that platelets have leucine rich repeats. Another protein of particular interest which has been reported to have leucine-rich repeats is the SLIT protein which has been reported to be useful in treating neuro-degecrativc diseasecs such as Alzheimer's disease, nrve damage such as in Parktinson's disease, and for diagriosis of cancer, see, Antavanistsakoms, S. and Rothberg. J. WO92lOS1S-AI by Yale University. Other saudie reporting on the biological functions of proteins having Icucine-rich repeats include: -Tayar. ct al.. Mgj Ce1lEndo2cdno. W- Ieand). 12(1-2):6S-70 (Dec. 1996) (gonadosroiin receptor involvement); Miura, et al., WN..EnahU~ (Japan), 54(7):1794-1789 (July 1996) (apoptosis involvement); Harris, P. et al., L-Am-..Soc.

NkphM.. 6(4):1125-1 133 (Oct. 1995) (kidney disease involvement); and Ruoslahti, E. et al.. W091 10727-A by La Jolla Cancer Research Foundation (decorin binding to tranforming growth factorfi involvement for treatment for cancer, wound healing and scarring).

Efforts are therefore being undertaken by both industy and academia to identify new proteins having leucine rich repeats to better understand protein-protein interactioas. Of particular interest are those proteins having Icucine rich repeat and homology to ktnown ncuronal leucx= rich repeat proteins. Many efforts are focused on the screening of mammnalian recombinant DNA Libraries to Idtfy the coding sequences for novel secreted and nMembrane-bound proteins having leucine rich repeats. Examples of screening methods and technique s are described in the literature [see, for example. Klein et al.. Proc. NaI Acad- Sci. 21:7108-7113 (1996); U.S. Patent No. 5.536.637)).

We describe herein the identification and characterization of a novel polypeptide which has homology to leucine rich repeat proteins, designated herein as PR0293.

39. PROW4 Prote n-protein interactions include receptor and antigen complexes and signaling rnechanisms. As more is known about the stnuctural and functional mechanisms underlying protein-protein interactiotts. protein-protein interactions can be more easily manipulated to regulate the particular result of the protein-protein interaction. Thus, the underlying mechanisms of protein-protein interactions arc of interest to the scientific and medical community.

All proteins containing leucine-iich repeats are thought to be involved in protein-protein interactions.

Lcucine-rich repeats arc short sequence motifs present in a number of proteins with diverse futions and cellular locations. The crystal structure of ribonuclease inhibitor protein has revealed that leuicine-rich repeats correspond to beta-alpha structural units. These units are arranged so that they form a parallel beta-sheet with one surface exposed to solvent, so that the protein acquires an untusual, oonglubular shape. These two featurs have been indicated as responsible for the protein-binding functions of proteins containing leucine-rich repeats. See, Kobe and Deisenhofer, Trends Biochem. Sci., 19(10):415-421 (Oct. 1994).

A study has been reported on leucine-rich proteoglycans which serve as tissue organizers, orienting and ordering collagen fibrils during ontogeny and art involved in pathological processes such as wournd healing, tissue repair, and tumnor stroma formation. lozzo. R. Crit Rev- 1iochem. Mot. iol., 32(2):141-174 (1997). Others studies implicating leucine rich proteins in wound healing and tissue repair are De La Salle, ct al., Ywix..Rev- Er. Hmatwl. (Germany). 37(4):215-222 (1995). reporting mutations In the leuciiie rich motif in a complex associated with the bleeding disorder Bernard-Souicr syndrome and Chlcmnetson. Y. 1bromb. Haen~. (Germany).

74(l):111-116 (July 1995). reporting that platelets have leucine rich repeats. Another protein of particular interest which has been reported to have leucine-rich repeats Is the SLIT protein which has been reported to be useful in treatin u-degenerative diseases such as Alzheimer's disease, nerve damage such as in Parkinson's disease, and for diagnosis of cancer. see, Aztavanistsakonas, S. and Rorhberg, 1. W09210518-Al by Yale University. Other studies reporting on the biological funrctions of proteins having leucirie-rich repeats include: Tayar, et al., MPL CeIl Edorinol., (Ireland), 125(1-2):65-70 (Dec. 1996) (gonadotropin receptor involvement): Miura, et al..

Myz n Rinlm (Japan), 54(7:1784-1799 (July 1996) (apoptosis involvement); Harris. P. et al.. 1.Am- So Neihro.. 6(4).1125-1133 (Oct. 1995) (kidney disease involvement), and Ruoslahti, E. L, et al., W091 10727-A by La Jolla Cancer Reseazrch Foundation (decorin binding to transforming growth factorfl involvement for treatment for cancer, wound healing and scarring).

Dcnsin is a glycoprotein which has been isolated from the brain which has all the hallmarks of an adhesion molecule. It is highly concentrated at synaptic sites in the brain and is expressed prominently in dendritic processes in developing neurons. Densin has been characterized as a member of the 0-inked sialoglycoproteins. Densin has relevance to medically important processes such as regencration. Given the physiological importance of synaptic processes and cell adhesion mechanisms in vivo. efforts are curenty being under taken to identify new, native proteins which are involved in synaptic machinery and cell adhesion. Densin is fuirther described in Kennedy, M.B, Trends Nmrosi. (England). 20(6)-.264 (1997) and Apperson, el al., 1L Nurosci. 16(21):6839 (1996).

Efforts are trfor being undertaken by both industry and academia to identify new proteins having leucine rich repeats to better understandi protein-protein interactions. Of particular interest arc those proteins havig leucine rich repeats and homnology to known proteins having leucine rich repeats such as KIAO31 and densin. Many efforts arc focused on the -cenn of mammalian recombinant DNA libraries to identify the coding sequences for novel secreted and membrane-bound proteins having leucinc rich repeats. Examples of screening methods and techniques arm described in the literature (sme for example, Klein e al., Proc. Nad Acad -Si 23:7108-7113 (1996); U.S. Patent No. 5,536,637)].

We describe herein the identification and characterization of a novel polypeptide which has homology to leucine rich repeat proteins. designated herein as PR0247.

40. PR0302. PR0303. rR0304. PRO307 and PR0343 Proteases are enzymatic proteins whiich are involved in a large number of very imiportant biological processes in mammalian and non-mammalian organisms. Numerous different protease enzymes fronm a variety of different -mmalan and non-manralian organismis have been both identified and characterized. The mammalian* protease enzymes play important roles in manty different biological processes including, for example, protein digestion activation, inactvation, or modulation of peptide hormone activity, and alteration of the physical properties of proteins and enzymes.

In light of the important physiological roles played by protease enzymes, efforts are currently being undertaken by both industry and academia to identify new, native protease hornologs. Many of these efforts are focused on the screening of mamnmalian recombinant DNA libraries to identify the coding sequences for novel :20 secreted and membrane-bound receptor proteins. Examples of screening methods and techniques are described in the literature (see, for example, Klein et al., Proc. Na] Acad. Sci, 91:7108-7113 (1996); U.S. Patent No.

5,536,637)]. We herein describe the identification of novel polypeptides having homology to various protease enzymes, designated herein as PR0302. PR0303, PR0304. PRO3O7 and PR0343 polypeptides.

41. PR32 The GLIP protein family has been characterized as comprising zinc-finger proteins which play important roles in emnbryogenesis. These proteins may function as transcriptional regulatory proteins and are known to be amplified in a subset of humn tumors. Gliorna pathogenesis protein is smictually related to a group of plant parlxigalesis-related prottins. It is highly expressed in glioblastorna. See US Pat. Nos. 5,582,981 (issued Dec. 1996) and 5.322.801 (issued June 21, 1996), Ellington. A.D. et al., ture, 3M~:818 (1990). Grindley, J.C. et al., Del. il 182):337 (1997), Marine. et al., Mech. f=21l (1997), The CRISP orecysteine rich secretory protein family are a group of proteins which ame also structurally related to a group of plant pathogenesis proteins. [Schwdetzky, inthem- n1:325 (1997), Pfisterer. hMaLCjL~inL. 1=(f:6160 (1996), Krarzschinrar. L. i. JR~eL 733l827 (1996)]. We describe herein the identification of a novel polypeptide which has homology to GLIP and CRISP, designated herein as PR0328 polypeptides.

42. PROMS5. PRO331 and PR0326 Protein-protein interactions include receptor arid antigen complexes and signaling mechanisms. As mare is known about dhe structural and functional mechanisms underlying protein-protein interactions, protein-protein inerctons can be momt easily manipulated to regulate die particular result of die protein-protein inieraction. Thus, the underlying mechanisms of protein-protein interactions are of interest to the scientific and medical cormmunity.

AllI proteins conaining leucine-rich repeats are thought to be involved in protein-protein interactions.

Leucine-rich repeats are short sequcnce. motifs present in a number of proteins with diverse functions and cellular locations. The crystal structure of ribonuclease inhibitor protein has rcvcaled that leircine-rich repeats correspond to beta-alpha structural units. These units are arranged to that they form a parallel beta-sheet with one surface exposed to solvent, so that dhe protein acquires an urnusual. nonglubular shape. These two features have been indicated as responsible for the protein-binding funactions of proteins containing kcucine-rich repeats. See, Kobe and Deisenhofer. Trends 134ocem. Sei, 19(10):415-421 (Oct. 1994).

A study has been reported on leucine-rich proteogycans which serve as tissue organizers. orienting and ordering collagen fibrils during ontogeny and are involved in pathological processes such as wound healing, tissue repair, andl tmnor stroma formation. lozzo, R. Crit. Rev. Biochem. Mol. Biol., 32(2):141-174 (1997). Other studies implicating leucine rich proteins in wound healing and tisse repair are De La Salle, et al., Vouv. v Er. Hematol. (Germany). 37(4)-215-m (1995), reporting imutations in t Icucine rich motif in a complex associated with the bleedirg disorder Beruard.Saulier syndrome, Chlemetson, K. 1hbz, Haemost. (Germany). 74(1): 111 116 (July 199:5), reporting that platelets have leucine rich repeats and Ruoslahti. E et al.. W09110727-A by La Jolla Cancer Research Foundlation reporting that decarin binding to transforming growth factorp has involvement in a treatment for cancer, wound healing and scarring. Related by function to this group of proteins is the insulin like growth factor (laP), in that it is useful in wtxrnd-healing and associated therapies concerned with re-growth of tissue, such as connective tissue, skin and bone; in promoting body growth in hums and animatls; and in stimulating other growth-related processes. The acid labile subunit of IGF (ALS) is also of interest in that it increases the half-life of IGF and is part of t IGF complex in vivo.

Another protein which has been reported to have leucine-rich repeats is dhe SLIT protein which has been reported to be useful in treating neuro-degenerative diseases such as Alzheimer's disease, nerve damage such as in Parkinson's disease, and for diagnosis of cancer, see, Artavanistsakonas, S. and Rothberg, J. W09210518-A I by Yale University. Of particular interest is UG-1. a membrane glycoprotein that is expressed specifically in glial cells in the mouse brain, and has leucine rich repeats and irnmunoglobulin-like domains. Suzuki, et al., 1. Bial.

Chem. 271(37)-27522 (1996). Other studies reporting on the biological functions of proteins having leucinc: rich repeats include: Tayar, et al.. Mol. Cell Endocrinol., (Ireland), 125(1-2):65-70 (De. 1996) (gonadotropin receptor involvement); Miura, et al., Nj22n..Rinj (Japan), 54(7):1784-1789 (July 1996) (apoptosis involvermn); Harris, P. et al, 1. Am. Soc. NeoRalW.. 1125-1133 (Oct. 1995) (kidney disease involvement).

Efforts are thereIxe being undertaken by both industry and academia to identify new proteins having leucine rich repeats to better understand protein-protein interactions. Of particular interest ame those proteins having leucine rich repeats and homology to known proteins having leucine rich repeas such as LIG-1. ALS and decorin. Many efforts ame focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel secreted and membrane-bound proteins having Icucine rich repeats. Exaimples of screening methods sand tochniques am descrbed in teliteratre see, for exntple. Klein et al.. Proc. Mdt. Acad. Sci. .2:7108-7113 (1996); U.S. Patent No. 5,536.637)).

We describe herein the identification and characterization of novel polypeptdecs which have homology to proteins of the Icucine rich repeat superfamily, designated herein as PROM.5 PRO331 and PR0326 polypeptides.

It% 43. PR0332 Secreted proteins comnprising a repeat characterized by an arrangement of conserved leuine residues (lwdeun-rich repeat rnoti) have diverse biological roles. Certain proteoglycans, such as biglycan, fibromodulin and decorin, are, for example. characterized by the presence of a leucine-sich repeat of about 24 amino adids [Ruoslahti, Ann. Rev. Cell. Bl-. 29-255 (1988): Oldberg e al., EN413 1. 2601-2604 In general, proteoglycans ame believed to play a role in regulating extracellular matrix, cartilage or bone function. The proteoglycan decorin binds to ccilagen type Iand and afetste rate of fibril formation. Fibromoodulin also binds collagen and delays fibril fomai Both fibronmodui and decorin inhibit the activity of transforming growth factor beta (TOFP3) (U.S.

Patent No. 5.583.103 issued December 10. 1996). TGF-P is knowni to play a key role in the induction of extracellular matrix and has been implicated in the development of fibrotic diseases. such as cancer and glomerulonephritis. Accordingly. proteoglycans have been proposed for the treatment of fibrotic cancer, based upon ~tir ability to inhibit TOP-'s growth stitmulating activity on the cancer cell. Proteoglycans have also been described as potenially useful in the treatment of other proliferative paftologies, including rheumatoid arthritis, arteriosclerosis, adult respiratory distress syndrome, cirrhosis of the liver, fibrosis of the hangs, post-myocardial infarction, cardiac 20 fibrois, post-angioplasty restenosis. renal interstitial fibrosis and certain dermal fibrotic conditions, such as keloids and scarring, which might result from burm inuries, other invasive skin injaies. or cosmetic or reconstructive surgery Patent No. 5,654.270. issued August 5, 1997).

:We describe herein the identification and cliaracterization of novel polypeptides which have homology to proteins of the leircine rich repeat superfamily, designated herein as PR0332 polypeptides.

44. PRO334 Microfibril bandles and proteins found in association with these bundles, particularly attachment molecuiles, -are of interest in the field of dermatology, particularly in the study of skin 'which has been damaged from aging, injuries or the sun. Fibriuin niicrofibrils define the continuous elastic network of skin. and are present in dermis as microfibril bundles devoid of measurable elastin extending from the dermal-epithelial Junction and as components of the tdick elastic fibre present in the deep reticular dermis. Moreover, Marfan syndrome has been linked to mutations which interfere with multimierization of fibrillin monomers or other connective tissue, elements.

Fibulin-l is a modular glycoprotein with amnino-terminal anaphlatoxin-lile modules followed by nine epidermal growth factor (EGP)-lie modules and, depending on alternative splicing, four possible carboxyl terminit.

Fibulin-2 is a novel extracellular matrix protein frequently found in close association with microfibrils containing either fibronectin or fibrillin. 7bus. fibriilin. fibulin. and molecules related thereto are of interest, particularly for the use of preventing skin from being damaged from aging, injuries or the sun, or for restoring skin damaged from sam. Moreover, these molecules arc generally of jitet in the study of connective tissue and attachment mnolecules andi related mechanisus. Fibrillin. filmlin and related inxoicse are further described in Adams. eta&L. J. MolL iol..

272(2):226-36 (1997); Kielty and Shutdcworth, Microse. Res- Tech.. 38(4):413-27 (1997); and Chil J .ad Silz&. 12(2Supp.):131.5 (1997).

Currently, efforts arc being undertaken by both industry and academia to identify new, native secreted and mnbrni-bound receptor proteins, particularly s ted proteins which have homology to fibulin ane-fibrillin. Many efforts ame focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel secreted and raenibram-boxui receptor proteins. Examples of screening methods and techniques arc described in the literature (sc. for examplc. Klein ct al., Proc- Nall Acad. Sci., 23:7108-7113 (1996); U.S. Patent No.

5.536,637)].

We herein describe the idevitification and characteriaion of novel polypptdes having homology to fibulin and fibrilin, designated herein as PR0334 polypeptides.

PRO346 71ie widespread occurnce of cancer has prompted the devotion of considerable resources and discovering new treatments of treaunent. One particular method involves the creation of tumor or cancer specific mnonoclonal antibodies (mAbs) which are specific to tumor antigens. Such mAbs, which can distinguish between normal and cancerous cells are useful in t diagnosis, prognosis and treatment of the disease. Particular antigens arc known to be associated with neoplastic diseases, such as colorectal and breast cancer. Since colon cancer is a widespread disease, early diagnosis and treatmen is an important medical goal. Diagnosis and treatment of cancer can be 20 implemented using mnonoclonal antibodies (muAbs) specific therefore having fluorescent, nuclear magnetic or ~.radioactive tags. Raidioactive genes, toxins and/or drug tagged mAbs can be used for treatment in sizu with minimal patient description.

:Carcinoembryonic antigen (CEA) is a glycoprotein found in human colon cancer and the digestive organs of a 2-6 month humanemnbryos. PEA is a known hruma aumor marker and is widely used in the diagnosis of neoplastic diseases. such as colon cancer. For example, when the serum levels of CEA am elevated in a patient, a drop of CEA levels after surgery would indicate the tumnor resection was successful. On dhe other hand, a subsequent rise in serum CEA levels after surgery would indicate that axiastases of the original tumor ay have formed or that 9. new primary tumors may have appeared. CEA may also be a target for antisense nucleotides 46. PR0269R Protein disullide isomersc is an enzymatic protein which is involved in the promnotion of correct refolding of proteins through the establishment of correct disulfide bond formation. Protein disulfide isomerase was inaitially identified based upqon its ability to catalyze the renaturation of reduced denatured RNAse (Goldberger et al., J. Bin!.

Chem. 239:1406-1410 (1964) and Epstein et al.. Col S)inHarbor Symp. Quant. Biol. 28:439-449 (1963)). Protein disulfide isomerase has been shown to be a resident enzyme of the endoplasmic recticuluim which is retained in the endoplasinic reticulum, via a -KDEL or -HDEL amino acid sequence at its C-terminus.

Given the importance of disulfide bond-forming enzymes and their Potential uses in a ruamber of different applications. for example in increasing the yield of correct refolding of recombinantly Produced proteins, efforts art currently being undertken by both industry and academia to identify new, nativc proteins having homology to protein disulfide isomerase. Many of these efforts are focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel protein disulfide isomerase homologs. We herein describe a novel polypeptide having homology to protein disulfidc isomerase. designated herein as PR0268.

47. PR0330D Prolyl 4-hydroxylase is an enzyme which functions to post-translationally bydroxylate proline residues a( the Y position of the amino acid sequence Gly-X-Y, which is a repeating three amino acid sequence found in both collagen, and procollagen. Hydroxylation of proline residues at the Y position of the Gly-X-Y amino acid triplet to form 4-hydroxyproline residues at those positions is required before newly synthesized collagen polypeptide chains ~~may fold int their proper the~iesmltriple-he~ical conformation. If bydroxylation does not occur. synthesized .~.collagen polypeptides remain non-helical, are poorly secreted by cells and cannot assemble into stable functional collagn fibrils. Vuozio et al.. Proc. NatL Acad. Sci. USA 89:7467-7470 (1992). Prolyl 4-hydroxytase is comprised of at least two different polypeptide subunits. alpha and beta.

Efforts are being undertaken by both industry and academia to identify new, native secreted and membrane- 6 bound receptor proteins. Many efforts are focused on the screening of mammalian recombinant DNA libraries to identify the coding sequences for novel secreted and membrane-bound receptor proteins. Examples of screening methods and techniques are described in the literature [see, for example, Klein et al.. Proc. Nati. Acad. Sci., 20 22:7108-7113 (1996); U.S. Patent No. 5,536.637)]. Based upon these efforts. Applicants have herein identified and describe a novel polypepide having homology to the alpha subunit of prolyl 4-hydroxylase. designated herein as PR0330.

*00048. PR0339 and PR0310 Fringe is a protein which specificall blocks serrate-mediated activation of notch in the dorsal compartment of dhe Drosophila wing imaginag disc. Fleming, et al.. Dyc1olgn n. 124(1S):2gr73-SI (1997). Therefore, fringe is of interest for both its role in development as well as its ability to regulate serrate, particularly serrate's signaling Os....abilities. Also of interest are novel polypeptides which may have a role in development and/or the regulation of serrate-lie molecules. Of particular interest are novel polypeptides having homology to fringe as identified and described herein, designated herein as PR0339 and PRO310 polypeptides.

49. EL02M Lectins ame a class of proteins comprising a region that hinds carbohydrates specifically and non-covalently.

Numerous lectins have been identified in higher animals, both membrane-bound and soluble, -and have been implicated in a variety of cell-recognition phenomena and tumor metastasis.

Most lectins can be classified as either C-type (calcium-dependent) or S-type (thiol -dependent).

34 Lectins are thought to play a role in regulating cellular events that are initiated at the level of the plasma membrane. For example, plasma membrane associated molecules are involved in the activation of various subsets of lymphoid cells, e.g. T-lymphocytes, and it is known that cell surface molecules are responsible for activation of these cells and consequently their response during an immune reaction.

A particular group of cell adhesion molecules, selectins, belong in the superfamily of C-type lectins.

This group.includes L-selectin (peripheral lymph node homing receptor (pnHR), LEC-CAM-1, LAM-1, gp 9 0 MEL, gp00EL, GPllO0EL, MEL-14 antigen, Leu-8 antigen, TQ-1 antigen, DREG S: antigen), E-selectin (LEC-CAM-2. LECCAM-2, ELAM-1), and Pselectin (LEC-CAM-3, LECAM-3, GMP-140, PADGEM). The structure of selectins consists of a C-type lectin (carbohydrate binding) domain, an epidermal growth factorlike (EGF-like) motif, and variable numbers of complement regulatory (CR) motifs. Selectins, are associated with 20 leukocyte adhesion, e.g. the attachment of neutrophils to venular endothelial cells adjacent to inflammation (Eselectin), or with the trafficking of lymphocytes from blood to secondary lymphoid organs, e.g. lymph nodes and Peyer's patches (L-selectin).

Another exemplary lectin is the cell-associated macrophage antigen, Mac-2 that is believed to be involved in cell adhesion and immune responses. Macrophages also express a lectin that recognizes Tn Ag, a human carcinomaassociated epitope.

Another C-type lectin is CD95 (Fas antigen/APO-l) that is an important mediator of immunologically relevant regulated or programmed cell death (apoptosis).

"Apoptosis" is a non-necrotic cell death that'takes place in metazoan animal cells following activation of an intrinsic cell suicide program. The cloning of Fas antigen is described in PCT publication WO 91/10448, and European patent application EP510691. The mature Fas molecule 34a consists of 319 amino acids of which 157 are extracellular, 17 constitute the transmembrane domain, and 145 are intracellular. Increased levels of Fas expression at T cell surface have been associated with tumor cells and HIVinfected cells. Ligation of CD95 triggers apoptosis in the presence of interleukin-1 (IL-2).

C-type lectins also include receptors for oxidized low-density lipoprotein (LDL). This suggests a possible role in the pathogenesis of atherosclerosis.

We herein describe the identification and characterization of novel polypeptides having homology to C-type lectins, designated herein as PRO244 polypeptides.

Throughout the description and claims of this specification, the word "comprise" and variations of the word, such as "comprising" and "comprises", means "including but not limited to', and is not intended to exclude other additives, components, integers or steps".

SUMMARY OF THE INVENTION 1. PRO211 and PR0217 20 Applicants have identified cDNA clones that encode novel polypeptides having homology to EGF, designated in the present application as "PRO211" and S* "PR0217" polypeptides.

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PRO211 or PR0217 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding EGF-like homologue PRO211 and PR0217 polypeptides of Fig. 2 (SEQ ID NO:2) and/or 4 (SEQ ID NO:4) indicated in Fig. 1 (SEQ ID NO:1) and/or Fig. 3 (SEQ ID NO:3), respectively, or is complementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PRO211 and PR0217 EGF-like homologue PR0211 and PR0217 polypeptides. In particular, the invention provides isolated native sequence PR0211 and PRO217 EGFlik bomologue polypeptides, which. in one embodiment. includes an amino acid sequence comprising residues: I to 353 of Fig. 2 (SEQ ID NO:2) or 1 to 379 of Fig. 4 (SEQ ID NO: 4).

2. PRO230 Applicants have identified a cDNA clone that encodes a novel polypeptidc. whcrein the polypeptide is designated in the present application as *PR0230". t' In one embodiment. the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0230 polypeptide. In one aspect. the isolated nucleic acid comprises DNA encoding the PR0230 polypeptide having amino acid residues 1 throuigh 467 of Figure 6 (SEQ ID NO: 12). or is complemientary to such encoding raaleic acid sequec, andi remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0230 polypeptide. In particular, t invention provides isolated naive sequence PR02230 polypeptide. wich in one embodiment. inchudes an amino acid sequence comprising residues I through 467 of Figure 6 (SEQ ED NO: 12).

:In another embodiment, the invention provides an expressed sequence tag (ES7) comprising the nuclcotide sequence of SEQ ID NO: 13 (Figure 7) which is herein designated as DNA20088.

3. PRO232 Applicants have identified a cDNA clone that encodes a novel polypeptide, wherein the polypeptide is designated in the present application as "PR0232.

20 In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding aR03 oyetd.I n set h sltdncecai opie N noigteP03 oyetd having amino acid residues I to14 of Figure9 (SEQ ID NO, 18). or is complementry to such encoding nucleic acid sequence. and remans stably bound toit under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0232 polypeptide. In particular. the invention provides isolated native sequence PR0232 polypeptide. which in one embodiment, includes an amino acid sequence comprising residues I to 114 of Figure 9 (SEQ ID NO:18).

4. PRO187 Applicants have identified a cDNA clone that encodes a novel polypeptide, designated in the present application as "PR0187*.

In one embodiment. the invention provides an isolated nucleic acid molecule comprising DNA encoding a PROl87polyppidc. [none aspect, the isolated nucleic acid comprises DNA encoding the PROWB polypepuide of Figure 11I (SEQ ID N0:23), or is complementary to such encoding nucleic acid sequence, and remains stably bound to it under at leat moderate, and optionally, under high stringency conditions. In another aspect, dhe invention provides a nucleic acid comprising the coding sequer of Figure 10 (SEQ ID NO=fl or its complement. In another aspect, the invention provides a nucleic acid of the full length protein of clone DNA27864-1155. deposited with the ATCC under aicesion mnber ATCC 209375. alternatively die coding sequence of clone DNA27864-11S5.

deposited under accession rnumber ATCC 209375.

In yet another einbodiarnt. te invenion proviles isolated PRO 187 polypeptide. ln particular, die invention provides isolated native sequence PROI 87 polypeptide. which in ont embodiment, inludes an amino acid sequence comprising residues I to 205 of Figure I11 (SEQ ID 140:23). Mcenativly. the invention provides a polyetdec encoded by the nucleic acid deposited under accession number ATCC 209375.

S. PRO265 Applicants have identified a cDNA clone that encodes a novel polypeptide, wherein the polypeptidc is designated in the present application as *PR026S'.

In ore embodiment. the invention provides an isolated nucleic: acid molecule comprising DNA encoding a PR0265 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding the PR0265 polypeptide having amino acid residues I to 660 of Figure 13 (SEQ ID NO:28). or is complementary to such encoding nucleic acid seqe.e and remain stably bound to it under at least moderate, and optionally, under high stringenicy ~.conditions.

In another embodiment. the invention provides isolated PR0265 polypeptide. In particular, the invention provids isolated native sequence PRO265 polypeptide. which in one embodiment, includes an amino acid sequence compising residues I to 660 of Figure 13 (SEQ ID 140:28). An additional embodiment of the present invention is directed to an isolated extraoelardomiain of a PRO265 polypeptide.

6. PRO219 20 Applicants have identified a eDNA clone that encodes a novel polypeptide. wherein the polypeptide is designated in the present application as *PR0219*.

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0219 polypeptide. In one aspect, the isolated nucleic adid comprises DNA encoding the PRO219 polypeptide having amino acid residues I to 915 of Figure 15 (SEQ ID NO:34). or is complementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodint,. the invention provides isolated PR0219 polypeptide. In particular. tie invention provides isolated native sequence PR0219 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues I to 915 of Figure 15 (SEQ ID 140:34).

7. PRQ246 Applicants have identified a eDNA clone that encodes a novel polyetide. wherein the polypeptide is designated in the present application as *PRO246'.

Inaone embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0246 polypeptide. Inaone aspect, the isolated nuecic acid comprises DNA encoding the PR0246 polypeptide having amino acid residues I to 390 of Figure 17 (SEQ ID 140:39), or is complementary to such encoding nucleic acid sequece,. and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0246 polypepade. In particular. dhe invention provides isolated native sequence PR0246 polypeptide, which in one embodiment, includes an amino acid sequence comprisinig residues I to 390 of Figure 17 (SEQ ID NO:39). An additional embodiment of the present invention is directed to an isolated extracellular domain of a PR0246 polypepdec.

S. PRO228 Applicants have identified a eDNA clone tha enmcds a novel polypeptide having homology to CD97, EMR1 and latrophilin, wherein the polypeptide is designated in the present application as *PR0228'.

In one embodiment, the invention provides an isolated nucleic acid woleailc comprising DNA encoding a PR0228 polypeptdce. In one aspect, die isolated nucleic ac-id comprises DNA encoding dhe PR0228 polypeptide having amino acid residues I to 690 of Figure 19 (SEQ ED NO:49), or is complementary to such encoding nucleic: acid seuewc, andl remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

:In another embodiment, die invention provides isolated PR0228 polypeptide. In particular, the invention provides isolated native sequence PR0228 polypeptide, which in one embodiment, includes an amnin acid sequence comprising residues I to 690 of Figure 19 (SEQ ID NO:49). An additional embodiment of the present invention is directed to an isolated extracellular domain of a PR0228 polypeptide.

In another embodiment, the invention provides an expresed sequence tag (ESi) comprising the nucleotide sequenc of SEQ MD NO:S0. designated herein as DNA21951.

ERQS3 Applicants have identified a cDNA clone (DNA49435-12 19) that encodes a novel polypeptide, designated in the present application as PR0533.

In one embodiment, the invention provides-an isolated nuecic acid molecule having at least about sequence identity to a DNA molecule encoding a PR0533 polypeptide comprising die sequence of amino acids 23 to 216 of Figure 22 (SEQ ID N0:59). or the complement of the DNA molecule of The sequence identity preferably is about 85%, more preferably about 90%, most preferably about 95%. ho one aspect, the isolated nucleic acid has at least about 80%, preferably at least about 85%, rer preferably at least about 90%, and most preferably at least about 95 sequence identity with a polypeptide having amin acid residues 23 to 216 of Figure 22 (SEQ MD NO:59). Preferably, the highest degree, of sequence identity occurs within the secreted portion (amino acids 23 to 216 of Figure 22, SEQ ID NO:59). In a further embodiment, the isolated nucleic acid molecule comprises DNA encoding a PR0533 polypeptide having amino acid residues 1 to 216 of Figure 22 (SEQ ID NO:59), or is comnplenntuary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and .pinly under high stringency coxitlions. In another aspect, the invention provides a nucleic; acid of the fuM length protein of clone DNA49435-1219. deposited with the ATCC under accession number ATCC 209480.

In yet another emnbodiment, the invention provides isolated PR0533 polypeptide. In particular. the invention provides isolated native sequence PR0533 polypeptide, which in one embodiment, includes an amino acid sequence comprising residus 23 io 216 of igure 22 (SEQ ID NO.59). Native PR0533 polypeptides with or without the native signal sequrc= (amino acids I to 22 in Figure 22 (SEQ ID NO:S9)). and with or without the initiating methioniu art specifically included. Alternatively, the invention provides a PR0533 polypeptide encoded by t nucleic acid deposited under accession umber ATCC 209480.

10. PRO24SIt Applicants have identified a cDNA clone that encodes a novel polypcptide, wherein the polypeptide is designated in the present application as 'PR024S".

In one embodiment. the invention provides an isolated nucleic acid mnolecule comprising DNA encoding a PR0245 polyepde. In one aspect. the isolated nucleic acid comprises DNA encoding the PR0245 polypeptide having amirnacid residues It u312 of Fig. 24 (SEQ ID NO:64), or is complementary to such encoding nuecic acd sequence and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0245 polypetdde. In particular, the invention provides isolated native sequenc PR0245 polypeptide, which in one embodiment, includes an amino acid sequence con~isingresidues 1 to 312 of Figure 24 (SEQ ID NO:64).

It. P110220. P110221 aind PR10227 Applicants have identified cDNA clones thant each ecode novel polypeptides, all having leucine rich repeats.

These polypeptides are designte in the present application as PR0220. PR0221 and PR0227.

In one embodiment. the invention provides isolated nucleic acid molecules comprising DNA respcctivcly :20 encoding PR0220. PR0221 and PR0227. respectively. In one aspect, provided herein is an isolated nucleic acid cmrISes DNA encoding the PR0220 polypeptide having amino acid residues I through 708 of Figure 26 (SEQ ID NO:69). or is complementary to such encoding nucleic acid sequece,. and remains stably bound to it under at leat moderate. and optionally, under high stringency conditions. Also provided herein is an isolated nucleic acid cmrssDNA etcoding the PR0221 polypeptide having amino acdd residues 1 throgh 259 of Figure 28 (SEQ ID NO:71), or is complementary to such encoding nucleic acid sequee and remains stably bound to-it under at least moderate, and optionally, under high stringency conditions. Moreover, also provided herein is an isolated nucleic aci comprises DNA encoding the PRO2W polypeptide having amino acid residues I through 620 of Figure 30 (SEQ IW NO:.73). or is complementary to such encoding ucleic acdd sequene, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another emnbodiment. the invention provides isolated PP10220. PR0221 and PRO227 polypeptides. In particular, provided herein is the isolated native sequence for the PR0220 polypeptide. which in one embodiment, includes an amino acid sequence comprising residues 1 to 708 of Figure 26 (SEQ ID NO: 69). Additionally provided herein is the isolated native "epm for the PRO221 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues I to 259 of Figure 28 (SEQ MD NO:? Moreover, provided herein is the isolated native sequence for ft PR0227 polypeptdec, which in one embodiment. includes an amino acid sequence comprising residues I to 620 of Figure 30 (SEQ ID NO:73).

12. PR02SB APPlicant have identified a cDNA clone th a odes a novel polypeptide having homology to CRTAM and poliovinus receptor precursors, wherein the polypeptide is designated in the present application as "PR02S8'.

In ocroebodiment, the invention provides an isolated aucleic acid molecule comprising DNA encoding a PR0258 polypeptide. In one aspect. the isolated nucleic acid comprises DNA encoding die PR0258 polypeptide having amino adid residues I to 398 of Figure 32 (SEQ ID NO:84). or is complemntary to suckaencading nucleic acid sequence, and remains stAlIy bound to it under at least moderate, and optionally, wnder high stringency conditions.

In another embodiment, the invention provides isolated PR0258 polypeptdec. In particular, the invention provides isolated native sequesice PR0258 polypeptide, which in one embodiment. includes an amino acid sequence comprising residues 1 to 398 of Figure 32 (SEQ ID NO:94). An additional embodiment of t present invention is directed to an isolated extracellular domain of a PR0258 polypeptide.

13. PRQ2( :Applicants have identified a cDNA clone that encodes a novel polypeptide. wherein the polypeptide is designated in the present application as *PR0266'.

In one embodiment. die invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0266 polypeptide. In one aspcct. the isolated nucleic acid comprises DNA encoding the PR0266 polypeptide having amino acid residues I to 696 of Figure 34 (SEQ ID NO:91), or is complementary to such encoding mucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0266 polypeptide. In particular, the invention provides isolated native sequesme PR0266 polypeptide. which in one embodiment, includes an amino acid sequence comprising residues I to 696 of Figure 34 (SEQ ID NO:91).

14. PROQZ2 Applicants have identified a cDNA clone that encodes a novel polypeptide, wherein the polypeptide is designated in the present application as PR0269.

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0269 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding the PR0269 polypeptide having amino acid reskdues I to 490 of Fig. 36 (SEQ ID NO:96). or is complemntary to such encoding nucleic; acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0269 polypeptide. In particular, the invention provides isolated native sequec PR0269 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues I to 490 of Figure 36 (SEQ ID NO:96). An additional embodiment of the present invention is directed to an isolated extracliu domain of a PRO269 polypeptide.

PRQ21 Applicants have identified a cDNA clone that encodes a novel polypeptide, wherein the polypeptide is designated in the present application as PR0287'.

in one embodiment, dhe invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0287 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding the PR0287 polypeptide having amino acdresidues I to 415 of Fig. 38 (SEQ ID NO:l104). or is complemntary to such encoding nuecec acid sequence. and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment the invention provides isolated PR0287 polypeptide. In particular. the invention provides isolated native sequence PR0287 polyetide, which in one enzbodiiat, includes an amino acid sequec comprising residues I to 4 15 of Figure 38 (SEQ ID NO: 104).

16. PRO214 Applicants have identified a cDNA clone that encodes a novel polypeptide, designated in the present application as "PRO214*.

one embodiment. the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0214 polyeptide. In one aspect, the isolated nucleic acid comprises DNA encoding the PR0214 polypeptide of Fig. 40 (SEQ ID NO: 109). or is complemrentary to such encoding nucleic acid sequence, amd remains stably bound to it under at least moderate, and optionally, under high stringency conditions. In another aspect, the invention provids a MUCleic acid comprising the coding sequence of Fig. 39 (SEQ ID NO: 108) or its complenxenL In another aspect, die invention provides a nucleic acid of the ful length protein of clone DNA32286-1191. deposited with 20 ATCC udracsinnumber ATCC 209385. isltdP04poypie.npatcarthivnin In yt aothe emodbrntthe invention providesisltdP 21 oyeie.Ipaicarthinnin provides isolated native sequence PR0214 polypeptde, which in one embodimert, includes an amino acid sequence comprising the residues of Figure 40 (SEQ ID NO: 109). Alternatively, the invention provides a polypeptide encoded by the nucleic acid deposited under accession number ATCC 209385.

17. PRO317 Applicants have identified a eDNA clone that encodes a novel polypeptide, designated in the present application as "PR0317*.

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding PRO317 polypeptide. In one aspect, the isolated nucleic acid comprises DNA (SEQ ID NO: 113) encoding PR0317 polypeptide having amino acid residues 1 to 366 of Fig. 42, or is complemcntary to such encoding nucleic acid sequence. and remains stably bound to it under at least moderate, and optionally, unde high stringency conditions.

In another embodiment. the invention provides isolated PR0317 polypetide. In particular, the invention provides isolated native-sequence PR0317 polypeptide, which in one eniodiment, includes an amino acid sequence comprising residues I to 366 of Figure 42 (SEQ ID NO: 114).

In yet another embodiment, the invention supplies a method of detecting the presence of PR03 17 in a sample, the method comprising: a) contacting a detectable anti-PRO3l7 antibody with a sample suspected of containing PRO3 17. and b) dewcesin binding of the antibody to die sample, wherein the sample is selected from the group consisting of a body fluid, a tissue sample. a cell extract, and a cell culture medium.

In a still further embodiment a method is provided tar determining the presence of PRO3 17 mRNA in a sample. the method comprising: a) contacting a sample suspected of containing PR0317 mnRNA with a detectable nuclelk acid probe that hybridizes under moderate to stringent conditions to PR0317 mRNA; and b) detecing hybridization of the probe to the samle.

Preferably, in this method the sample is a tissue sample and the detecting step is by in rint hybridization.

or the sample is a cell extract and detection is by Norther analysis.

Further, the invention provides a method for treating a PR0317-usociated disorder comprising administering to a mmmal an effective amount at the PR0317 polypeptide or a composition thereof containng a carrier, or with an effective amount of a PR0317 agonist or PR0317 antagonist. such as an antibody which binds ~.specifically to PR0317.

is. .RQ301 Applicants have Identified a cDNA clone (DNA40628-1216) that encodes a navel polypeptide. designated in the present application as PR0301.

In one embodiment. the invention provides an isolated nucleic acid molecule having at least about S0% sequence identity to a DNA molecule encoding a PRO301 polypeptide comprising the sequence af amino acids 28 to 258 of Fig. 44 (SEQ ID NO: 119), or dhe complement oftthe DNA molecule oft(a). Mwe sequence identity preferably is about 85%, more preferably about 90%,ms preferably about 95 In one aspect, the isolated nucleic acid has at least about 80%. preferably at least about 85 morm preferably at least about 90 and most preferably at least about 95% sequence identity with a polypeptide having amino acid residues 28 to 258 of Fig. 44 (SEQ MD NO:119). Preterably, the highest degree of sequence identity occmr within the extracellular domains (amino acids 28 to 258 of Fig. 44, SEQ IM NO:1 19). In afurther embodimient, the isolated nucleic acid molecule comprises DNA encoding a PRO301 polypeptide having amino acid residues 28 to 299 of Fig. 44 (SEQ ID NO:119). or is complernentay to such aeoding nucleic acid sequence. and remains stably bound to It under at least moderate, and optionally, und1 high stringency conditions. In another aspect, the invention provides a nucleic acid of the full length protein of clone DNA40628-1216, deposited with the ATCC under accession number ATCC 209432, alternatively the coding sequence ofcdone DNA40628-1216, deposited under accession number ATCC 209432.

In yet another embodiment, the invention provides isolated PRO301 polypeptide. In particular, the invention provides isolated native sequence PRO301 palypeptide, which in one embodiment, includes an amino acid sequence comprising the extracelkilar domain residues 28 to 258 of Figure 44 (SEQ ID NO: 119). Native PRO301 polypcptides with ofrwirbothemnaive signal sequence (amino acids 1 to 27 in Figure 44 (SEQ MDNO: 119), and with or without the initiating mneth onine ame specifically included. Additionally, thc sequences of the invention may also comprise the transimbranC domain (residues 236 to about 258 in Figre 44; SEQ ID NO: 119) and/or the intracellular domain (about residue 259 to 299 in Figure 44; SEQ ID .NO:119). Alternatively, the invention provides a PRO301 potypeptide encoded by tie nucleic acid deposited under accession riumber ATCC 209432.

19. PRQ224 Applicants have identified a cONA clone that encodes a novel polypeptidc, wherein the polypeptide is designated in the present application as *PR0224'.

In one embodiment, the invention provides an isolated nuceic acid molecule eomprising-DNA encoding a PR0224 polypeptide. In one aspect, the isolated nudeic acid comprises DNA encoding the PR0224 polypcptide having m acid residues I to 282 of Figure 46 (SEQ MD NO:127). or is comp lemnittary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, amid optionally, under high stringency conditions.

In another embodiment. the invention provides isolated PRO224 polypeptide. In particular, the invention provides isolated native sequecec PR0224 polypeptide. which in one embodment, includes an amino acid sequenoe ::comrising residues I to 282 of Figure 46 (SEQ ID NO: 127).

EEQ22 Applicants have identified a eDNA dlone that encodes a novel polypepuide. wherein the polypeptide is designated in die present application as 'PR222'.

In one embodiment. the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0222 polypeptide. In one aspect, the isolated =ucic acid comprises DNA encoding the PR0222 polypeptide having amino acid residues I to 490 of Fig. 48 (SEQ WD NO: 132), or is complementary to such encoding nucleic acid :20 sequence, andi remains stably bound to it under at least moderate. and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated MR0222 polypeptide. In particular, the invention provides isolated native sequence PR0222 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues I to 490 of Figure 48 (SEQ ID NO: 132).

21. PR23 Applicants have identified a cDNA dlone that encodes a novel lectin polypeptide molecule, designated in the present application as OPR0234'.

In one embodiment, the invention provides an isolated nucleic acid encoding a novel lectin comprising DNA encoding a PR0234 polypeptide. In one aspect, the isolated nucleic acid comprises the DNA encoding PR0234 polyqdcdes having ammo acid residue I to 382 of Fig. 50 (SEQ ID NO:137), or is coamplemnentary to such encoding nucei acid sequate and remains stably bound to it under at least moderate, and optionally, under high stringency conditions. In another aspect, the invention provides an isolated nucleic acid molecule comprising die nucleotide sequence oftFig. 49 (SEQ ID NO: 136).

In another embodiment. the invention provides isolated novel PR0234 polypeptides. In particular, the invention provides isolated native sequence PR0234 polypeptdde. which in one embodiment, includes an amino acid sequcnce comprising residues I to 382 of Figure 50 (SEQ ID NO: 137).

In yet anothier emodiment. dhe invention provides oligomicleotide probes useful for isolating genomic and cDNA nucicotide sequences.

22. PRO231 Applicants have identified a cDNA clone that encodes a novel polypeptide having homology to a putative acid phosphatase, wherein the polypeptide is designated in thic present application as 9 PR0231 In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0231 polypeptde. In one aspect, the isolated nucleic acid comprises DNA encoding die PR0231 polypeptide having amino acid reidiies 1 to 428 of Fig. 52 (SEQ ID NO- 142). or is complementary to such emod Ong nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally. under high stringency conditions.

In another embodiment, dhe invention provides isolated PR0231 polypeptide. In particular, the invention provides isolated native sequence PR0231I polypeptide. which in one embodiment. includes an amino acid sequenice *compriing residues I to 428 of Figure 52 (SEQ ID NO:.142).

*23. PROZ2 Applicants have identified a cDNA clone that encodes a novel polypepdde having homology to scavenger OLeS receptors wherein the polypeptide is designated in the present application as 'PR0229".

In one emnbodimen, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0229 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding thc PR0229 polypeptide having amninn acid residues It to 347 of Figure 54 (SEQ ID NO: 148). or is complementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringerncy *..*conditions.

In another embodiment, dhe invention provides isolated PR0229 polypeptide. In particular, the invention prov~ides Msoated native sequence PR0229 polypeptide, which in one embodiment, includles an amino acid sequence comprising residues 1 to 347 of Figure 54 (SEQ ID NO: 148).

24. IPRO23 Applicants have identified a eDNA clone that encodes a novel polypeptdec having homology to reductase, wherein die polypeptide is designated in the present application as "PR0238*.

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0238 polypeptide. Inone aspect, the isolated nucleic acid comprises DNA encoding the PR0238 polypeptide having amofno acid residues I to 3 10 of Figure 56 (SEQ ID NO: 153). or is complementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0238 polypeptide. In particular, the invention provides isolated native sequence PR0238 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues I to 3 10 of Figure 56 (SEQ ID NO: 153).

PRO233 Applicants have identified a cDNA clone that encodes a novel polypeptide. wherein the polypeptide is designated in the present application as "PR0233'.

In one emtbodimtent, the invention provides an isolated micleic acid molecule comprising DNA encoding a PR0233 polyetide. In one aspect, the isolated nucleic acid comprises DNA encoding the PR0233 polypeptide havingamno acid reidues 1 to 300of FigureS58 (SEQ ID NO: 159), or is complementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally. uinder high stringency conditions.

In another embodiment, the invention provides isolated PR0233 polypeptide. In particular. the invention provides isolated native sequence PRO233 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues 1 to 300of Figurc 58(SEQ ID NO: I 9).

26 .RO22 Applicants have identified a cDNA clone that encodes a novel polypeptide having homology to wcine carboxypeptidase polypeptides, wherein the polypeptidc is designated in the present application as 'PR0223*.

In one embodiment. the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0223 polypeptide. In one aspect. the isolated nucleic acid comprises DNA encoding the PR0223 polyetide having amino acid residues 1 to 476 of Figure 60 (SEQ ID NO: 164). or is complementary to such encoding nucleic acid sequence. and remains stably bound to it under at least moderate, and optionally. under high stringncy 20 conditions.

In another embodiment, the invention provides isolated PR0223 polypeptide. In particular, the invention provides isolated native sequence PR0223 polypeptide, which in one embodiment, includes an amino acid sequence comprising reidue I to 476 of Figure 60 (SEQ ID NO 164).

.27. PRM Applicants have identified a cDNA clone that encodes a novel polypeptide, wherein the polypeptide is designated in the present application as *PR0235' In one embodiment, the invention provides an isolated nucleic; acid molecule comprising DNA encoding a PR0235 polypeptide. In one aspect. the isolated nucleic acid comprises DNA encoding the PR0235 polyetide having am- acid residues 1 to 552 of Figure: 62 (SEQ ID NO: 170), or is complementary to such encoding nucleic acid sequence, and remiains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0235 polypeptide. in particular, the invention provides isolated native sequence PR0235 polypeptide. which in one embodiment, includes an amino acid sequence comprising residues I to 552 of Figure 62 (SEQ ID NO: 170).

28. PRO236 and PRO262 Applicnts have idenified eDNA clones that encode novel polypeptides having homology to P-gaiactosidasc, wherein those polypeptides ame designated in the present application as 'PR0236' and *PR0262'.

In omnemuodimet the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0236 pokfpeptide. In one aspect, the isolated nuceic acid comprises DNA encoding the PR0236 polypeptdec having amino acid rsidues I to 636 of Figure 64 (SEQ ID NO: 17S), or is coimplementary to such encoding nucleic acid sequece. and remains stably bound to it under at least moderate, and optionally. under high stringency conditions.

In another embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0262 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding the PR0262 polypeptide having amino acid reidues 1 to 654 of Figure 66 (SEQ ID NO: 177). or is complementary to such encoding nuclexc acid sequence, and remains stably bound to it under at least moderate, anid optionally. under high stringency conditions.

In another emjbodiment, the invention provides isolated PR0236 polypepuide. In particular, the invention provides isolated native sequence PR0236 polypeptide, which in one embodimen, includes an amino acid sequence comprising residucs 1 to 636 of Figure 64 (SEQ ID NO: 175).

In another embodiment, the invention provides isolated PR0262 polypeptide. In particular. the invention provides isolated native sequence PR0262 polypeptide. which in one embodiment, includes an amino acid sequence comprising residues 1 to 654 of Figure 66 (SEQ ID NO: 177).

29. PREQZ39 Applicants have identified a cDNA clone that encodes a novel polypeptide, wherein the polypeptide is designated in the present application as 'PR0239'.

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0239 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding the PR0239 polyetide having amino acid rcsidues 110o 501 of Figure 68 (SEQ ID NO- 185). or is complementary to such encoding nucleic acid sequence. and remains stably bound to it under at Icast moderate, and optionally, under high stringency conditions.

In another embodimrent, the invention provides isolated PR0239 polypeptide. In pa nocular, the invention provides isolated native sequence PR0239 polypeptide, which in one embodimn~t, includes an amino acid sequence comprising residues I to 501 of Figure 68 (SEQ ID NO: 185).

Applicants have identified a eDNA clone that encodes a novel polypeptide, wherein the polypcptide is designated in the present applicaionM as "PR0257'.

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0257 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding the PR0257 polypeptide having amino nod residues I to 607 of Figure 70 (SEQ ID NO: 190), or is complementary to such encoding nucleic acid sequence. and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

in another emibodimient. the invention provides isolated PR0257 polypeptidc. In particular. the invention provides isolated native sequence PROMS polypeptide. which in one embodiment. includes an amino acid Sequence compriing ridues Ito 607of Figurc 70(SEQ ID NO:190). An additional embodiment of the present invention is directed to an isolated extraceflular domain of a PRO2S7 polypepuide.

31. PRO2602 Applicants have identified a cDNA clone that encodes a novel polypeptidc, wherein the polypeptide is designated in the present application as 'PR0260'.

I one emibodimnent, die invention provides an isolaed trucleic acid molecule comprising DNA encoding a PR0260 polypetide. In one aspect, the isolated nucleic acid comprises DNA encoding the PR0260 polyetide having anuno acid residues I to 467 of Figure 72 (SEQ MD NO: 195). or is complementary to such encoding nucleic :.:acid sequence. and remain stably bound to it under at least moderate, and optionally, underhigh stringency conditions.

I another embodiment, the invention provides isolated PR0260 polypeptide. In particular. the invention provides isolated native sequence PR0260 polypeptide. which in one embodiment, includes an amino acid sequence comprising residues I to 467 of Figure 72 (SEQ W NO: 195).

032. PR26 Applicants have identified a cDNA clone that encodes a novel polypeptide having homology to CD44 antigen, wherein the polypeptide is designated in the present application as 'PRO263".

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a *.:PR0263 polypeptide. In one aspect. the isolated nucleic acid comprises DNA encoding the PR0263 polypeptide having amino acid residues I to 322 of Figure 74 (SEQ ID NO:201). or is complementary to such encoding nuecic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0263 polypepude. In particular, dhe invention provides isolated native sequence PR0263 potypeptide, which in one embodiment, includes an amino acid sequence comprising residues I to 322 of Figure 74 (SEQ ID NO.201). An additional embodiment of the present invention is directed to an isolated extrAcellular domain of a PR0263 polypeptide.

33. PRQZ270 Applicants have identified a cDNA clone that encodes a novel potypeptide. wherein the polypetide is designated in the present application as "PR0270".

In one embodiment. the invention provides an isolated micleic acid molecule comprising DNA encoding a PR0270 polypeptide. I one aspect, tie isolated nucleic acid comprises DNA whiveti includes the sequence encoding the PR0270 polypeptide having amino acid residues I to 296 of Fig. 76 (SEQ ID) NO:207), or is complementary to such encoding nucleic acid sequence. and remains stably bound to it under at least moderate, and optionally, under high tringency conditions.

In another embodiment, the invention provides isolated PR0270 polypeptidc. In particular, the invention provides isolated native sequence PR0270 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues 1 to 296 of Figure 76 (SEQ ID NO:207).

34. PRO271 Applicants have identified a cDNA clone that encodes a novel polypeptide having homology to the protcoglycan link protein, wherein the polypeptide is designated in the present application as "PRO271'.

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0271 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding the PRO271 polypeptide having amino acid residues 1 to 360 of Figure 78 (SEQ ID N0:213). or is complementary to such encoding nucleic acid sequence, and remains stably bound to it unde at least moderate, and optionally, under high stringency •conditions.

In another embodiment, the invention provides isolated PR0271 polypeptide. In particular, the invention 15 provides isolated native sequence PRO271 polypeptide. which in one embodiment, includes an amino acid sequence comprising residues 1 to 360 of Figure 78 (SEQ ID NO:213).

PROQ22 2Applicants have identified a cDNA clone that encodes a novel polypeptidc, wherein the polypcptide is 20 designated in the present application as "PRO272".

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0272 polypeptide. In one aspect, the isolated nucleic acid comprises DNA encoding the PRO272 polypeptide having amino acid residues I to 328 of Figure 80 (SEQ ID NO:221), or is complementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0272 polypeptide. In particular, the invention provides isolated native sequence PRO272 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues 1 to 328 of Figure 80 (SEQ ID NO:211).

36. PRO294 Applicants have identified a cDNA clone that encodes a novel polypcptide, wherein the polypeptide is designated in the present application as 'PR0294".

In one embodiment, the invention provides an isolated nucleic acid molecule comprising DNA encoding a PR0294 polypeptide. In one aspect. the isolated nucleic acid comprises DNA encoding the PR0294 polypeptide having amino acid residues I to 550 of Figure 82 (SEQ ID N0:227), or is complementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

48 In another enbodinent, the invention provides isolated PR0294 polypeptide. In particular, the invention provides isolated native sequence PR0294 polypeptide, which in one enTbodinent, includes an amino acid sequence ccnprising residues 1 to 550 of Figure 82 (SEQ ID N3:227).

37. PRO295 Applicants have identified a cENA clone that encodes a novel polypeptide, wherein the polypeptide is designated in the present application as "PR0295".

S 10 In one embodiment, the invention provides an isolated nucleic acid molecule caoprising ENA encoding the PRO295 polypeptide. In one aspect, the isolated nucleic acid ccmprises INA encoding the PRO285 polypeptide having amino acid residues 1 to 350 of Figure 84 (SEQ ID N0:236), or is carplementary to such encoding nucleic acid sequence, and 15 remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another ebodiment, the invention provides isolated PRO295 polypeptide. In particular, the invention provides isolated native sequence PRO295 polypeptide, which in one embodiment, includes an amino 20 acid sequence caoprising residues 1 to 350 of Figure 84 (SEQ ID NO:236).

38. PR0293 Applicants have identified a cENA clone that encodes a novel human neuronal leucine rich repeat polypeptide, wherein the polypeptide is designated in the present application as "PR0293".

In one embodiment, the invention provides an isolated nucleic acid molecule ccnprising INA encoding a PR0293 polypeptide. In one aspect, the isolated nucleic acid ccaprises IMA encoding the PRO293 polypeptide having amino acid residues 1 to 713 of Figure 86 (SEQ ID N3:244), or is carplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0293 polypeptide. In particular, the invention provides isolated native sequence PR0293 polypeptide, which in one embodiment, includes an amino acid sequence carprising residues 1 to 713 of Figure 86 (SEQ ID ND:244).

An additional embodinent of the present invention is directed to an isolated extracellular domain of a PR0293 polypeptide.

49 39. PR247 Applicants have identified a cENA clcne that encodes a novel polypeptide having leucine rich repeats wherein the polypeptide is designated in the present application as "PI)247'.

In one enbodinent, the invention provides an isolated nucleic acid molecule conprising ENA encoding a PRO247 polypeptide, In one aspect, the isolated nucleic acid carprises EA encoding the PR0247 polypeptide having amino acid residues 1 to 546 of Figure 88 (SED ID NO:249), or is caplenentary to such encoding nucleic acid sequence, and remain stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0247 polypeptide. In particular, the invention provides isolated native sequence PRO247 polypeptide, which in one erbodimrent, includes an amino 15 acid sequence carprising residues 1 to 546 of Figure 88 (SB ID )NO:249).

An additional etbodinent of the present invention is directed to an isolated extracellular daain of a PRO247 polypeptide.

POD302, PRO303, PRO304, PRO307 and PRO343 20 Applicants have identified cMA clones that encode novel polypeptides having hanology to various proteases, wherein those polypeptide are designated in the present application as "PR0302", *PRO303", "PRO304-, -PR0307' and "PRO343" polypeptides.

In one enbodinent, the invention provides an isolated nucleic acid nolecule carprising Onk encoding a PRO302 polypeptide. In one aspect, the isolated nucleic acid carprises A encoding the PRO302 polypeptide having amino acid.residues 1 to 452 of Figure 90 (SEQ ID N:254), or is carplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another enbodirrent, the invention provides an isolated nucleic acid nolecule carprising DEA encoding a PR0303 polypeptide. In one aspect, the isolated nucleic acid carprises IA encoding the PRO303 polypeptide having amino acid residues 1 to 314 of Figure 92 (SEQ ID ND::256), or is carplermntary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In yet another enbodiment, the invention provides an isolated nucleic acid molecule carprising MNA encoding a PR0304 polypeptide. In one 50 aspect, the isolated nucleic acid molecule co prises ENA encoding a PRO304 polypeptide having amino acid residues 1 to 556 of Figure 94 (SE ID N0:258), or is ccuplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides an isolated nucleic acid nolecule ccuprising ENA encoding a PRO307 polypeptide. In one aspect, the isolated nucleic acid ccuprises DNA encoding the PR0307 polypeptide having amino acid residues 1 to 383 of Figure 96 (SEQ ID 1N:260), or is cacplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another arbodinent, the invention provides an isolated nucleic acid molecule ccrprising ENA encoding a PR0343 polypeptide. In one oo .15 aspect, the isolated nucleic acid ccnprises DNA encoding the PRO343 polypeptide having amino acid residues 1 to 317 of Figure 98 (SEQ ID NO::262), or is cmaplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

20 In another embodiment, the invention provides isolated PR0302 polypeptide. In particular, the invention provides isolated native sequence PR0302 polypeptide, which in one embodiment, include an amino acid sequence ccnprising residues 1 to 452 of Figure 90 (SEQ ID ND:254).

S" In another embodiment, the invention provides isolated PRO303 polypeptide. In particular, the invention provides isolated native sequence PR0303 polypeptide, which in one embodiment, includes an amino acid sequence cmaprising residues 1 to 314 of Figure 92 (SEQ ID ND:256).

S In another embodiment, the invention provides isolated PRO304 polypeptide. In particular, the invention provides isolated native sequence PRO304 polypeptide, which in one embodiment, includes an amino acid sequence ccnprising residues 1 to 556 of Figure 94 (SEQ ID N3:258).

In another embodiment, the invention provides isolated PR307 polypeptide. In particular, the invention provides isolated native sequence PR0307 polypeptide, which in one embodiment, includes an amino acid sequence ccaprising residues 1 to 383 of Figure 96 (SEQ ID NO:260).

In another embodiment, the invention provides isolated PR0343 polypeptide. In particular, the invention provides isolated native sequence PR0343 polypeptide, which in one embodiment, includes an amino acid sequence ccaprising residues 1 to 317 of Figure 98 (SEQ ID NO:262).

51 41. PR0328 Applicants have identified a cINA clone that encodes a novel polypeptide, wherein the polypeptide is designated in the present application as "PRO328".

In one embodiment, the invention provides an isolated nucleic acid molecule ccprising EMA encoding a PR0328 polypeptide. In one aspect, the isolated nucleic acid cacprises ENA encoding the PR0328 polypeptide having amino acid residues 1 to 463 of Figure 100 (SEQ ID NO:284), or is complementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0328 polypeptide. In particular, the invention provides isolated native 15 sequence PRO328 polypeptide, which in one embodiment, includes an amino acid sequence caomprising residues 1 to 463 of Figure 100 (SEQ ID ND:284).

An additional embodiment of the present invention is directed to an isolated extracellular domain of a PR0306 polypeptide.

42. PRO335, PRO331 and PRO326 S:Applicants have identified three cNA clones that respectively encode three novel polypeptides, each having leucine rich repeats and horology to LIG-1 and ALS. These polypeptides are designated in the present application as PRO335, PRO331 and PRO326, respectively.

In one embodiment, the invention provides three isolated nucleic acid molecules camprising INA respectively encoding PR0335, PR0331 and PRO326, respectively. In one aspect, herein is provided an isolated nucleic acid ccaprising ENA encoding the PRO335 polypeptide having amino acid residues 1 through 1059 of Figure 102 (SeQ ID ND:289), or is carplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions. Also provided herein is an isolated nucleic acid ccnprises EM encoding the PRO331 polypeptide having amino acid residues 1 through 640 of Figure 104 (SBE ID NO:291), or is ccaplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions. Additionally provided herein is an isolated nucleic acid cmaprises ENA encoding the PR0326 polypeptide having amino acid residues 1 through 1119 of Figure 106 (SEQ ID NO: 293), or is ccmplementary to such encoding nucleic acid sequence, and remains 52 stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another erbodiment, the invention provides isolated PR0335, PRO331 and PR0326 polypeptides or extracellular domains thereof. In particular, the invention provides isolated native sequence for the PR0335 polypeptide, which in one embodiment, includes an amino acid sequence ccmprising residues 1 through 1059 of Figure 102 (SEQ ID ND:289). Also provided herein is the isolated native sequence for the PR0331 polypeptide, which in one enbodiment, includes an amino acid sequence cunprising 1 through 640 of Figure 104 (SEQ ID NO: 291). Also provided herein is the isolated native sequence for the PR0326 polypeptide, which in one embodiment, includes an amino acid sequence carprising residues 1 through S* 1119 of Figure 106 (SEQ ID NO: 293).

15 43. PR0332 Applicants have identified a cENA clone (MIA40982-1235) that encodes a novel polypeptide, designated in the present application as "PR0332".

In one embodiment, the invention provides an isolated nucleic 20 acid molecule ccnprising NA having at least about 80% sequence identity to a ENA molecule encoding a PR0358 polypeptide carprising the sequence of amino acids 49 to 642 of Fig. 108 (SEQ ID N):309), or the ccplement of the fMA molecule of The sequence identity preferably is about nore preferably about 90%, most preferably about 95%. In one aspect, the isolated nucleic acid has at least about 80%, preferably at least about nore preferably at least about 90%, and nost preferably at least about 95% sequence identity with a polypeptide having amino acid residues 1 to 642 of Fig. 108 (SEB ID NO:309). Preferably, the highest degree of sequence identity occurs within the leucine-rich repeat domains (amino acids 116 to 624 of Fig. 108, SEQ ID ND:309). In a further uebodiment, the isolated nucleic acid nolecule comprises INA encoding a PR0332 polypeptide having amino acid residues 49 to 642 of Fig. 108 (SEQ ID NO:309), or is ccmpletentary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another enbodiment, the invention provides isolated PR0332 polypeptides. In particular, the invention provides isolated native sequence PRO332 polypeptide, which in one embodiment, includes an amino acid sequence ccmprising residues 49 to 624 of Figure 108 (SEQ ID ND:309).

53 Native PRO332 polypeptides with or without the native signal sequence (amino acids 1 to 48 in Figure 108, SEQ ID NO: 309), and with or without the initiating methionine are specifically included.

44. PRo334 Applicants have identified a cCNA clone that encodes a novel polypeptide having hoxology to fibulin and fibrillin, wherein the polypeptide is designated in the present application as "PR0334'.

In one embodiment, the invention provides an isolated nucleic acid molecule coprising DNA encoding a PRO334 polypeptide. In one aspect, the isolated nucleic acid ccznprises DNA encoding the PR0334 polypeptide having amino acid residues 1 to 509 of Figure 110 (SEQ ID. N:314), or is carplementary Sto such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

15 In another embodiment, the invention provides isolated PRO334 polypeptide. In particular, the invention provides isolated native sequence PR0334 polypeptide, which in one entodiment, includes an amino acid sequence conprising residues 1 to 509 of Figure 110 (SEB ID N:314).

20 45. PR0346 Applicants have identified a cErA clone (ENA44167-1243) that encodes a novel polypeptide, designated in the present application as "PR0346".

In one embodiment, the invention provides an isolated nucleic acid molecule having at least about 80% sequence identity to a UNA molecule encoding a PR0346 polypeptide conprising the sequence of amino acids 19 to 339 of Fig. 112 (SEQ ID NO:319), or the carplement of the [NA molecule of The sequence identity preferably is about 85%, more preferably about most preferably about 95%. In one aspect, the isolated nucleic acid has at least about 80%, preferably at least about 85%, more preferably at least about 90%, and most preferably at least about 95% sequence identity with a polypeptide having amino acid residues 19 to 339 of Fig. 112 (SEQ ID N: 319). Preferably, the highest degree of sequence identity occurs within the extracellular damains (amino acids 19 to 339 of Fig. 112, SEQ ID NO:319). In alternative enbodiments, the polypeptide by which the horology is measured carprises the residues 1-339, 19-360 or 19-450 of Fig. 112, SEQ ID ND:319). In a further embodiment, the isolated nucleic acid molecule carprises UA encoding a PR0346 polypeptide having amino acid residues 19 to 339 of Fig. 112 (SEQ ID NO: 319), alternatively residues 1-339, 19-360 54 or 19-450 of Fig. 112 (SEQ ID NO: 319) or is ccplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions. In another aspect, the invention provides a nucleic acid of the full length protein of clone ENA44167-1243, deposited with the ACC under accession number ACC 209434, alternatively the coding sequence of clone ENA44167-1243, deposited under accession number A'TI 209434.

In yet another embodiment, the invention provides isolated PRO346 polypeptide. In particular, the invention provides isolated native sequence PRO346 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues 19 to 339 of Figure 112 (SEQ ID NO:319).

Native PR0346 polypeptides with or without the native signal sequence (residues 1 to 18 in Figure 112 (SEQ ID NO: 319), with or without the initiating methionine, with or without the transmeibrane damain (residues 15 340 to 360) and with or without the intracellular domain (residues 361 to 450) are specifically included. Alternatively, the invention provides a PRO346 polypeptide encoded by the nucleic acid deposited under accession number ATOC 209434.

S. 20 46. PR0268 Applicants have identified a cMNA clone that encodes a novel polypeptide having harology to protein disulfide iscmerase, wherein the polypeptide is designated in the present application as *PRO268'.

In one embodiment, the invention provides an isolated nucleic acid olecule carprising MA encoding a PRO268 polypeptide. In one aspect, the isolated nucleic acid comprises ]NA encoding the PRO268 polypeptide having amino acid residues 1 to 280 of Figure 114 (SEQ ID NO:324), or is comaplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PRO268 polypeptide. In particular, the invention provides isolated native sequence PRO268 polypeptide, which in one enbodiment, includes an amino acid sequence comrprising residues 1 to 280 of Figure 114 (S3E ID NO:324).

An additional embodiment of the present invention is directed to an isolated extracellular domain of a PR0268 polypeptide.

47. PRO330 Applicants have identified a cENA clone that encodes a novel 54a polypeptide having homology to the alpha subu.nit of proly-I 4-hydroxylase, wherein the polypeptide is designated in the present application as PRO33O'.

in one embxdiirent, the invention provides an isolated nucleic acid nolecule ccuprising IIPA encoding the PR0330 polypeptide. In one aspect, the isolated nucleic acid uolecule comiprising EM ecoding a PR0330 polypeptide having amino acid residues 1 to 533 of Figure 116 (SW) ID NO: 331), or is carpleentaxy to such encoding nucleic acid sequence, and rem~ains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another erboxit, the invention provides isolated PR0330 polypeptide. In particular, the invention provides isolated native seqence PR0330 polypeptide, which in one enibodiment, includes an amino acid sequence ccxnprising residues 1 to 533 of Figure 116 (SE2 ID JN):331).

48. PRO339 and PRO310 Aplicants have identified two~ crWM clones wherein each clone encodes a novel polypeptide having haiology to fringe, wherein the polypeptides are designated in the present application as "PRO339, and "PR310.

In one enbodimnit, the invention provides isolated nucleic acid mrolecules conprising USA encoding the PR0339 arid/or a PR0310 polypeptide.

In one aspect, the isolated nucleic acid ccrprises EVlA encoding the PR0)339 polypeptide having amino acid residues 1 to 772 of Figure 118 (=EX ID NO: 338), or is carplemeitary to such encoding nucleic acid sequence, and remains stably bound to it under at least nnderate, and optionally, under high stringency conditions. in another aspect, the isolated nucleic acid caiprises 1iM encoding the PRO310 polypeptide having amino acid residues 1 to 318 of Figure 120 (SEX2 ID) ND:340), or is canplementary to such encoding nucleic acid sequence, and remains stably bound to it under at least moderate, and optionally, under high stringency conditions.

In another ertbodi-et, the invention provides isolated PR03 39 as well as isolated PR0310 polypeptides. In particular, the invention provides isolated native sequence PR0339 polypeptide, which in one eatbodimm~t, includes an amino acid sequence cczrprising residues 1 to 772 of Figure 118 (SEQ ID ND: 338). Th -e invention further provides isolated native sequence PRO310 polypeptide, which in one effbodirit, includes an amino acid sequence ccuprising residues 1 to 318 of Figure 120 (SE)2 ID NJ: 340).

54b 49. PER244 Applicants have identified a clNA clone that encodes a novel polypeptide, designated in the present application as "PRO244".

In one embodiment, the invention provides an isolated nucleic acid nolecule conprising INA encoding PRO244 polypeptide. In one aspect, the isolated nucleic acid ccmprises INA encoding PRO244 polypeptide having amino acid residues 1 to 219 of Fig. 122 (SEQ ID NO:376), or is camplementary to such encoding nucleic acid sequence, and remains stably bound to it under at last moderate, and optionally, under high stringency conditions.

In another embodiment, the invention provides isolated PR0244 polypeptide. In particular, the invention provides isolated native sequence PR0244 polypeptide, which in one embodiment, includes an amino acid sequence comprising residues 1 to 219 of Figure 122 (SEQ ID NO:376).

50. Additional bodiments In other embodiments of the present invention, the invention provides vectors carprising INA encoding any of the above or below described polypeptides. A host cell ccnprising any such vector is also provided. By way of exanple, the host cells may be CHO cells, E. coli, or yeast. A process for producing any of the above or below described polypeptides is further provided and ccnprises culturing host cells under conditions suitable for expression of the desired polypeptide and recovering the desired polypeptide fran the cell culture.

25 In other enbodiments, the invention provides chimeric molecules cacprising any of the above or below described polypeptides fused to a heterologous polypeptide or amino acid sequence. An exanple of such a chimeric nolecule comprises any of the above or below described polypeptides fused to an epitope tag sequence or a Fc region of an immunoglobulin.

In another etbodiment, the invention provides an antibody which specifically binds to any of the above or below described polypeptides.

Optionally, the antibody is a monoclonal antibody.

In yet other embodiments, the invention provides oligonucleotide probes useful for isolating gencnic and cMNA nucleotide sequences, wherein those probes may be derived from any of the above or below described nucleotide sequences.

54c For the purposes of this specification it will be clearly understood that the word "comprising" means "including but not limited to', and that the word "comprises" has a corresponding meaning.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 shows a nucleotide sequence (SEQ ID NO:1) of a native sequence PRO211 cDNA, wherein SEQ ID NO:1 is a clone designated herein as "UNQ185" and/or "DNA32292-1131.

Figure 2 shows the amino acid sequence (SEQ ID NO:2) derived from the coding sequence of SEQ ID NO:1 shown in Figure 1.

4 Figure 3 shows a nucleotide sequence (SEQ ID NO:3) of 15 a native sequence PR0217 cDNA, wherein SEQ ID NO:3 is a clone designated herein as "UNQ191" and/or "DNA33094-1131".

Figure 4 shows the amino acid sequence (SEQ ID NO:4) derived from the coding sequence of SEQ ID NO:3 shown in Figure 3.

20 Figure 5 shows a nucleotide sequence (SEQ ID NO:11) 6* of a native sequence PR0230 cDNA, wherein SEQ ID NO:11 is a weer clone designated herein as "UNQ204" and/or "DNA33223-1136".

Figure 6 shows the amino acid sequence (SEQ ID NO:12) derived from the coding sequence of SEQ ID NO:11 shown in Figure 25 Figure 7 shows a nucleotide sequence designated *ses: herein as DNA20088 (SEQ ID NO:13).

•bOO oooo Figurc 8 shows a nucleotde sequesice (SEQ MD NO: 17) of a native sequence PR0232 cDNA. wherein SEQ ID NO: 17 is a clone designated herein as "UNQ206* and/or 'DNA3443541140-.

Figure 9 shows the amino acid sequence (SEQ ID 140:18) derived from the coding sequence of SEQ ID NO: 17 shown in Figure B.

Figure 10 shows a taidnodtde: sequer (SEQ MD NO:22) of a native sequence PR0187 eDNA. wherein SEQ ID NO:22 is a clone designated herein as "U1Q161' and/or "DNA27864-1155*.

Figure 11 shows (he amino acid sequence (SEQ MD NO:23) derived from the coding sequene of SEQ ID NO022 shown in Figure Figure 12 shows a nucklotide sequence (SE ID 14027) of a native sequence PR0265 eDNA, wherein SEQ ID NO:27 is a clone designated herein as "UN1Q232' and/or "D1A36350-1 158'.

Figure 13 shows the amino acid sequence (SEQ ID) NO:28) derived fr-om dhe coding sequence oftSEQ ID 140:27 shown in Figure 12.

Figures 14A-B show a rncleodde sequence (SEQ ID NO:33) of a native sequence PRO21 9 eDNA. wherein SEQ ID N0:33 is a clon designated herein as "UNQ193' arid/or 'DNA32290-l 164*.

Figure 15 shows the amino acid sequence (SEQ ID NO:34) derived from the coding sequence of SEQ ID NO:33 shown in Figures 14A-B.

Figure 16 shows a m1icleaide sequence (SEQ IID N&-38) of a naive sequence PR0246 cDNA, wherein SEQ ID NO:38 is a clone designated herein as OUNQ220" and/or ODNA35639-1172'.

140:38Figure 17 shows the amin acid sequence (SEQ ID NO:39) derived from the coding sequence of SEQ ID *O3 shown in Figure 16.

Figur 18 shows anuiclecode sequenc (SEQ ID 140:48) of a native sequence PRO228 eDNA, wherein SEQ ID NO:48 is a clone designated herein as 'UNQ2OV" and/or 'DNA33092-1202".

Figure 19 shows dhe amino acid sequence (SEQ ID 140:49) derived from the coding sequence of SEQ ID 140:48 shown in Figure 18.

Figure 20 shows a nucleotade sequence designated herein as DNA21951 (SEQ ID 140:50).

Figure 21 shows a mideoddec sequence (SEQ M NO.58) of a native sequence PR0533 cDNA, wherein SEQ ID 140:58 is a clone designated herein as OUNQ3440 and/or *D1A49435-1219'.

.0..Figure 22 shows die amino acid sequence (SEQ ID 10:5 9) derived from the coding sequence of SEQ ID 140:58 shown in Figure 21.

Figure 23 shows a raicloodde sequence (SEQ ID 10:63) of a native sequence PR0245 cDNA, wherein SEQ ID 14063 is a clone designated herein as 'UNQ219' and/or *DNA35638-I 141".

Figure 24 shows the amino acid sequence (SEQ ID 10:64) derived from the coding sequence of SEQ ID 140:63 shown in Figure 23.

Figure 25 shows a nucleotide sequence (SEQ ID NO:68) of a native sequence PR0220 cDNA. wherein SEQ ID 140:68 is a clone designated herein as 'UNQ194" and/or 'DNAM298.I 132'.

Figure 26 shows the amino acid sequence (SEQ ID 10:69) derived from the~coding sequence of SEQ ID 140:6 shown in Figure .Figure 27 shows a -zcloodde sequenc (SEQ ID NO:70) of a native sequence PR0221 I DNA. wherein SEQ ID NO:70 is a clone designated herein as 'UNQI95" and/or OD1A33089-l 132'.

Figure 28 shows the amino acid sequence (SEQ MD NOM7) derived from dhe coding sequence of SEQ ID shown in Figure 27.

Figure 29 shows a mxleodde sequnc (SEQ ID NO:72) of a native scquece= PR0227 cDNA. wherein SEQ M I NO:72 is a clone designated herein as IUNQ2Or and/or 'DNA33786-I 132'. I' Figure 30 shows the amino acid sequence (SEQ ID NO:73) derived from the coding sequence of SEQ ID NO:72 shown in Figure 29.

Figure 31 shows a micleooide sequence (SEQ U) NO:83) of a native sequence PR0258 cDNA, wherein S EQ [D NO:83 is a clone designated hercin as ULNQ22S' aior DNA3S9l8-1174*.

Figure 32 shows the amino acid sequence (SEQ ID NO:84) derived from the coding sequence of SEQ ID NO:83 shown in Figure 31.

Figurc 33 shows a inicloodde sequence (SEQ ID NO:90) of a native sequence PR0266 cDNA. wherein SEQ ID NO:90 is a clone designated herein as OUNQ233* and/or 'DNA37ISO0-1178'.

Figure 34 shows the amino acid sequence (SEQ ID NO-.91) derived from dhe coding sequence of SEQ ID NO:90 shown in Figure 33.

Figure 35 shows a micleodde sequence (SEQ ID NO:95) of a native sequence PR0269 cDNA, wherein SEQ MD NO:95 is a clone designated herein as *UNQ236' and/or *DNA38260-1190".

NO:95Figure 36 shows the amino acid sequence (SEQ ID NO:96) derived from the coding sequence of SEQ ID in Figure Figure 37 shows a nucleotide sequence (SEQ ID NO:103) of a native sequence PR0287 cDNA. wherein SEQ ID NO: 103 is a clone designated herein as 'UNQ2SO" and/or "DNA39969-1 185'.

:Figure 38 shows di amino acid sequence (SEQ ID NO: 104) derived from the coding sequence of S EQ ID NO: 103 shown in Figure 37.

Figure 39 shows a nucleozide sequence (SEQ ID NO:108) of a native sequence PR0214 cDNA, wherein SEQ ID NO:108 is a clone designated herein as 'UNQ188' and/or 'DNA32286-1 191"*.

Figure 40 shows t amino acid sequence (SEQ ID NO: 109) derived from the coding sequence of SEQ ID NO: 108 shown in Figure 39.

Figure 41 shows a nucleotide sequence (SEQ ID NO: 113) of a native sequenice PR0317 cDNA. wherein SEQ ID NO:113 is a clone designated herein as OUNQ278' and/or 'DNA33461-1 199'.

Figure 42 shows the amino acid sequence (SEQ ID NO: 114) derived from the coding sequence of SEQ ID NO: 113 shown in Figure 4 1.

Figure 43 shows a nucleodde sequence (SEQ ID NO: 18) of a native sequence PRO301 cDNA, wherein SEQ ID NO:1 18 is a clone designated herein as "UNQ264' and/or 'DNA40628-1216'.

Figure 44 shows the amino acid sequence (SEQ ID NO: 119) derived from the coding sequence of SEQ ID NO:118 shown in Figure 43.

Figure 45 shows a nucleotide sequence (SEQ ID NO: 126) of a native sequence PR0224 cDNA. wherein SEQ ID NO: 126 is a clone designated herein as "UNQ 198' and/or "DNA33221-1133'.

Figure 46 shows the amino acid sequence (SEQ ID NO:127) derived from the coding sequence of SEQ ID NO: 126 shown in Figure Figure 47 shows a nucleodde sequence (SEQ ID NO: 131) of a native sequence PRO222 cDNA. wherein SEQ ID NO:131 is a clone designated herein as "UNQI96" and/or "DNA33107-1135".

Figure 48 shows the amino acid sequence (SEQ ID NO:132) derived from the coding sequence of SEQ ID NO:131 shown in Figure 47.

Figure 49 shows a nucleodde sequence (SEQ ID NO:136) of a native sequence PRO234 cDNA, wherein SEQ ID NO:136 is a clone designated herein as *UNQ208' and/or "DNA35557-1137'.

Figure 50 shows the amino acid sequence (SEQ ID NO:137) derived from the coding sequence of SEQ ID NO:136 shown in Figure 49.

Figure 51 shows a nucleotide sequence (SEQ ID NO: 141) of a native sequence PRO231 cDNA, wherein SEQ ID NO: 141 is a clone designated herein as "UNQ205" and/or "DNA34434-1139".

Figure 52 sows the amino acid sequence (SEQ ID NO: 142) derived from the coding sequence of SEQ ID NO:141 shown in Figure 51.

Figure 53 shows a nucleotde sequence (SEQ ID NO:147) of a native sequence PRO229 cDNA, wherein 15 SEQ ID NO: 147 is a clone designated herein as "UNQ203" and/or "DNA33100-1159".

Figure 54 shows the amino acid sequence (SEQ ID NO:148) derived from the coding sequence of SEQ ID NO:147 shown in Figure 53.

Figure 55 shows a nucleodde sequence (SEQ ID NO: 152) of a native sequence PRO238 cDNA, wherein SEQ ID NO:152 is a clone designated herein as "UNQ212" and/or "DNA35600-1162".

20 Figure 56 shows the amino acid sequence (SEQ ID NO: 153) derived from the coding sequence of SEQ ID NO:152 shown in Figure Figure 57 shows a nucleotide sequence (SEQ ID NO: 158) of a native sequence PRO233 cDNA, wherein SEQ ID NO:158 is a clone designated herein as *UNQ207" and/or "DNA34436-1238".

Figure 58 shows the amino acid sequence (SEQ ID NO:159) derived from the coding sequence of SEQ ID 25 NO: 158 shown in Figure 57.

*Figure 59 shows a nucleotide sequence (SEQ ID NO: 163) of a native sequence PR0223 cDNA, wherein SEQ ID NO:163 is a clone designated herein as "UNQ197" and/or "DNA33206-1165".

Figure 60 shows the amino acid sequence (SEQ ID NO: 164) derived from the coding sequence of SEQ ID NO:163 shown in Figure 59.

Figure 61 shows a nucleotide sequence (SEQ ID NO: 169) of a native sequence PRO235 cDNA. wherein SEQ ID NO:169 is a clone designated herein as 'UNQ209" and/or "DNA35558-1167".

Figure 62 shows the amino acid sequence (SEQ ID NO: 170) derived from the coding sequence of SEQ ID NO:169 shown in Figure 61.

Figure 63 shows a nucleotide sequence (SEQ ID NO: 174) of a native sequence PRO236 cDNA, wherein SEQ ID NO: 174 is a clone designated herein as "UNQ210" and/or "DNA35599-1168".

Figure 64 shows the amino acid sequence (SEQ ID NO: 175) derived from the coding sequence of SEQ ID NO:174 shown in Figure 63.

Figure 65 shows a nucleodde sequnc (SEQ ID NO:176) of a native sequence PRO262 cDNA. wherein SEQ ID NO: 176 is a clone designated herein as *UNQ229 and/o r 'DNA36992-1168*.

Figure 66 sbows die amino acid sequence (SEQ ID NO: 177) derived from the coding sequence of SEQ MD NO: 176 shown in Figure Figure 67 shows a nucleotide sequee (SEQ ID NO: 184) of a native sequence PR0239 cDNA. wherein SEQ ID NO:184 is a clone designated herein as 'UNQ213" and/or "DNA3440Y7-1169*. It Figur 68 shows the amino acid sequence (SEQ 11) NO: 18S) derived from the coding sequence of S EQ I D NO: 184 shown in Figure 67.

Figure 69 shows a nucleotide sequece~ (SEQ ID NO: 189) of a native sequence PRO25'7 cDNA.7 wherein SEQ ID NO: 189 is a clone designated herein as "UNQ224* and/or ODNA35M41-1173'.

Figure 70 shows the amnino acid sequence (SEQ ED NO:190) derived from the coding sequence of SEQ ID NO: 189 shown in Figure 69.

Figure 71 shows a nucleotde sequence (SEQ ID NO: 194) of a native sequne PR0260 cDNA, wherein SEQ ID NO:194 is a clone designated herein as 'UNQ227* and/or "DNA33470-117S".

rW 72 &bows. the amin acid sequecc (eQ ID N:9)derived frmtecdigsqec of SEQ ID NO:I194 shown in Figure7 1.

Figure 73 shows a nucleotide sequence (SEQ ID 140:20) of a native sequence PR0263 cDNA, wherein -SEQ ID NO0:200 is a dlone designated herein as 'UNQ230" and/or *D1A34431-117r.

:Figure 74 shows the amino acid sequence (SEQ WD 10:201) derived from the coding sequence of SEQ ID 140:200 shown in Figure 73.

Figure 75 shows a nucleotde sequence (SEQ ID 140:206) of a native sequence PR0270 cDNA. wherein SEQ MD NO:206 is a clone designated herein as *IUNQ237' and/or *D1A39510-1181'.

Figure 76 shows the amino acid sequence (SEQ H) NO:207) derived from the coding sequence of SEQ ID NO:206 shown in Figure Figure 77 shows a nucleotde sequence (SEQ ED 140:212) of a native sequence PR0271 cDNA, wherein SEQ ID 140:212 is a clone designated herein as 9.1NQ238" and/or 'DNA39423-11ST'.

Fgre78 shows t amrin acid sequence (SEQ ID NO:213) derived from, the coding sequence of SEQ ID NO:212 shown in Figure 77.

.Figure 79 shows a nucleotide sequence (SEQ ID NO:220) of a native sequence PR0272 cDNA, wherein SEQ ID NO:220 is a done designated herein as "UNQ239' and/or 'DNA40620-1193.

Figure S0 shows the amino acid sequence (SEQ ID NO:221) derived from dhe coding sequence of SEQ ID NO:220 shown in Figure 79.

Figure 81 shows a nucleotide sequence (SEQ ID 140:226) of a native sequence PR0294 cDNA, wherein SEQ ID 140:226 is a clone designated hercin as wUNQ2S7' and/or "DNA40604-1l7".

Figure 82 shows the amino acid sequence (SEQ ID 140:227) derived from the coding sequence of SEQ ID 140:226 shown in Figure 81.

Figure 83 shows a nucleotide sequence (SEQ ID NO:235) of a native sequence PR0295 cDNA. wherein SEQ ID 140:235 is a done designated herein as "U1Q258' and/or "DNA38268-1188.

59 Figure 84 shows the amino acid sequence (SE)2 ID NO7:236) derived fromn the coding sequence SE2 ED) NO:235 -shown in Figure 83.

Figures 85A-B show a nucleotide sequence (SE2 ID NW:243) of a native sequence PR293 cMA@, wherein S ID NO:244 is a clone designated herein as "UNQ256- and/or "ENAU3715-1193-.

Figure 86 shows the amino acid sequence (SEQ mD-N 244) derived frcin the coding sequence of SE2 ID N):244 shown in Figures Figures 89A-B show a nucleotide sequence (SEB2 ID NOJ: 248) of a native sequence ROf247 cEN'A, wherein SE2 ID IN):248 is a clone designated herein as "UND221" and/or Irta35673-1201V.

Figure 88 shows the amino acid sequence (SE2 ID D:2 49) derived fromn the coding sequence of SB2 ID M~1:248 shown in Figure 87.

Figure 89 shows a nucleotide sequence (SM) MD NO:253) of a native sequence FIR302 ca4A, whierein SEQ ID ND7:253 is a clone designated herein as IR265" and/or "MUL4370-27".

Figure 90 shows the amino acid sequence (SFQ ID NO~: 254) derived fran the coding sequence of SEQ ID NO:253 shown in Figure 89.

Figure 91 shows a nucleotide sequence (SE2 ED NO7:255) of a :native sequence PR0303 cENA, wherein SEQ ID 1N):255 is a clone designated herein as "UNQ266' and/or "nM@4255-1217'".

Figure 92 shows the aminom acid sequence (SEQ ID N:256) derived :fran the coding sequence of SE2 ID NO:2SS shown in Figure 91.

Figure 93 shows a nucleotide sequence (SE2 ID NJ:257) of a native sequence PR0304 cENA, wherein SE)2 ID ND:257 is a clone designated herein as 'UNQ267' and/or 'tTNA39520-1217-.

Figure 94 shows the amiLim acid sequence (SEQ ID 17:258) derived from the coding sequence of SEQ ID ND2S7 shown in Figure 93.

Figure 95 shows a nucleotide sequence (SEQ ID 7:259) of a native sequence PRO307 cEMA, wherein SE)Q ID NO:259 is a clone designated herein as IUrR2701 and/or "E41225-12171.

Figure 96 shows the aminom acid sequence (SE)2 ID ND:260) derived frat the coding sequence of SE2 ID M7:259 shown in Figure Figure 97 shows a nucleotide sequence (SE2 ID 173:261) of a native sequence PRO343 cMA wherein SEQ ID ND3:261 *is a clone designated herein as "tI1R302, and/or "a1k43318-12l71.

Figure 98 shows the amino acid sequence (SEe ID 17:262) derived fron the coding sequence of SEXQ ID 17:262 shown in Figure 97.

Figure 99 shows a nrucleotide sequence (SEp ID N73:283) of a native sequenoe PR328 wherein SEYQ MD M:283 is a clone designated 60 herein as "UNQ289" and/or "E1N40587-1231'.

Figure 100 shows the amino acid sequence (SEQ ID W:284) derived frcn the coding sequence of SEQ ID NO:283 shown in Figure 99.

Figures 101A-B show a nucleotide sequence (SEQ ID NO:288) of a native sequence PR0335 cI1a, wherein SEQ ID ND:288 is a clone designated herein as "'UN287" and/or "I-41388-1234.

Figure 102 shows the amino acid sequence (SEQ ID E:289) derived frcn the coding sequence of SEQ ID ND:288 shown in Figures 103A-B.

Figure 103 shows a nucleotide sequence (SEQ ID ND:290) of a native sequence PR0331 cla-, wherein SEQ ID ND:290 is a clone designated herein as 'UNQ292" ard/or "URA40981-1234'.

Figure 104 shows the amino acid sequence (SEQ ID N:291) derived frmn the coding sequence of SEQ ID ND:290 shan in Figure 103.

Figure 105A-B show a nucleotide sequence (SEQ ID NO: 293) of a native sequence PR0326 CErA, wherein SEQ ID N1D:293 is a clone designated **herein as "'UNQ287" and/or "Ij37140-1234".

Figure 106 shows the amino acid sequence (SEQ ID ND:294) derived frczn the coding sequence of SEQ ID NO:293 shown in Figures 105A-B.

Figures 107A-B show a nucleotide sequence (SEQ ID N:309) of a 20 native sequence PR0332 c1A, wherein SEQ ID NJ:309 is a clone designated herein as "UNQ293' or "[Er40982-1235'.

Figure 108 shows the amino acid sequence (SEQ ID NO: 310) derived frcm the coding sequence of SEQ ID NO: 309 shown in Figure 107.

Figure 109 shows a nucleotide sequence (SEQ ID 10:314) of a native sequence PR0334 c1lA, wherein SEQ ID ND3:314 is a clone designated herein as "'UNQ295' or "UA41379-1236'.

Figure 110 shows the amino acid sequence (SEQ ID N:315) derived fran the coding sequence of SEQ ID ND:314 shown in Figure 109.

Figure 111 shows a nucleotide sequence (SEQ ID NO:319) of a native sequence PR0346 cMA, wherein SEQ ID I:318 is a clone designated herein as "UNQ305' or "lA44167-1243.

Figure 112 skhows the amino acid sequence (SEQ ID 13:319) derived frcm the coding sequence of SEQ ID ND:318 shown in Figure 111.

Figure 113 shows a nucleotide sequence (SEQ ID ND:323) of a native sequence PRO268 ct1A, wherein SEQ ID ND:323 is a clone designated herein as "N'L235" or 'EW39427-1179.

Figure 114 shows the amino acid sequence (SEQ ID ND:324) derived frman the coding sequence of SEQ ID NO:323 shown in Figure 113.

Figure 115 shows a nucleotide sequence (SEQ ID N3:330) of a 61 native sequence PR330 cEhA, wherein SEQ ID N3:330 is a clone designated herein as "Ug290" or *IEM40603-1232'.

Figure 116 shows the amino acid sequence (SEi ID O: 331) derived fran the coding sequence of SE2 ID ND:330 shown in Figure 115.

Figure 117 shows a nucleotide sequence (SEQ ID ND:337) of a native sequence PRO339 crMA, wherein SED ID 1D:337 is a clone designated herein as "UNQ229 or "CA43466-1225".

Figure 118 shows the amino acid sequence (SEQ ID No: 338) derived fraom the coding sequence of SEQ ID NO: 337 shown in Figure 117.

Figure 119 shows a nucleotide sequence (SED ID EN:339) of a native sequence PRO310 cEA, wherein SP ID ND:339 is a clone designated herein as "UMI273" or "IEA43046-1225".

Figure 120 shows the amino acid sequence (SE ID N0:340) derived frcam the coding sequence of SEQ ID ND: 339 shown in Figure 119.

15 Figure 121 shows a nucleotide sequence (SED ID N:375) of a native sequence PR0244 cENA, wherein SEQ ID ND:375 is a clone designated herein as "UNQ218 or "r'IA35668-11711.

Figure 122 shows the amino acid sequence (SE2 ID O: 376) derived fromn the coding sequence of SE IM NO:375 shown in Figure 121.

IE.. XIPICN OF TME PREER E MBDIMENMS I. Definiticm s The terms "PRO polypeptide" and 'PRO" as used herein and when inediately followed by a numerical designation refer to various polypeptides, wherein the crplete designation PRO/numbrrer) refers to specific polypeptide sequences as described herein. The ternms "PRO/number polypeptide" and "PRO/number" as used herein enccpass native sequence polypeptides and polypeptide variants (which are further defined herein). The PRO polypeptides described herein may be isolated fran a variety of sources, such as fran human tissue types or from another source, or prepared by reccbinant or synthetic methods.

A "native sequence PRO polypeptide' ccprises a polypeptide having the same amino acid sequence as the corresponding PRO polypeptide derived fran nature. Such native sequence PRO polypeptides can be isolated fran nature or can be produced by recorbinant or synthetic neans. The term "native sequence PRO polypeptide" specifically encapasses naturallyoccurring truncated or secreted formns of the specific PRO polypeptide and extracellular dcimnain sequence), naturally-occurring variant fons of alternatively spliced formis) and naturally-occurring 61a allelic variants of the polypeptide. In various enbodinents of the invention, the native sequence PRO211 is a mature or full-length native sequence PRO211 polypeptide ccnprising amino acids 1 to 353 of Figure 2 (SEQ ID N3:2), the native sequence PRO217 is a mature or full-length native sequence PR0217 polypeptide cunprising amino acids 1 to 379 of Figure 4 (SEQ ID ND:4), the native sequence PRO230 is a mature or full-length native sequence PR0230 polypeptide carprising amino acids 1 to 467 of Figure 6 (SEQ ID ND:12), the native sequence PRO232 polypeptide is a mature or fulllength native sequence PRO232 polypeptide carprising amino acids 1 to 114 of Figure 9 (SEQ ID ND:18). the native sequence PRO187 is a mature or fulllength native sequence PR187 carprising amino acids 1 to 205 of Figure 11 S: (SEQD2 ID the native sequence PRO265 polypeptide is a mature or full- .length native sequence PRD265 polypeptide is an extracellular dcnain of the full-length PRO265 protein, wherein the putative transrbrane donain of the full-length PR0265 protein is encoded by nucleotides beginning at nucleotide 1969 of SEQ ID NO:31, the native sequence PR0219 polypeptide is a mature or full-length native sequence PR0219 polypeptide cciprising amino acids 1 to 915 of Figure 15 (SEJ ID NO:34), the native sequence PRO246 is a mature or full-length native sequence PRO246 polypeptide carcnprising amino acids 1 to 390 of Figure 17 (SEe ID NO:39) or the native sequence PRO246 polypeptide is an extracellular donain of the full-length PRO246 protein, wherein the putative transmabrane dcnain of the full-length PR246 protein is encoded by nucleotides beginning at nucleotide 855 as shown in Figure 16, the native sequence PR0228 polypeptide is a mature or full-length native sequence PR0228 polypeptide ccnprising amino acids 1 to 690 of Figure 19 (SEQ ID NO: 49) or the native sequence PRO228 polypeptide is an extracellular darin of the full-length PRO228 protein, the native sequence PR0533 is a mature or full-length native sequence PRO533 carcnprising amino acids 1 to 216 of Figure 22 (SEQ ID MD:59), with or without the N-termnninal signal sequence, and with or without the initiating methionine at position 1, the native sequence PRO245 polypeptide is a mature or full-length native sequence PRO245 polypeptide coyisn amin acdds I to 312 of Figure 24 (SEQ ID N0:64). the native sequence of each PR0220. PR0221 and PR 0227 potypeptides is a mature or full-length native sequence PR0220. PR0221 and PR027 polypeptide comprising amizi acids I through 708 of Figure 26 (SEQ ID N0:69), 1 through 259 of Figure 28 (SEQ ID NO:7 1).

and I through 62D of Figure 30 (SEQ ID NO.73). the native sequence PRO258 polypeptde Jis a mature or ftalength native sequence PR0258 polypepride comprising amino acids I to 398 of Figure 32 (SEQ ID) 10:84) or the native sequence PR0258 polypepude Ls an emtaceliular domain of the full-length PRO258 protein, whe~en the putative transmenibnno domain of the full-Icngth PR0258 protein is encoded by nucteotides beginning at nucleotide 1134 of SEQ ID NO:83. the native sequence PR0266 polypeptide is a mature or full-length native sequence PR0266 polypeptide comprising amino acids I to 696 of Figure 34 (SEQ ID NO:91) or the native sequec PR0266 polypeptide is an exaocchilar domain of die fdl-lerth MR0266 protein, wherein the putative traanamembrane domain of the ful-length, P0266 protein is encod by micloddes beginning at about nucleotide 2009 of SEQ rD NO: 104.

the native sequence PR0269 po4ytide is a mature or full-length native sequence PR0269 polyetide comprising :antinoacids I o40o iue3 SQMN:6 rtentv eunePO6 oyetd sa xrclua domain of the fuall-length PR02,69 protein, wherein the putative tranisrinbrane domain of the full-length PR0269 protein is encoded by ileotides beginning at mucleotide 1502 as shown in Figure 35. the native sequence PRO2W 15 polypeptide is a mature or full-length native seqenc PR0287 polypeptide comprising amino acids 1 to 415 of Figure 38 (SEQ ID NO: 104), the native sequence PR0214 is a mature or full-length native sequence PR0214 comprising aisno acids 1 to 420 of Fig. 40 (SEQ ID NO: 109), the native-sequenice PRO317 is a full-length native-pre-sequence PR0317 comprising amin acids 1 to 366 of Fig. 42 (SEQ WD NO:114) or a mature native-sequenc PR0317 comprising amino acids 19 to 366 of Fig. 42 (SEQ ID NO. 114). the native sequence PRO301 is a mature or fulllength native sequence PRQ301 comprising amino acids I to 299 of Fig. 4 (SEQ ID NO: 119). with or without the N-terminal signal sequence, with or without the initiating methionine at position 1, with or without the potential tuansmembrane domain at position 236 to about 258. and with or without the intracellular domain at about position 259 to 299, the native sequence PR0224 polypeptide is a mainr or full-length native sequence PR0224 polypeptide comprising amino acids I to 282 of Figure 46 (SEQ ID NO:127). the native sequence PROM2 polypeptide is a mature or full-length native sequence PROM2 polypeptide comprising amino acids I to 490 of Figure 48 (SEQ ID NO: 132). t native sequence PR0234 is a mature or full-length native sequence novel lectin comprising amino acids I to 382 of Fig. 50 (SEQ MD NO: 137), the native sequence PR0231 polypeptide is a mature or full-length native soquecec PR0231 polypetd comprising amnino acids 1 to 428 of Figure 52 (SEQ ID NO: 142). the native sequence PRO229 polypieptide is a mature or fuil-laigt native sequence PR0229 polypeptde comprising amino acids 1 to 347 of Figure 54 (SEQ M NO:148), the native sequence PR0239 polypeptide is a mature or full-length native sequence PR0238 pobypeptide comprising amino adids I to 310 of Figure 56 (SEQ ID NMAD5). the native sequence PR0233 polypeptide ita-mreor full-legth nativeseqiEPR0233 polypeptd cromprising amin acids I to 300 of Figure 58 (SEQ ED NO: 159). the native sequence PR0223 polypeptide is a mature or full-length native sequence PR0223 polypeptide comprising amin acids I to 476 of Figure 60 (SEQ ID NO:164), the native sequence PR0235 polypeptide is a mature or full-lngth native seqixc P110235 polypeptide comprising amin acids 1 to 552 of Figure 62 (SEQ ID NO: 170), the native sequence PR0236 polypeptide is a mature or full-length native sequence PR0236 polypeptide comprising amino acids 1 to 636 of Figure 64 (SEQ ID NO:175), dhe native sequence PR0262 63 polypeptide is a mature or full-length native sequence PRO262 polypeptide ccrprising amino acids 1 to 654 of Figure 66 (SEQ ID 0:177) the native sequence PRO239 polypeptide is a mature or full-length native sequence PR0239 polypeptide carprising amino acids 1 to 501 of Figure 68 (SEQ ID N1:185), the native sequence PRO257 polypeptide is a mature or full-length native sequence PRO257 polypeptide ccuprising amino acids 1 .to 607 of Figure 70 (SEQ ID W:190) or the native sequence PRO257 polypeptide is an extracellular dmain of the full-length PRO257 protein, wherein the putative transmxabrane darin of the full-length PRO257 protein is encoded by nucleotides beginning at nucleotide 2668 as shown in Figure 69, the native sequence PR0260 polypeptide is a mature or full-length native sequence PRO260 polypeptide ccaprising amino acids 1 to 467 of Figure 72 (S1EQ ID NO:195) the native sequence PRO263 polypeptide is a mature or full-length native sequence PRO263 polypeptide caprising amino acids 1 to 322 of Figure 74 (SEQ ID rN:201) or the native sequence PRO263 polypeptide *is an extracellular darain of the full-length P1RO263 protein, wherein the putative transmentrane dmarin of the full-length PRO263 protein is encoded by nucleotides beginning at nucleotide 868 of SEQ ID N10:200, the native sequence PRO270 polypeptide is a mature or full-length native sequence PRO270 polypeptide caprising amino acids 1 to 296 of Figure 76 (S3EQ ID ND:207), the native sequence PR0271 polypeptide ccaprising amino acids 1 to 296 of Figure 76 (SEQ ID N.:207), the native sequence PRO271 polypeptide is a mature or full-length native sequence PRO271 xmprising amino acids 1 to 360 of Figure 78 (SEQ ID N10:213), the native sequence PRO272 polypeptide is a mature or full-length native sequence PR0272 polypeptide carprising amino acids 1 to 328 of Figure 80 (SEQ ID NO:221), the native sequence PRO294 polypeptide is a mature or full-length native sequence PRO294 polypeptide ccmprising amino acids 1 to 550 of Figure 82 (SEQ ID 10:227), the native sequence PRO295 polypeptide is a mature or full-length native sequence PRO0295 polypeptide ccurprising amino acids 1 to 350 of Figure 84 (SEQ ID 10:236), the native sequence PRO293 polypeptide is a nmature or full-length native sequence PR0293 polypeptide cacprising amino acids 1 to 713 of Figure 86 (SEQ ID ND:244) or the native sequence PRO293 polypeptide is an extracellular dmain of the full-length PRO293 protein, wherein the putative transmetbrane dmrain of the full-length PM293 protein is encoded by nucleotides beginning at nucleotide 2771 of SED ID ND:243, the native sequence PRO247 polypeptide is a mature or full-length native sequence PRO247 polypeptide ccunprising amino acids 1 to 546 of Figure 88 (SE2 ID 10:249), the native sequence PR0302 polypeptide is a mature or full-length 64 native sequence PR0302 polypeptide ccprising amino acids 1 to 452 of Figure 90 (SED ID ND:254), the native sequence PR0303 polypeptide is a nature or full-length native sequence PR0303 polypeptide cuprising amino acids 1 to 314 of Figure 92 (SEQ ID NO:256), the native sequence PR0304 polypeptide is a mature or full-length native sequence PR0304 polypeptide ccmprising amino acids 1 to 556 of Figure 94. (SEQ ID the native sequence PRO307 polypeptide is a mature or full-length native sequence PRO307 polypeptide caprising amino acids 1 to 383 of Figure 96 (SEQ ID NO:260), the native sequence PRO343 polypeptide is a mature or full-length native sequence PRO343 polypeptide cacprising amino acids 1 to 317 of Figure 98 (SEQ ID NO:262), the native sequence PRO328 polypeptide is a mature or full-length native sequence PR0328 polypeptide ccTprising amino ;acids 1 to 463 of Figure 100 (SEQ ID Nl:284) or the native sequence PRO306 polypeptide is an extracellular darain of the full-length PRO306 protein, 15 wherein the putative extracellular danAin of the full-length PRO306 protein, the native sequence PRO335 polypeptide is a nature or full-length native sequence PR0335 polypeptide caxprising amino acids 1 through 1059 of Figure 102 (SEQ ID ND:289), the native sequence PR0331 polypeptide is a 2 nature or full-length native sequence PR0331 polypeptide conprising amino acids 1 through 640 of Figure 104 (SEp ID NO:291), the native sequence PR0326 polypeptide is a nature or full-length native sequence PR0326 polypeptide crrprising amino acids 1 through 1119 of Figure 106 (SEQ ID N:293). wherein additional enbodinents include wherein the transmenbrane regions are deleted or the peptides are truncated, so as to not include the transarane regions for each of PR0335, PRO331, and PR0326, the native sequence PRO332 is a nature or full-length native sequence PRO332 ccprising amino acids 49 to 642 of Figure 180 (SED ID ND:309), with or without the N-terminal signal sequence, and with or without the initiating methionine at position 1, the native sequence PRO334 polypeptide is a mature or full-length native sequence PR0334 polypeptide carprising amino acids 1 to 509 of Figure 110 (SEQ ID ND:314), the native sequence PRO346 is a mature or full-length native sequence PRO346 carprising amino acids 19 to 339 of Figure 112 (SEQ ID ND:319), with or without the N-terminal signal sequence, with or without the initiating nethionine, with or without the transrabrane dcnain at positions 340 to 360 and with or without the intracellular domain at positions 361 to 450, the native sequence PRO268 polypeptide is a mature or full-length native sequence PRO268 polypeptide ccprising amino acids 1 to 280 of Figure 114 (SEB ID N3:324) or the native sequence PRO268 polypeptide is an extracellular domain of the full-length 64a 9e 0e a. *j a a .tC 0 awa a S. 0 Oa a PR0268 protein, wherein the putative tranamenrbrane domain of the fulllength PR0268 protein is encoded by nucleotides beginning at nucleotide 559 as shown in Figure 113, the native sequence PRO330 polypeptide is a mature or full-length native sequence PR0330 polypeptide comprising amino acids 1 to 533 of Figure 116 (SB ID N0:331), the native sequence PR0339 polypeptide is a mature or full-length native sequence PRO33.9 polypeptide conprising amino acids 1i-to 772 of Figure 118 (SEQ ID NO:338), the native sequence PR0310 polypeptide is a mature or full-length native sequence PRO310 polypeptide ccmprising amino acids 1 to 318 of Figure 120 (SBQ ID N0:340) and the native sequence PR0244 is a mature or full-length native sequence PR0244 comprising andmino acids 1 to 219 of Figure 122 (SEQ ID N0:376), wherein the mature, full-length native sequence PR0244 protein ccnprises a cytoplasmic domain (about amino acid positions 1 to 20), a transwmtnrane domain (about amino acid positions 21 to 46), and an extracellular domain (about amino acid positions 47 to 219). Within the extracellular domain, the C-lectin domain is between about amino acid positions 55 and about amino acid position 206. Native sequence PR0244 as shown in Figure 122 maps to chromosome 12, bands p12-p1 3 "PRO polypeptide variant" means an active PRO polypeptide as defined above or below having at least about 80% amino acid sequence identity with the full-length native sequence PRO polypeptide sequence as disclosed herein. Such PRO polypeptide variants include, for instance, PRO polypeptides wherein one or more amino acid residues are added, or deleted, at the N- or C-terminus of the full-length native amino acid sequence.

25 Ordinarily, a PRO polypeptide variant will have at least about 80% amino acid sequence identity, more preferably at least about 90% amino acid sequence identity, and even more preferably at least about 95% amino acid sequence identity, with the amino acid sequence of the full-length native amino acid sequence as disclosed herein.

30 "PR0317 variants" or PR0317 sequence variants" as defined herein mean biologically active PRO317s as defined below having less than 100% sequence identity with the PRO317 isolated from recomibinant cell culture or from nemammalian fetal kidney tissue having the deduced sequence described in Figure 42. Ordinarily, a biologically active PRO317 variant will have an amino acid sequence having at least about 70% -amino acid sequence identity with the PRO317 of Figure 42, preferably at least about more preferably at least about 80%, still more preferably at least about 85%, even ore preferably at least about 90%, and most preferably at least about 95% 65 70-100%, 75-100%, 80-100%, 85-100%, 90-100%, and 95-100% sequence identity, respectively). These variants include covalently modified polypeptides, as well as PRO317 fragments and glycosylation variants thereof. PRO317 fragments have a consecutive sequence of at least 10, 15, 20, 25, 30, or 40 amino acid residues, preferably about 10-150 residues, that is identical to the sequence of the PR0317 shown in Figure 42.

Other preferred PRO317 fragments include those produced as a result of chemical or enzymatic hydrolysis or digestion of the purified PR0317.

A "chimeric PR0317" is a polypeptide comprising fulllength PR0317 or one or more fragments thereof fused or bonded to a second protein or one or more fragments thereof. The chimera will typically share at least one biological property in 15 common with PRO317. The second protein will typically by a cytokine, growth factor, or hormone such as a neurotrophic or angiogenic factor such as GDNF or VEGF, or another member of the TGF-superfamily such as EBAF-1. Another exemplary preferred PRO317 chimera is a "domain chimera' that consists of the N- 20 terminal residues substituted with one or more, but not all, of the residues of the human EBAF-1. In this embodiment, the PR0317 chimera would have individual or blocks of residues from the human EBAF-1 sequence added or substituted into the PRO317 sequence. For example, one or more of those segments of EBAF-1 that are not homologous could be substituted into the corresponding segments of PRO317. It is contemplated that this "PRO317-EBAF-1 domain chimera" will have an agonist biological activity.

"Percent amino acid sequence identity" with respect to the PRO polypeptide sequences identified herein is defined as the percentage of amino acid residues in the PRO sequence which are identical with the amino acid residues in a candidate polypeptide sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software 65a such as BLAST, ALIGN or Megalign (DNASTAR) software. The preferred software alignment program is BLAST. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared.

"Percent nucleic acid sequence identity* with respect to PRO-encoding nucleic acid sequences identified herein is defined as the percentage of nucleotides in the PRO sequence of interest which are identical with the nucleotides in a candidate nucleic acid sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum *'percent sequence identity. Alignment for purposes of i. determining percent nucleic acid sequence identity can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software such as BLAST, ALIGN or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared.

20 "Isolated", when used to describe the various polypeptides disclosed herein, means polypeptide that has been identified and separated and/or recovered from a component of its natural environment. Contaminant components of its natural environment are materials that would typically interfere with diagnostic or therapeutic uses for the polypeptide, and may include enzymes, hormones, and other proteinaceous or nonproteinaceous solutes. In preferred embodiments, the polypeptide will be purified to a degree sufficient to obtain at least 15 residues of N-terminal of internal amino acid scquecec by use of a spinnig cup sequemlr. or a2) to homogeneity by SDS-PAGE under nonreducing or reducing conditions using Coomassie blue or. preferably. silver stain. Isolated polypeptide includes polypqnide In situ within recombiznant cells, sinc at least oe component of the PRO polypeptide natural environment will not be present. Ordinarily. however. isolated polypepsde will be prepared by at least one puification step.

As 'isolated" PRO polypeptide nucleic acid is a nucleic acid molecule that is identified and separated frm at least one contateirum nucleic acid molecule with which it is ordinarily associated in the natural 3-urc of dhe PRO polypept~le nucleic acid. An isolated PRO polypeptide nucleic acid molecule is other than in the form or setting in which it is found in nature. Isolated PRO polypeptide nucleic acid molecules therefore are distinguished from the specific PRO polypeptide nucleic acid molecule as it exists in nanual cells. However, an isolated PRO po lypcptide inidac acid molecule incles PRO polypepide nucleic acid molecules contained in cells that ordinarily express the PRO polypeptide where, for example, the nucleic acid molecule is in a chrmosomal location different from that of natural cells.

'Southern analysis" or "Southern blotting* is -a method by which tie presence of DNA sequeces in a restriction endazaiceas digest of DNA or a DNA-comalning composition is confirmed by hybridization to a known, labeled oligonucleotide or DNA fragment. Southern analysis typically involves electrophorctic: separation of DNA digests on agarose gels.. denaturation of dhe DNA after electrophoretic separation, and transfer of the DNA to nitrocellulose. nylon, or anothcr suitable mim rawi stqlou far analysis with a radiolabeled. biotinylated, or enzymelabeled probe as described in sections 937-9.52 of Sambrook ei Molecular Cloning: A Laboratory Manual (New :York: Cold Spring Harbor Laboratory Press. 1989).

*'Northern analysis'0 or "Northern blotting* is a method used to identify RNA sequences that hybridize to a keown probe such as an oligoncloodde, DNA fragmnt. cDNA or fragment thereof, or RNA *fragment. The probe is labeled with a radioisotope such as 3P. or by biotinytation, or with an enzyme. The RNA to be analyzed is usually electrophoretically separated on an agarose or polyacrylainide gel, ansferred to nitrocellulose. nylon, or other suitable membrane, and hybridized with die probe, using standard techniques well known in die art such as those described in sections 7.39-7.52 of Sambrook et al.. supm'.

The term "control sequences" refers to DNA sequences necessary for the expression of an operably linked *..coding sequence in a partilar hos organism. The control sequences that are suitable for prokaryotes. for example, include a promoter, optionally an operator sequence, and a ribosomne binding site. Eukaryotic cells are known to utilize promoters. polyatlenylation signals, and enhancers.

Nucleic adid is "operably linked' when it is placed into a functional relationship with another nucleic acid sequence. For example, DNA for a presequence or secretory leader is operably linked to DNA for a polypeptide if It is expressed as a preprotein that participates in the secretion of the polypeptide; a promoter or enhancer is operably linked to a coding sequ ee if it affects the transcription of the sequence: or a ribosome binding site is operably linked to a coding sequence if it is positioned so as to facilitate translation. Generally, 'operably linked" means that the DNA sequences being linked are contiguous, arnd, in the case of a secretory leader, contiguous and in reading phase. However, enhancers do not have to be contiguous. L inking is accomplished by ligation at convenient restriction sites. If such sites do not exist, the synthetic oligonucleotide adaptors or linkers are used in accordance with conventional practice.

The term 'antibody" is used in the broadest sense arid specifically covers single an-PRO polypepdde monoclonal antibodies (ncluding against, antagonist. and oeniurlzig antibodies) and anti-PRO polypeptide antibody compositious with polyepitopic specificity. The term 'mnowclonal antibody* as used herein refers to an antibody obtained from a population of substanatially homogeneous antibodies. the individual antibodies comprising the population are identical except for possible naturally-occuning mutations that may be present in mnr amounts.

Active' or "activity" for the purposes herein refers to formis) of PRO polypeptide which ftsain the biologic and/or immunologic activities of the specific native or naturally-occurring PRO polypeptide. The activity of a PR0332 polypeptide preferably involves the regulation of exuceUular matrix. cartilage, or bonc fuanction.

OPR0317-asociated disorder' refers to a pathologcal condition or disease wherein PR0317 is over- or unlerexpressed. Such disorders include diseases of the female genital tract or of the endometrium of a mnammal.

including hyperplia. eadometritis. endonietriosis, wherein the patient is at risk for infertility due to endometrial factor. endometrioma. and endometia cancer, especially those diseases involving abnormal bleeding such as a gynecological diseate. They also include diseases involving angiogcEesis, whercin the angiogenesis results in a :4 patbolocal condition, such as cancer involving solid oinors (the therapy for the disorder would result in decreased vasculadization and a decline in growth and metastasis of a variety of tumors). Alternatively, the angiogenesis may be beneficial, such as for ischemia, especially coronary ischemia. Hcec, these disorders include tse found in patits whose hearts ame functioning but who have a blocked blood supply due to atherosclerotic coronary artery disease and those with a functioning but underperfused heurt, including patients with coronary arterial disease who are not optimal candidates for angioplasty and coronary artery by-pass surgery. The disorders also include diseases .*:involving the kidney or originating from the kidney tissue, such as polycystic kidney discase and chronic and acute renal failuire.

*Treatment" or *treating" refers to both therapeutic treatment and prophylactic or preventative measures.

ixThoe in need of treatment include those already with the disorder as well as those prone to have the disorder of those -in which the disorder is to be prevented.

'Mammal" for purposes of treatment refers; to any animal classified as a mammal, including humans, domestic and farm animals, and zoo, sports, or pet animals, such as shecp. dogs, horses, cats, cows, and the like.

Preferaby. the mammal2 herein is 2 human.

44*4'Carriers' as used herei n include pharmaceutically. acceptable carriers, excipients, or stabilizers which are nontoxic to the cell or mammal being exposed thereto at the dosages and concentrations employed. Often the physiologically acceptable carrier is an aqueous PH buffered solution. Examples of physiologically acceptable carniers include buffers; such as phosphate, citrate, anid other organic acids. antioxidants including ascorbic acid; low molecular weight (less than about 10 residues) polypeptide;, proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycinec. gltamin asparagine, arginine or lysirE: monosaccharides, disaccharides, and other carbohydrates including glucose, mannoe, or dextrins; chelating agents such as EDTA; sugar alcohols such as manhol or sorbitol; salt-forming counterions such as sodium; and/or nonionic surfactants such as TWEEN-W, polyethylene glycol (PEG). and PLURONICST".

The term "agonist* is used to refer to peptide and uion-peptide analogs of the native PRO polypeptides (where native PRO polypeptide refers to pro-PRO polypeptide, pre-PRO polypeptide, prepro-PRO polypeptide, or imature PRO polypeptide) of the present invention and to antibodies specifically binding such native PRO polypeptdes,. provided diat they retain at least one biological activity of a native PRO polypeptidc. Preferably. the agnit of dft present invention retain the qualitative binding recognition properties and receptor activation properties of the native PRO polypeptide.

The term 'antagonist' is used to refer to a molecule inhibiting a biological activity of a native PRO polypeptide of the presen invention wherein native PRO polypeptide refers to pro-PRO poitpeptide. pre-PRO polypeptide. prepro-PRO polypeptide. or mature PRO polypeptde. Preferably, die antgonists herein inhibit the binding of a native PRO polypeptide of the present invention. Preferred antagonists essentially completely block the binding of a native PRO317 polypeptide to a PR0317 polypcptide receptor to which it otherwise binds. Such receptors may inchide the Type I and Type U1. and possibly Type MI rceptors identified for the TOF- superfamily.

KolodzieJczyk and Hal, supra. A PRO polypeptide "antagonist* is a molecule which prevents, or interferes with, a PRO antagonist effector function a molecule which prevents or interferes with binding and/or activation of a PRO polypeptde receptor by PRO polypptde). Such molecules can be screened for their ability to competitively inhibit PRO polypeptide receptor activation by monitoring binding of native PRO polypeptide in dhe presence and absence of die test antagonist molecule, for example. Examples of PR0317 polypeptide antagonists include neutralizing antibodies against F-2. An antagonist of the invention also encompasses an mntisense polyntucleotide against the PRO polypeptide gene, which anisense polynucleotide blocks transcription or translation of the PRO polypeptide gene, thereby inhibiting its expression and biological activity.

*Stringent condition" means employing low ionic strength and high temperature for washing. for example. 0.015 sodium chloridei0.0015 M sodium citrateV.1 sodium dodecyl sulfate at 500C. or employing 20 during hybridization a denaturing agent. such as formamide, for example. 50% (volvol) formamride with 0. 1% bovine serum albuminl0.1I FicoUIO.1 polyvinylpyrrolidione/5O nM sodium phosphate buffer at pH 6.5 with 750 tnM :sodium chloride, 75 mnM sodium citrate at 42'C. Another example is use of 50% formamide, 5 x SSC (0.75 M NaCI. 0.075 M sodium citrate), 50 mM sodium phosphate (pH 0.1 sodium pyrophosphate, 5 x Denhardt's soluion, sonicated salmon sperm DNA (50 jug/ml). 0. 1% SDS, and 10% dextran sulfate at 42*C. with washes at 42*Cin 0.2 xSSC and 0.1% SDS. Yet another example is hybridization using a buffer of 10% dextran sulfate, 2 x SSC (sodium chlouide/sodium canla) and 50% formamide at 55 followed by a high-stringency wash consisting of0.l x SSC containing EDTA at 556C.

Moderately stringent conditions" are described in Samabrook et al.. sqr,. and include the use of a washing solution and hybridization conditions temperanire ionic strength. and %SDS) less stringast than described above. An example of moderately stringent conditions is a condition such as overnight incubation at 37*C in a soluton comprising: 20% fomide, 5 x SSC (150 mM NaCI, 15 mM trisodium citrate), 50 mM sodium phosphate (pH 5 x Denhardt's solution, 10% dextran sulfate, and 20 mg/niL dentured sheared salmon sperm DNA, followed by washing the filters in I x SSC at about 37-50*C. The skilled artisan will recognize how to adjust the temperature, ionic strength, etc., as necessary to accommodate factors such as probe length and die like.

It. Comoodftions i Methods of the invcntion* 1. FalIlengpth PR0211 and PR27Plotde The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred toin the present application as PRO211I and PRO2 17. In particular. Applicants have identifie-d and isolated cDNA encodig PRO211 and PR0217 polypeptides. as disclosed in further detail in the Examples below. Using BLAST (FastA format) sequence alignmnent computer programs. Applicants found that eDNA s-oquenme encoding fuI-length native sequence PRO211I and PR0217 have homologies to known proteins having EGF-like domins.

Specifically, te cDMA sequne DNA32292-1 131 (Figure 1. SEQ H) NO:l) has 36% idenuify' and a Blast score of 209 with PAC6 RAT and 31 identify and a Blast score of 206 with Fibulin-I, isoform c precursor. The cDNA sequence DNA33094-1131 (Figure 3. SEQ ID NO.3) has 36% identity and a Blast score of 336 with easter newt Imin. and 37% identity and a Blast score of 331 with to-ni tcoascipa-X precursor. Accordingly, itis presently V'0 believed that PRO211 arnd PR0217 polypeptides disclosed In the present application are newly identified members of the EGF-like family and possesses properties typical of the EGF-Ukc protein family.

2. Full-leagpth PRO230 Fobvuntdes The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides 0 referred to in the present application as PROM3. In particular, Applicants have identified and isolated cDNA encoding a PR0230 polypqpifr. as disclosed in further detail in the Examples below. Using known programs such as BLAST and FastA sequence alignment computer program. Applicants found that a eDNA sequence encoding fulllength native sequence PR0230 has 48% amino acid identity with the rabbit tubuloizuerstitiai nephritis antigen precursor. Accordingly. it is presently believed that PR0230 potypeptide disclosed in the present application is a newly identified member of the tubulointerstitial nephritis antigen family and possesses the ability to be recognized :by human autoantibodies in certain forms of tubulointerstitial nephritis.

3. Full-leneth PRO232 Polvoentides 0Go:25 The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PROM3. In particular, Applicants have identified and'isolated, eDNA 000*00encoding a PR0232 polypeptide, as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignment computer programs, Applicants found that a portion of the ful-lngth native sequence PR0232 (shown in F7igure 9 and SEQ ID NO:lIS) has 35% sequence identity with a stem cell surface antigen from Gallus gallus.- Accordingly, it is presently believed that the PR0232 polypeptide disclosed in tie present application may be a newly identified stem cell antigen.

4. Full-lenp'th PRO197 Pohopetides The present invention provides newly identified and isolated nueleotdec sequences encoding polypeptides referred to in the present application as PROIS7. In particular. Applicants have identified and isolated cDNA encoding a PRO187 polypeptide. as disclosed in further detail in the Examnples below. Using BLAST and FastA sequence ahgtunent comptster programs. Applicant fixind d=a a full-length native sequence PRO 187 (shown in Figure IS) has 74% amrino acdd sequence identity and BLAST score of 310 with various androgen-nduced growth factors and FGF-8. Accordingly, it it presently believed tdn PRO0137 potypptde disclosed in the present application is a newly identified member of the FGP-8 protcin family and may possess identif activity or property typical of the FGF-8-like protein family.

S. FuII-lenath PR0265 Pobypntldes The present invention provides newly iden~tified and isolated nucleotide sequences encoding polypctidcs referred to in the present application as PRO26S. In particular, Applicants have identified and isolated cDNA encoding a PR0265 polypeptide. as disclosed in fitrther detail in the Examples below. Using programs such as BLAST and FastA sequence alignment computer programs. Applicants found! that various portions of the PR0265 polypeptide have significant homology with the flbronmdulin protein and fibromodulin precursor protein. Applicants *have also found that the DNA encoding the PR0265 polypeptide has significant homology with platelet glycoprotein V. a meber of the leudzie rich related protein family involved in din and woinal repair. Accordingly, it Is presently believed that PRO26S polypetide disclosed in the present application is a newly identified member of the leucinc rich repeat family andi possesses protein protein binding capabilities, as well as be involved in skin and wound repair as typical of this family.

6. Full-tengh PRO219 Potyptides refrredThe present invention provides newly identified and isolated nuecotide sequences encoding polypeptides reere to in the present application as PR0219. In particular. Applicants have identified and isolated cDNA *coo 20 encoding a PR0219 polypeptide. as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignment computer programs. Applicants found that various portions of the PR0219 polypeptide have signifieant homology with the mouse and human niatrilin-2 precursor polypeptdecs. Accordingly, it is presently believed that PR0219 polypeptide disclosed in the present application is related to the matrilin-2 precursor polypeptide.

7. Full-lenoth PRO246 Polvpptides *The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PR0246. In particular. Applicants have identified and isolated cDNA eneoding a PR0246 polypeptide. as disclosed in further detail in the Examples below. Using BLAST and FastA seqtlutme alignment computer program. Applicants found that a portion of the PR0246 polypeptide has significant homology with the btman cell swfac protein HCAR. Accordingly, it is presently believed that PR0246 polypeptide disclosed in the present application may be a newly identifled mnezzbrane-bound virus receptor or tmor cell-specific antigen.

8. Full-length PR228 P-dyoepides Tbe present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PR0228. In partilar. Applicants have identified and isolated cDNA encoding a PR0228 polyrpeptide. as disclosed in futher detail in the Examples below. Using BLAST and FastA sequence alignment computer programs, Applicant found that various portion of the PR0228 polypeptidc have significant homology with the EMR1 protein. Applicants have also found that die DNA encoding the PR0228 polypeptide has significant homology with Iatrophilin. snacrophage-restricted cell surface glycoprotein. B0457.1l and leucocyte anogen CD97 precursor. Accordingly, it is presently believed that PR0228 polypeptide disclosed in the present applicatin is a newly identified member of the seven tranismenmbrane suerfamily.-and possesses characteristics and functional properties typical of this ftmily. In particular, it is believed that PR0228 is a new member of the subgroup within this family to which CD97 and EMRI belong.

9. Full-leni'th PRO533 Polvoptides The present invention provides newly identified and isolated micleodde sequences encoding polypeptides referred to in the present application as PROM3. In particular, Applicants have identified and isolated cDNA aending a PROS33 polypeptide. as disclosed in further detail in the Examples below. Using BLAST-2 and FastA V sequee alinctit computer programs. Applicants found that a full-length native setluence PR0533 (shown in Figure *22 and SEQ ID NO-.59) has a Blast soore of 509 and 53 amio acid sequence identity with fibroblast growth factor (FOP). Accordingly. It is presently believed diat PRO533 disclosed in the present application is a newly identified member of the fibroblast growth factor family and may possess activity typical of such polypeptides.

Full-length PRO245 PoyneotIde The present invention provides newly identified and isolated ncleotide sequences encoding polypeptides referred to in the present application as PROM24. In particular. Applicants have identified and isolated cDNA encoding a PR0245 polypeptide. as disclosed in further detail in the Examples below. Using BLAST and FastA :sequence aligiment computer programs, Applicants found that a portion of the amino acid sequence of the PR0245 polypeptide has 60% amino acid identity with the humnan c-myb protein. Accordingly, it is presently believed that the PR0245 polypeptide disclosed in the pree, application my be a newly identified member of the transmembrane protein tyrosine kinas family.

11. Full-length PRO=,0 PR221 and PRO227 PoLyUenIddes The present invention provides newly identified and isolated ucleotide sequences encoding polypeptides referred to in the present application as PR0220. PR0221 and PR0227. In particular. Applicants have identified and isolated cDNAs encoding a PR0220, PR0221 and PPR0227 polypeptdec. respectively, as disclosed in further deail in tdi Examples below. Using BLAST and FastA sequence alignment computer programs, PRO220 has armno acid idenity witlh the amino acid seqenc of a leucir rich protein wherein the identity is 87 PR0220 additionally has amino acdd identity with the neuronal latrine rich protein wherein the identity is 55 The neuronal leucine rich protein ft further described in Taguchi, et al.. Mol. Bain.Res., 35:31-40 (1996).

PR0221 has am~io acid identity with the SIT protein precursor, wherein different portions of these two proteins have die respective percent identities of 39%. 38%. 34%, 31 and PRO22 has amino acid identity with the amn acid sequene of platlet glycoprotein V precursor. The sam reults were obtained for hurmn glycoproccin V. Different portions of these two proteins show the following percent identities of 30%, 28%. 28%. 31%. 35%, 39% and 27%.

Accordingly, it is presently believed that PR0220. PR0221 and PRO2W polypeptdecs disclosed in the present application are newly Identified members of the leucine rich repeat protein superfamily and that each possesses protean-protein binding capab~ilities typical of t leucine: rich repeat protein superfaily. It is,.also believed that they have capabilities simnila to those of SLIT, the leucine rich repeat protein and human glycoprotein V.

12. Full4eng'th PRO259 Polyptides The present invention provides newly identified and isolated nucleoidde sequences encoding polypeptides referred to in the present application as PR0258. In partilar. Applicants have identified and isolated cDNA encoding a PR0259 polypeptide. as disclosed in futher detail in the Examples below. Using BLAST and FastA seqiuence aliginment computer programs. Applicants found that various portions of the PRO258 polypeptide have signffcan homology with the CRTAM and poliovirus receptors. Accordingly, it is presently believed that PRO258 polpeptidc disclosed in the present application is a newly identified snunber of the Ig superfamily and possesses virus receptor capabilities or regulates imimune (unction as typical of this family.

13. Full4ength PR0266 PolXMentides The present invention provides newly identified and isolated macleode sequences encoding polypeptides referred to in the present application as PR0266. In particular. Applicants have identified and isolated cDNA encoding a PR0266 polypeptide. as disclosed in furlier detail in the Examples below. Using BLAST and FastA sequence alignment compuiter programs. Applicants found that various portions of the PR0266 polypeptide have significant homology with the SUJT protein from Drosophilia. Accordingly, it is presently believed that PR0266 polypeptide disclosed in the present application is a newly identified member of the leucine rich repeat family and possesses ligand-ligand biniding activity and neuronal developmen typical of this family. SU1T has been shown to be useful in the study and tretment of Atzheinier's disease, supra, and thus, PR0266 may have involvement in the study and cure: of this disease.

14. Full-length P!R0269 Polyneotide9 Tbe present invention provides newly identified and isolated niacleodde; sequnce encoding polypeptides referred to in the present application as PR0269. In particular, Applicants have identified and isolated eDNA encoding a PRO269 polypeptide, as disclosed in further detail in the Examples below. Using BLAST. FastA and sequence alignment computer programs, Applicants found that the amino acid sequence encoded by macleotides 314 to 1783 of the ful-length native seqene PR0269 (shown in Figur 35 and SEQ MD NO:95) has significant homology to human urinary thrombomodulin and various trmbomochalin analogues respectively, to which it was aligned.

Acodigy it is prescotty believed that PR0269 polypcptidc disclosed in the present application Is a newly identified member of the thromnbomodulin family.

IS. Fyll~en~h PRO287 Folvueptides The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PROW9. In particular, Applicants have identified and isolated cDNA encoding a PROW8 polypptde, as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignment computer programs, Applicants found that various portions of the PRO2W polypeptide have signficant homology with the type I procollagen C-protemnase enhanicr protein precursor and type I 'procoulagen Cprowinase enhancer prowin. Accordingly, it is presently believed that PRO28 polypcptidc disclosed in te present application is a newly identified member of the C-protcinase enhancer protein family.

16. ftl14length PR0214 Polvmentldes The present invention provides newly identified and isolated nwuceotide sequences encoding polypeptides referred to in the present application as PRO2 14. In particular, Applicants have identified and isolated cDNA encoding a PR0214 polypetdec, as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignment computer programs, Applicants found that a full-length native sequenc PR0214 polypeptide (shown in Figure 40 and SEQ ID NO: 109) has 49% amino acid sequence identity with HIT protein, a known member of the EGF-family. The comparison resulted in a BLAST score of(920, with 150 mnatching mitcleotides. Accordingly, it is presently believed that the PR0214 polypetde disclosed in the present application is a newly idied member of the family corrarising EGF domains and may possess activities or properties typical of the EGF-domain containing family.

17. Fnll-lenglh PR0317 Paolv ides The present invention provides newly identified and isolated micleotide sequences encoding polypeptides referred to in the present application as PR0317. In particular. cDNA encoding a PR0317 polypeptide has been idcritified and isolated, as disclosed in further detail in the Examples below. Using BLAST" and FastK" sequence alignmnt computer programs, it was found tha a full-length native-sequence PR0317 (shown in Figure 42 and SEQ ID NO:114) has 92% amrino acid sequence identity with EEAF-I. Further, it is closely aligned with many other members of the TGF- superfamily.

Accordingly, it is presently believed that PR0317 disclosed in the present application is a newly identified member of the TGP- superfamily and IMay Possess properties that arc therapeutically useful in conditions of uterine bleeding, etm. Hence, PR0317 may be useful in diagnosing or treating abnormal bleeding involved in gynecological diseases, for example, to avoid or lessen the need for a hysterectomy. PR0317 may also be useful as an agent that affects angiogenesis in general, to PR0317 may be useful in anti-tumor indications, or conversely, in treating coronary ischemic conditions.

Library sources reveal cdat EMSswed to obtain the consensus DNA for generating PR0317 primers and probes were found in normal tissues (uterus, prostate, colon, and pancreas), in several tmors (colon, brain (twice), 35 pancreas, and mnullerian cell), and in a heart with isdtmria. PR0317 has shown up in several tissues as well, but it does look to have a greater concentration in uterus. Hence, PR0317 may have a broader use by the body than EAF-I. It is contemplated that, at least for some indications. PR0317 my have opposite effects from EAF-I.

18. Full-length PRO301 Polvuenti&e The present hmventon provides newly identified and isolated rnvcicotide sequences encoding polypeptides referred to in dhe present application as PRO301. in Particular, Applicants have identified and isolated cDNA encoding a PR0301 polypcptide. as disclosed in further detail in die Examples below. Using BLAST and FastA aeqnm aligaiwint compter program. Applicants found doa a ful-length native sequence PRO301 (shown in Figure 44 and SEQ ID NO:119) has a Blast score of 246 correspondting to 30% amin acid sequence idaty with human A33 antigen precursor. Accordingly. it is-presently believed that PRO301 disclosed in the present application is a newly identi em tnber of lt A33 antigen protein hardly and may be expressed in human ncoplastic diseases such as colorectal cancer.

19. Dill-length PRO224 Polvuentde The present invetion provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PR0224. In particular, Applicants have Identified and isolated cDNA encoding a PRC)224 polypeptie. as disclosed in further detail in the Examples below. Using known programs such as BLAST and FastA sequence alignment computer programs. Applicants found that full-length native PR0224 (Figure 46, SEQ MD NO:127) has amino add identity with apolipoprotein E receptor 2906 from hoino sapiens. The alignmets of different portions of these two polypeptides show amino acid identities of 37%, 36%. 30%. 44%. 44 and 28% respectively. Full-length native PRO224 (Figure 46. SEQ ID NO:127) also has amino acid identity with very low-density lipoprotein receptor precursor from gall. The alignments of different portions of these two polypeptides show amino acid identities of 38%, 37%. 42%. 33%, and 37% respectively. Additionally, full-length native PR0224 (Figure 46. SEQ ID NO: 127) has amino acid identity with the chicken oocyte receptor P95 from Gallus gallus. The alignmens of different portions of these two polypeptides show amino acid identities of 38%6, 42%, 33%. and 37% respectively. Moreover, full-length native: PR0224 (Figure 46, SEQ ID NO:127) has am acid identity with very low density lipoprotein receptor short form precursor from humans. The alignments of different portions of these two polypeptides show amino acid Identities of 32%, 38%, 34%, 45%, and 31%, respectively. Accordingly, it is presently believed that PR0224 polypeptide disclosed in the present application is a newly Ideniffied erof the low density lipoprotein receptor family and possesses the structural characteristics required to have the fianctional ability to recognize and endocytose low density lipoproteins typical of the low density lipoprotein receptor faily. CMh alignments described above used the following scoring parameters: T=7. S S2=36, Matrix: BLOSUM62.) Full-length PRO222 PolMICVtIdes The present invention provides newly identified and isolated nuwceotide sequences encoding polypeptides referred to in the present application as PR0222. In particular. Applicants have identified and isolated cDNA encoding a PR0222 polypeptide, as disclosed in further deta i n the Examples below. Using BLAST and FastA sequence alignment computer programs. Applicants foundi that a sequence encoding fulecngth native sequence PR0222 (shown in Figure 48 and SEQ ID NO: 132) has 25-26 amino acid identity with mouse complement factor h prcaursor. has 27-299,% amino ad Idenuity with completacnt receptor, has 25-47 amino acid identity with mouse cosrilemn*t Ob receptor tWp 2 long form precursor, has 40% amino acid identity with human hypothetical protein kiaaO247. Accordingly, it is presently believed that PR0222 polypeptide disclosed in the present application is a newly identified member of the comiplemnt receptor family and possesses activity typical of the complement receptor family.

21. Ful.letveth PRO234 Polvuentide The present invention provides newly identified and isolated nucleotde sequences encoding polypeptides referred to in the present application as PR0234. In particular. Applicants have identified and isolated cDNA enooding a PR0234 polyptde. as d&closed in fiute detail in the Examples below. Using BLAST (FastA-format) sequence alignment computer programs, Applicants kuxnd that a cDNA sequence encoding full-length native sequence PRO234 has 31 identity and Blast score of 134 with E-sclectin precursor. Accordingly, it is presently believed that the PRO234 polypeptides disclosed in the present application are newly identified members of the lectin/selectin .4 famnily and possess activity typical of the lectin/selectin family.

22. Full-length PR10231 Polvnepaidc 15 The present invention provides newly identified and isolated nucleoide sequences encoding polypeptides referred to in the present application as PR0231. In particular, Applicants have identified and isolated cDNA encoding a PR0231 polypeptide, as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignment computer programs. Applicants found that the full-length native sequence PRCY231I polypeptiode *:(shown in FgureS52adxlSEQID NO:l142) has 30 and 31 amino acid identity with human and rat prostaic acid phosphatase precursor proteins, respectively. Accordingly, it is presently believed that the PR023 1 polypeptide disclosed in the present application may be a newly identified member of the acid phosphatase protein family.

23. FutI-Ienffth PR10229 Polypenides The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PR0229. In particular. Applicants have identified and isolated cDNA encoding a PR0229 polypeptide, as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignmnt computer programs. Applicants found that various portions of the PR0229 polypeptide have significant homology with antigen wcl.1, M130 antigen. T cell Surface glycoprotein CD6 and CD6. It also is related to Spalpha. Accrdingly. it is presently believed that PR0229 polypeptidc disclosed in the present application is a newly identified umember of the family containing scavenger receptor homology, a sequence motif found in a number of proteins involved in immune function and thus possesses immune function and /or segments which resist degradation., typical of this family.

24. Full-length PR10239 Polvnentide The present invention provides newly identified and isolated nucleotdec sequences encoding polypeptides referred to in the present application as PR0238. In particular, Applicants have identified and isolated cDNA encoding a PR0238 polypeptide. as disclosed in fiurher detail in the Examples below. Using BLAST and FastA sequence alignment computer programs. Applicants found that various portions of the PR0238 polypeptide have significant homology with reductases, including oxidoreductase and fatty acyl-CoA reductase. Accordingly, it is presently believed that PR0238 polypeptide disclosed in the present application is a newly Identified member of the reductase family and possesses reducing activity qTical of the reductase: family.

25. Full-length PRO233 Potvnentldes The present invention provides newly identified and isolated nucicoode sequences encoding polypptdes referred to in the present application as PR0233. In particular. Applicants have identified and isolated cDNA encoding a PR0233 polypeptide, as disclosed in further detail in dhe Examples below. Using BLAST and FastA sequence-aligrunent computer programs, Applicants found that various portions of the PR0233 polyctide have significant homology with the reductase protein. Applicants have also found that the DNA encoding the PR0233 .polypeptide has significant homology with proteins from Caenorhabditis elegans. Accordingly, It is presently believed dhat PR0233 polyetide disclosed in the present application is a newly identified member of te reductase family and possesses die ability to effect fte redox state of the cell typical of the reductase 6anuly.

15 26. Full-lenrth PR0223 Poynsptides The present invention provides newly identified and isolated nucleotide sequences encoding polypcptides; referred to in the present application as PR0223. In particular. Applicants have identified and isolated cDNA encoding-a PR0223 polypeptide. as disclosed in further detail in the Examples below. Using BLAST and FastA sequence.aliginent comnputer program, Applicants foundl dwa the PR0223 polypeptide has significant homology with various serine carboxypeptidase polypeptides. Accordingly. It Is presently believed tha PR0223 polypeptidc disclosed in the present application is a newly identified seine carboxypeptidase.

27. Full-length PR235 PoluentiduK The present invention provides newly identified and isolated nuclcotide sequences encoding polypetides; referred to in the present application as PROM3. In particular, Applicants have identified and isolated cDNA encoding a PR0235 polypeptide. as disclosed in further detail in die Examples below. Using BLAST and FastA sequence alignment computer program, Applicants found that various portions of the PR0235 polypeptide have significant homology with the various plexin proteins. Accordingly, it is presently believed that PR0235 polypeptide disclosed in the present application is a newly Idenitified member of the plexin family and Possesses cell adhesion properties typical of the plexin family.

28. Full-teogth PRO736 and PRO262 PolXVoenides The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referre to in the present application as PR0236 and PR0262. In particular, Applicants have identified and isolated cDNA encoding PR0236 and PR0262 polypeptides, as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignmnrt computer programs, Applicants found that various portions of the PR0236 and PR0262 polypeptides have significant homology with various P-galactosldase and P-galactosidise precursor PolypcpddCS. Accordingly. ft is Presently believed that the PP.0236 and PR0262 polypeptides disclosed in the present application are newly identified P-galactosidase homologs.

29. FulI-lengill PRO239 Polvueptides The present invention provides newly identified and isolated nucleotide Sequences encoding polypeprades referred to in the present application as PR0239. In paricular. Applicants have identified andrisolazed cDNA encoding a PR.0239 polypeptide, as disclosed in further detail in the Examples below. Using BLAST and PastA sequene alignment computer programs. Applicants found that various portions of the PR.0239 polypeptide have significant homology with densin proteins. Accordingly, it is presently believed that PR.0239 polypeptide disclosed in the present applicationi is a newly identified member of the derisin family and possesses cell adhesion and the ability to effect synaptic processes as is typical of the densin family.

Full-length PROM5 Polvpetldes The present invention provides newly identified and isolated mucleotide sequences encoding polypeptides referred to in the present application as PROMS. In particular, Applicants havw identified ant! isolated cDNA encoding a PR0257 polypeptide, as disclosed in further detal in dhe Examples below. Using BLAST and FastA sequece alignment computer programs, Applicants found that various portions of the PRO2S7 polypeptide have significant homology with the ebnerin precursor and ebnerin protein. Accordingly, it is presently believed that PRO257 polypeptide disclosed in the present application is a newly identified protein member which is related to the eboerin protein.

31. Full~len,!th PR10260 Povunti&gm Thie present invention provides newly identified and isolated nucleotide sequences encoding polypeptides *44*referred to in the present application as PR0260. In particular, Applicants have identified and isolated eDNA encoding a PR0260 polypeptide. as disclosed in further detail in the Examples below. Using programs such as BLAST and FastA sequence alignment computer programs. Applicants found that various portions of dhe PR0260 polypeptde have significant homology with the alpha-l-fucosidase prectusr. Accordingly, it is presently believed that PR0260 polypeptide disclosed in the present application is a newly identified member of the fucosidase famnily andi possesses enzymatic activity related tofucose residues typical of the fucosidase family.

32. Full-lcngth P-RO263 Polnptides The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PRO026. In particular, Applicants have identified and isolated cDNA encoding a PR.0263 polypeptide, as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignment computer programs. Applicants found that variouis portions of die PR0263 polypeptide have significant homology with the CD44 antigen and related proteins. Accordingly, it is presently believed that PR0263 polypeptdec disclosed in the present application is a newly identified member of the CD44 antigen family and possesses at least one of the properties associated with these antigens. cancer and HIV marker, cell-cell or cellmatrix iteractions, regulating ceLl boaffc, lymph node homing. transmission of growth signals, and presentation of' chemoines and growth facors to traveling cells.

33. FulL~cngth PRO270 Pollplptides 31hc present invention provides newly identified and isolated nucleodde sequences encoding polypeptides3 referred to in the present application as PROM7. In particular. Applicants have identified and-isolated cDNA encoding a PR0270 polypeptide. as disclosed in further detail in dhe Examples below. Using BLAST. FastA and sequence aligrmci compme programs. Applicants found dian that various portions of dhe PR0270 polypeptide have significant homology with various thioredoxin proteins. Accordingly, it is presently believed that PR0270 polypeptide disclosed in the present application is a newly identified member of the thioredoxin family and possesses the ability to effect reduction-oxidation (redox) state typical of the thioredoxin family.

:34. Fulleangth PR271 Polvnentldes The present invention provides newly identified and isolated rtucleotide sequences encoding polypeptides referred to in the present application as PR0271. In particular. Applicants have identified and isolated cDNA encoding a PR0271 polypeptide. as disclosed in fuirther dtai in the Examples below. Using BLAST and FastA sequience aligrnat couputer programs, Applicant fcund diat the PR0271 polypeptide has significant homology with various link proteins aund precurors thereof. Accordingly, it is presently believed that PR0271 polypeptide disclosed in the present application is a newly identified link protein hosnolog.

3S. Full-length PR 0272 Polynentides The present invention provides newly identified and isolated ucleotide sequences encoding polypeptides referred to i the present application as PROM'7. In particular, Applicants have identified and isolated cDNA encoding a PR0272 polypqxide. as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignment computer programs. Applicants found that various portions of the PR0272 polypeptide have significant homology with the human reticulocalbin protein and its prccursors. Applicants have also found that the DNA encoding the PRO272 polypeptide has significant homology with the mouse reticulocalbin precursor protein.

Accordigl, it is presently believed that PRO272 polypeptde disclosed in the present application is a newly identified member of die reticulocalbin family and possesses the ability to bind calcium typical of the redilocalbin family.

36. Full-length PRO294 Povnetides The present invention provides newly identified ancl isolated nucleotide sequences encoding polypeptides referred to in the present application as PROM9. In particular. Applicants have identified and isolated cDNA encoding a PR0294 polypcptidc. as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignment computer programs. Applicants found that various portions of the PR0294 polypeptide have significan homology with dhe various portions of a murber of collagen proteins. Accordingly, tt is presently believed that PR0294 palypeptdec disclosed in the present application is a newly identified member of the collagen family.

37. FulI= R29S P b~de The present invention provides newly identified and isolated uiucleotide sequences encoding polypeptides referred to in the present application as PR0295. In particular. Applicants have identified and isolated cDNA encoding a PR0295 polypeptide. as disclosed in further derail an the Examples below. Using BLAST and FastA sequence ;lignmnt computer programs, Applicants found that various portions of the PR0295 polypeptdec have significant homology with irtegrin prmeuans. Accordingly. it is presently believed that PRCV295 polypptde disclosed in the present application is a newly identified member of the integrin family and possesses cell adhesion typical of the integrin family.

38. Full-leng'th PRO293 Potirenides Th mp nt invention provides newly identified and isolated nucleotdde sequences encoding polypeptides referred to in the present application as PR0293. In particular. Applicants have identified and isolated cDNA encoding a PR0293 polypepide. as disclosed in further detail in t Examples below. Using BLAST and FastA sequence alignment computer programs. Applicants found that portions of the PR0293 polypeptide have significant homology with the neuronai leucine rich repeat proteins 1 and 2, (NLRR-l and NLRR-2), particularly NLRR-2.

Accordingly. it is presently believed that PR0293 polypde disclosed in die present application is a newly identified r of the miuronal leucine rich repeat protein family and possesses ligand-ligand binding activity typical of the NRLL protein family.

39. Full-lencth PRO247 Polvotdes The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PROW4. In particular. Applicants have identified and isolated cDNA :encoding a PR0247 polypeptide. as disclosed in further detail in the Examples below. Using BLAST and FastA ***.*sequece alignment computer programs, Applicants found that various portions of the PRO24 polypeptide have significan homology with dens in. Applicants have also found that the DNA encoding dhe PR0247 polypeptide has significant homology with a number of other proteins. including KIAA023 I. Accordingly, it is presendy believed don PR0247 polypeptide disclosed in dhe present application is a newly identified member of the leucine rich repeat family and possesses ligand binding abilities typical of this family.

Fullength PROMZ PR0303. PR0304. PRO307 and PRO343 Polvucptides The present invention provides newly identified and isolated nucleotide sequences encodintg polypeptides refnul o in the present application as PR0302, PR0303. PR0304. PR0307 and PROW4. In particular, Applicants have identified and isolated cDNA encoding PR0302. PROM0, PR0304. PRO307 amd PR0343 polypeptides. as disclosed in fikhr derail in the Examples below. Using BLAST and FastA sequence alignment computer programs, Applicants found that various portions of the PR0302. PR0303. PR0304. PR0307 and PR0343 polypeptides have significant homology with various protease protens Accordingly, it is presently believed that the PR0302. PR0303, PR0304. PR0307 and PR0343 polypeptides disclosed in the present application are newly identified proteasc proteins.

80 41. Full-langth PR0328 Polypeptidea The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PR0328. In particular, Applicants have identified and isolated cWNA encoding a PRO328 polypeptide, as disclosed in further detail in the Exanples below. Using BLAST and FastA sequence alignment carputer programs, Applicants found that various portions of the PR0328 polypeptide have significant homology with the human glioblastamna protein ("GLIP") Further, Applicants found that various portions of the PR0328 polypeptide have significant horology with the cysteine rich secretory protein ('CRISP") as identified by BLAST hanology ([ECRISP3_., S68683, and CRS3_HUMAN] Accordingly, it is presently believed that PRO328 polypeptide disclosed in the present application is a newly identified marber of the GLIP or CRISP families and possesses transcriptional regulatory activity 15 typical of the GLIP or CRISP families.

42. Full-Length PB0335, PR0331 and PRO326 Polypeptides The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present 20 application as PR0335, PRO331 or PRO326. In particular, Applicants have identified and isolated cENA encoding a PRO335, PRO331 or PR0326 polypeptide, as disclosed in further detail in the Examples below. Using BLAST and FastA sequence alignment ccmputer programs, Applicants found that various portions of the PRO335, PR0331 or PRO326 polypeptide have significant hamology with LIG-1, ALS and in the case of PRO331, additionally, decorin. Accordingly, it is presently believed that the PRO335, PRO331 and PRO326 polypeptides disclosed in the present application are newly identified mnbers of the leucine rich repeat superfamily, and particularly, are related to LIG-1 and possess the biological functions of this family as discussed and referenced herein.

43. Full-Length PRD332 Polypeptides The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PR0332. In particular, Applicants have identified and isolated cEM encoding PRO332 polypeptides, as disclosed in further detail in the Exanples below. Using BLAST and FastA sequence alignment computer programs, Applicants found that a full-length native sequence PRO332 (shown in Figure 108 and SBE ID N3:309) has about 30-40% amino acid sequence 80a identity with a series of knon proteoglycari sequences, including, for exanpe, f ibramdulin and f ibrcxmdulin. precursor seuences of various species (FC)D BOVIN, EMDJ-IC, FMOD-WA, FMMDJ)EXSE, FMVDjLtPAN, PJU36773), ostecitodulin seqjuences (ABOO01lll, AB007848_1), decorin sequences (CFL13141_1, 0W033941, P R42266, P R42267, P R42260, PR89439), keratan sulfate proteoglycans (B1U48360_1, AF022890..L), corneal proteoglycan IAF22256l1), and bone/cartilage proteoglycans and proteoglycane precuLrsors (PGSLWhINE, P=S_MDJSE, PGS2j1U-VN).

Accordingly, it is presently believed that PIR)332 disclosed in the present application is a new proteoglycan- type molecule, and nray play a role in regulating extracellular matrix, cartilage, and/or bane function.

44. FLA14-ength PRO334 Polvptlde The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PR0334. In particular. Applicarits have ientified and isolated cDNA encoding a PR0334 polypeptide. as disclosed in further deta i n dhe Examples below. Using BLAST and FastA sequence alignment computer programs. Applicants found that various portions of the PR0334 polypepdde have significant homology with fibulin and fibrillin. Accordingly, it is presently believed that PRO334 polypcptide d&closed in the present application is a ncwly identified member of the epidermal growth factor family and possesses properties and activities typical of this family.

FutIllength PRO346 Pol=Cpncntd 10 The present invention provides newly identified and isolated nucleoride sequences encoding polypeptides referred to in the present application as PR0346. In particular. Applicants have identified and isolated cDNA encoding a PR0346 polypeptide. as disclosed in further detail in die Exampqles; below. Using BLAST and FastA sequence aligzmint computer programs, Applicants found that a full-ength native sequence PR0346 (shown in Figure 112 and SEQ ID NO:320) has 28 amino acid sequence identity with carcinuecmbmyonic antigen. Accordingly. it is presently believed that PR0346 disclosed in the presenit application is a newly identified member of the carcinoembryortic protein family and may be expressed in association with neoplastic: tissue disorders.

4. Full-lenath PR0269 Polypsgtides The present invention provides newly identified and isolated nucleotide sequences enicoding polypeptides referred to in the present application as PR0268. In particular. Applicants have identified and isolated cDNA encoding a PR0268 polypeptide. as disclosed in further detail in the Examples below. Using BLAST and FastA sequence augment conmputer programs, Applicants found that portions of the PR0268 polypeptide have significant homology with die various protein disulfide isomerase proteinis. Accordingly. it is presently believed that PRO268 polypeptide disclosed in the present application is a homolog of the protein disulfide isomerase: p5 protein.

47. EulI-jength PRO330 Polynentides The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides referred to in the present application as PR0330. In particular. Applicants have identified and isolated cDNA encoding a PR0330 polypeptide, as disclosed in further detail in the Examples below. Using BLAST and PastA sequence alignment computer programs. Applicants found that various portions of the PR0330 polypeptide have significant homology with die nmumne prolyl 4-hydroxylase alpha-Il subunit protein. Accordingly, it is presently believed that PR0330 polyetdec disclosed in the present application is a novel prolyl 4-hydroxytase subunit palypeptide.

48. Full-l.ngth PR0339 and PRO310 Polyuientides The present invention provides newly identified and isolated nucleotide sequences encoding polypeptides; referred to in the present application as PR0339 and PRO3 10. In particular, Applicants have identified and isolated 82 cNA encoding a PR0339 polypeptide, as disclosed in further detail in the Exanples below. Applicants have also identified and isolated cDA encoding a PRO310 polypeptide, as disclosed in further detail in the Exanples below.

Using BLAST and FastA sequence alignment cmputer programs, Applicants found that various portions of the PR0339 and PR0310 polypeptides have significant hanology with small secreted proteins from C. elegans and are distantly related to fringe. PR0339 also shows horology to collagen-like polymers. Sequences which were used to identify PRO310, designated herein as ENA40533 and ENA42267, also show homology to proteins fran C. elegans.

Accordingly, it is presently believed that the PRD339 and PRO310 polypeptides disclosed in the present application are newly identified nmaber of the family of proteins involved in development, and which may have regulatory abilities similar to the capability of fringe to regulate serrate.

15 49. Full-Length PRO244 Polypeptides The present invention provides newly identified and isolated nucleotide sequences encoding C-type lectins referred to in the present application as PRO244. In particular, applicants have identified and 20 isolated cEA encoding PRO244 polypeptides, as disclosed in further detail in the Exanples below. Using BLAST and FastA sequence aligrment canputer programs, Applicants found that a full-length native sequence PRO244 (shown in Figure 122 and SED ID ND:376) has 43% amino acid sequence identity with the hepatic lectin gallus gallus (LECH-CHICK), and 42% amino acid sequence identity with an HIV gpl20 binding C-type lectin (A46274). Accordingly, it is presently believed that PRO244 disclosed in the present application is a newly identified member of the C-lectin superfamily and may play a role in immune function, apcptosis, or in the pathogenesis of atherosclerosis. In addition, PRO244 may be useful in identifying tumor-associated epitopes.

PRO Polypeptide Variants In addition to the full-length native sequence PRO polypeptides described herein, it is contemplated that PRO polypeptide variants can be prepared. PRO polypeptide variants can be prepared by introducing appropriate nucleotide changes into the PRO polypeptide NA, or by synthesis of the desired PRO polypeptide. Those skilled in the art will appreciate that amino acid changes may alter post-translational processes of the PRO polypeptide, such as changing the nuber or position of glycosylation sites or altering the nmmbrane anchoring characteristics.

82a Variations in the native full-length sequence PRO polypeptides or in various domains of the PRO polypeptides described herein, can be made, for exanple, using any of the techniques and guidelines for conservative and non-conservative nutations set forth, for instance, in U.S. Patent No. 5,364,934. Variations mnay be a substitution, deletion or insertion of one or nore codons encoding the PRO polypeptide that results in a change in the amino acid sequence of the PRO polypeptide as ccnpared with the native sequence PRO polypeptide. Optionally the variation is by substitution of at least one amino acid residue with any other amino acid in one or nore of the domains of the PRO polypeptide. Guidance in determining which amino acid residue may be inserted, substituted or deleted without adversely affecting the desired activity may be found by ccparing the sequence of the PRO polypeptide with that of humologous known protein molecules and minimizing the number of amino acid sequence changes uade in regions of high harology. Amino acid substitutions can be the result of replacing one amino acid with another amino acid having similar structural and/or chemical properties, such as the replacement of a leucine with a serine, i.e., e consevative amino acid replacements. Insertions or deletions may optionally be in the range of 1 to 5 amino acids.

The variation allowed may be determined by systematically mnaking insertions, deletions or substitions of amino acids in the sequence and testing the resuting variants for activity in the in ierro assay described in the Examples below.

TIe variations can be made using methods known in the art such as oligotuiclonde-nediated (site-directed) inutagenesis, alanine scanning. and PCR mutageneis. Site-directed nuagensis (Carter et al., Nacl.Acids Res., 13-4331 (1986); Zollcr et al.. Nut AisE~ Re -W:6487 (19M7). cassette mutageniesis [Wells et al., 34:315 (1985)1. restriction selection nutagencsas [Wells et al.. Phios TranS- R. Soc, London SerA, Wf:415 (1986)) or other known techntiques can be performed on the cloned DNA to produce the desired PRO polypeptide variant DNA.

Scanning amino acid analysis can also be employed to identify one or more amino acids along a contiguous sequenc. Among the preferred scanning amino acids are relatively small, neutral amino acids. Such amino acids V. inchud alain, glycioc, serne, and cysteine. Manine is typically a preferred scanning amino add among this group :becasse it eliminates the side-chain beyond the beta-carbon and is less likely to alter the main-cliain conformation of tie variant. Alanine is also typically preferred because it is die most common amino acid. Further, it is frequently found in both buried and exposed positions [Creighton, Ibc Protins. Freeman Co., Chothia, L.

M& HioL. 150:1 (1976)). if alanine substitution does not yield adequate amounts of variant, an isoteric amino acid can be used.

51. Modifications of-PRO Polvwcptides Covalent modifications of PRO polypeptides are included within the scope of this invention. One typc of 20 covalent modification includes reacting targeted amino acid residues of the PRO polypeptide with an organtic derivatirzing agent that is capable of reacting with selected side chains or the N- or C- terminal residues of the PRO a: polypeptide. Derivatiztion with bifunctional agents is useful, for instance, for crosslinking a PRO polypeptide to a water-hroluble support mat&i or suface for use in the method for purifing anti-PRO polypeptide antibodies, and vice-versa. Commonly used crosslinking agents include, I ,l-bi(diazoacetyl)-2-phenylethane glutaraldehyde, N-hydroxysuccitnmide esters, for example, esters with 4-azidosalicylic acid, homobifunctional imdoesters, including ~disuccinimidyl ester such as 3,3'-ditmlobis(succinimidlpropionate), bifunctional maleimides such as bis-Nmaleimido-l .8-octane and agenits such as methyl-3-[(p-azidophcnylrdthiojpropioiniidatr.

Other modifications include deaniidation of glutaminyl and asparaginyl residues to the corresponding ghzzamyl id! aspartyl residues, respectively. hydroxyLation of prolmne and lysine, phosphorylation of hydroxyl groups of seryl or threonyl residues, methylation of the a-amino grouips of lysine, arginine, and histidine side chains IT.E_ Cegton. Pfai Structure and Molecular Proneniet W.H. Freeman Co., San Francisco, pp. 79-86 (1983)), acetylation of the N..terminal amine, and arnidation of any C-terminal carboxyl group.

Another type of covalent modification of the PRO polypeptides included within the scope of this invention comprises altering the native glycosylation pattern of the polypieptide. 'Altering the native glycosylation panecrnis intended for purposes herein to mean deleting one or more carbohydrate moieties found in a native sequence PRO polypeptide, and/or adding one or more glycosylation sites that are not present in the native sequence PRO polypeptide.

Addition of glyoxylation sites to the PRO polypeptide may be accomplished by altering the amino acid sequence. The alteration may be made, for example. by the addition of, or substitution by, one or morm scrine or threnninc residues to the native sequence PRO polypeptide (for 0-linked glycosylation sites). The PRO polypcptidc amino acid sequence may optionally be altered through changes at the DNA level. particularly by mutating the DNA cecoding the PRO polypeptide at preselected bases such that codons are generated that will translate into the desired amino adds. t Another means of increasing the number of carbohydrate moieties on the PRO polypeptide polypeptide is by chemical or enzymatic coupling of glycosides to the polypetide. Such methods are described in the art, in WO 87/05330 published 11 September 1987, and in Aptin and Wriston. CRC Crit. Rev. Bioebem-, pp. 259-306 (1981).

10 Removal of carbohydrate moieties present on the PRO polypeptide may be accomplished chemically or enzymatically or by mutational substitution of codons encodin~g for amino acid residues that serve as- targets for glycosylatin Chemical deglycosyladon modmlques are known in the art and described, for instantce, by Hakiniuddin.

et al.. Arch. Reochem- fli2Vhvs W:52 (1987) and by Edge et al.. Ana iohem. 111:131 (1981). Enzymatic cleavage of carbohydrate moieties on polypeptides can be achieved by the use of a variety of cado- and exoglycosidases as described by Thotakura et al., Meth. Enzymol. fl1:350 (1987).

Another type of covalent modification of PRO polypeptides of the invention comprises linidng the PRO polypeptide to one of a variety of nonproteinaceous polymers. polyethylene glycol. polypropylene glycol, or polyoxyalkylenes. in the manner set forth in U.S. Patent Nos. 4,640,835; 4.496,689; 4,301,144; 4,670,417; 4,791,192 or 4.179.337.

The PRO polypeptides of the present invention tnay also be modified in a way to form a chiimeric molecule comprising a PRO polypeptide fused to another, heterologous polypeptide or amino acid sequence. In one embodiment, such a chimeric molecule comprises-& M~son of the PRO polypeptide with a tag polypeptide which provides an epitope to which an anti-tag antibody can selectively bind. The epitope tag is gienerally placed al the amino- or carboxyl- cerminus of the PRO polypeptide. The presence of such epitope-segged forms of the PRO polypeptde can be detected using an antibody against the tag polypeptide. Also, provision of the epitope tag enables the PRO polypeptide to be readily purificid by affinity puification using an anti-tag antibody or another type of affinity matrix that binds to the epitope tag. In an alternative enbodiment, the chim ric molecule may comprise a fusion of the PRO polypeptide with an imrmmoglobulin or a particular region of an immunoglobulin. For a bivalent form of the chimeric molecule, such a fusion could be to the Fc region of an IgG molecule.

Vaious tag polypeptides and their respective antibodies are well known in the art. Examples include polyhistidint (poly-his) or poly-bistidine-gtycine (poly-his-gly) tags; the flu HA tag polypeptide and its antibody (Field et al., MoI. Cell. fliol.. 1-:2159-2165 (1988); t c-rnyc tag and the 8F9, 3C7. 61110. G4, B7 and 9E10 antibodies thereto [Evan et al., Molecular and Cellular Biology, 1:3610-3616 (1985)]; and t Herpes; Simplex virus glycoprotein D (gD) tag and its antibody (Paborsky et al., ,rti EnidM (6):547-553 (1990)). Other tag polypeptides include the Flag-peptder (Hopp et al., Bia~no~zlfgy, 6l:1204-1210 (1988)]; the KT3 epitope peptide (Martin et al., Sre= M:192-194 (1992)1; an ri-tubulin epitope peptdec (Skinnr et al.. 1LBi2ol Qm. 2:15163- 1516 6 (1991); and the 77gene 10 protein pepide tag [Lutz-Fmyyemnuthct al., Proc. Nad.- Acad. Sci. USA, 12:6393- 6397 (1990)).

52. Modification of PRO317 Amino acid seqc variants of PRO317? are prepared by Introducing appropriate zuicleotide changes into t PROW1. DNA, or by i vitro synthesis of die desired PR0317 polypeptide. Such variants include, for examplc.

deletions from, or inserfions or subs titutions of. residues within the amino acid sequene shown forbhuman PRO317 In Figure 42. Any combination of deletion, insertion, and substitution is made to arrive at the final constrict, provied that the final consmics possesses the desired characteristics. The amino acid changes also may alter posttranslational processes of the PRO3 17. such as changing the mnber or position of glycosylation sites. Moreover, like most mammalian gemes PRO317 is presumably encoded by nulti-exon genes. Alternative suRNA constructs which may be attributed to different mRNA splicing events following transcription. and which share large regions of identity with the cDNAs claimed herein, are considered to be within the scope of the present invention.

For t design of amin acid sequence variants of PROM 1. the location of the mutation site and the nature of the mutation wll depend on the PR0317 characteristic(s) to be mnodified. For example, candidate PR0317 antagonists or agonists will be initially selected by locating sites that are identical or highly conserved among PROW,7 EBIAF-l. LEFTy, and other members of the TGF- superfamily. The sites for mutation can be modified individually or in series, by substituting first with conservative amino acid choices and then with more radical selections dependting upon the results achieved. deleting the target residue, or inserting residue of thc same or a different class adjacent to the located site, or combinations of options 1-3.

A useful method for identification of certain residues or regions of the PR0317 polypepude that are preferred locations for mutagenesis is called "alanizn scanning niutagenesis,* as described by Cunningham and Wells, Scine 2_4: 1081-10115 (1989). Here, a residue or group of target residues are identified charged residues such as arg, asp, his, lys, and glu) and replaced by a rieutral or negatively charged amino acid (most preferably alanine or polyalanine) to affect the interaction of the amino acids with the sturrouinding aqueous environment in or outside the cell. Those domains demonstrauing fumctianial sensitivity to the substitutions then are refined by introducinglartd=or other varants at or for thesites of substitution. Thus, while the site forintroducing an amino acid sequece variation is predetermiztd. the nature of the fmuuation per se need not be predetermined. For example.

*000 to optimize the performance of a mutation at a given site. alanine scanning or random mutagenesis is conducted at the target codon or region and t PR0317 variants produced are screened for t optimal combination of desired activity.

There are two principal variables in the construction of amino acid sequence variants: the location of the mutation site and the nature of the mutation. These; are variants front t Figure 42 sequence, and may represent naturally occurrintg alleles (whiich will not require manipulation of the PRO3 17 DNA) or predetermined mutant forms made by tautating t DNA, either to arrive at an allele or a variant not found in nature. In general. the location and nature of the mutation chosen will depend upon the PR0317 characteristic to be modified.

Amino acid sequerice delein generally range from about I to 30 residues, more preferably about I to residues, and typically are contiguous. Contiguous deletions ordinarily are made in even naumbers of residues, but single or odd numbers of deletions are within the scope hereof. Deletions mnay be introduced into regions of low homology among PRO317. EBAF-1. and other members of the TGF- superfamily which share the most sequence identity to the human PRO317 amino acid sequence to modify the activity of PR0317. Deletions from PR0317 in aras of subsntial homology with one of the receptor binding sites of other members of the TGF- superfamily will be more likely to modify the biological activity of PRO317 more significantly. The number of consecutive deletions will be selected so as to preserve the tertiary structure of PR0317 in the affected domain, beta-pleated sheet or alpha helix.

Amino acid sequence insertins include amino- and/or carboxyl-terminal fusions ranging in length from one residue to polypeptidcs containing a hundred or more residues, as well as intrasequence insertions of single or multiple amino acid residues. Intrasequencc insertions insertions within the mature PR0317 sequence) may range generally from about I to 10 residues, more preferably 1 to 5. most preferably 1 to 3. Insertions are preferably made in even nmbers of residues but this is not required. Examples of terminal insertions include mature PRO317 with an N-terminal methionyl residue, an artifact of the direct production of mature PR0317 in recombinant cell c**ure, and fusion of a heterologous N-terminal signal sequence to the N-terminus of the mature PRO317 molecule Sto a facilitate te secrtion of mature PRO317 from recombinant hosts. Such signal sequences may be obtained from, and thus homologous to, the intended host cell species, but also may be from other members of the TGFsuperfamily. Suitable sequences include STII or Ipp for E. coli, alpha factor for yeast, and viral signals such as herpes gD or the native EBAF-I sequence for mammalian cells.

Other insertional variants of the PRO317 molecule include the fusion to the N- or C-terminus of PRO317 of immunogenic polypeptides, bacterial polypeptides such as beta-lactamase or an enzyme encoded by the E.

<coi' p lous. or yeast protein, and C-terminal fusions with proteins having a long half-life such as immunoglobulin constant regions (or other immunoglobulin regions), albumin, or ferritin, as described in WO 89/02922 published 6 April 1989.

A third group of variants are amino acid substitution variants. These variants have at least one amino acid residue in the PR0317 molecule removed and a different residue inserted in its place. The sites of greatest interest for substitutional mutagenesis include sites identified as the active site(s) of PRO317 and sites where the amino acids found in the known analogues are substantially different in terms of side-chain bulk. charge, or hydrophobicity, but where there is also a high degree of sequence identity at the selected site within various animal PR0317 species, or where the amino acids found in known members of the TGF- superfamily and novel PR0317 are substantially different in terms of side-chain bulk, charge, or hydrophobicity, but where there also is a high degree of sequence identity at the selected site within various animal analogues of such members among all the animal EBAF-1 molecules). This analysis will highlight residues that may be involved in the modulation of endometrial tissue or angiogenesis, and therefore, variations at these sites may affect such activities.

Other sites of interest are those in which particular residues of the PRO317 obtained from various species arc identical among all animal species of PRO317 and other members of the TGF- superfamily, this degree of conservation suggesting importance in achieving biological activity common to these cytokines. These sites, especially those falling within a sequence of at least three other identically conserved sites, are substituted in a relatively conservative manner. Such conservative substitutions are shown in Table I under the heading of preferred substitutions. If such substitutions result in a change in biological activity, then more substantial changes, denominated exemplary substitutions in Table 1. or as further described below in reference to amino acid classes.

are introduced and die products screened.

origina Ala (A) Arg (R) Asan(N) Asp (D) Cys (C) Gin (Q) Glu (E) 15 Gly (G) His le (1) Exernplary vaR; ICU; H~e lys; gin; asn gin, his; lys; arg ser asn asp pro; ala asn; gin; lys; arg leu; val; met;, ala.. phe; norleucine norleucine; le; val; met; ala; phe arg;, gIn; asn Ieu; phe; le leu; val; ile; ala; tyr ala ftr setyr; pbre trp. plie; thr; ser lic; 3wu; met. phe: ala; norleucine Preferred LCU (L) Lys (K) Miet (M) Phe (F) Pro (P) Ser (S) MW (1) TrP(WM Tyr M Val MV Substantial rnodifications in function or inununalogical identity of the PR0317 are accomplished by selectinig substitutions that differ significantly in their effect on maintaining the structure of the polypeptidi backbone in the area of the substitution, for example. as a sheet or helical conformation, the charge or hydrophobicity of the miolecule at the target site, or the bulk of the side chain. Naturally occurring residues are divided into groups based on common side-chain properties: hydrophobic: norleucine. met. ala. val. let', le; neutral hydrophilic: cys. scr. dir; acidic: asp. glu; basic: asn, gin, his, lys. arg; residues that influence chain orientation: gly, pro; and aromatic: tup, tyr, phe.

Non-conservative substitutions will entail exchanging a member of one of these classes for another class.

Such substituted residues also may be introduced into the conservative substitution sites or, more preferably, into the remaining (non-conserved) sites.

In ame embodiment -of the invention, it is desirable to inactivate one or more protease cleavage sites that are present in the molecule. These sites are ikdeified by inspection of the encoded amnino acid sequence. in the case of Irypsin for an arginyl or lysinyl residue. When protease cleavage sites are idetified. they ame rendered inactive to proteolytic cleavage by substituting the targeted residue with another residue, preferably a baskc residue such as giuzarnr or a hydropbilic residue such as serine; by deletin t residue; or by inertng a prolyl residue immediately after the residue.

1la another embodiment. any methionyl residues other than the starting methionyl residue of the signal sequence, or any residue located within about three residues or C-terminal to each such methonyl residue. is substituted by another residue (preferably in accord with Table 1) or deleted. Alternatively, about 1-3 residues are inserted adjacent to such sites.

Any eysteine residues not involved in maintaining the proper confornation of PR0317 also may be substituted, generally with serine. to inprove the oxidative stability of the molecule and prevent aberra crosslinking.

Nucleic acid molecules encoding amino acid sequence variants of PR0317 are prepared by a variety of mctheds known in t art fTse methods include, but ame not limited to. isolation from a natural source (in the case of naturally occurring amino acid sequence variants) or preparation by oligomzcleotde-mediated (or site-directed) musagcnesis. PCR mutagenesis, and cassette mutagenesis of an earlier prepared variant or a non-variant version of PROM 1.

Oligonucleotide-inediated nintageneis is a preferred method for preparing substitution, deletion, and *insetion variants of PR0317 DNA. This technique is well known in the art as described by Adelman et aL. DNA, 2: 183 (1983). Briefly, PRO317 DNA is altered by hybridizing an oligonrucleotide encoding the desired mutation to a DNA template, where the template is the single-stranded form of a plasmid or bacteriophage containing the unaltered or native DNA sequence of PROM1. After hybridization, a DNA polyntrase is wsed to synthtesize an entire second complementary strand of the template that will thus incorporate the oligonucleotide primer, and will code for the selected alteration in the PR0317 DNA.

:Generally, oligonucleotides; of at least 25 nucleohides in length are used. An optimal oligonucleodde will have 12 to 15 ntuceotides that art completely comaplenetary, to the template on either side of the nucleotide(s) coding for the mutation. This ensures that the oligonucleotide will hybridize properly to the single-stranded DNA template nolecule. lT oligomucleotides are readily synthesized using techniques known in the art such as that described by Crea etrat.. Pzoc- Natl. Acad- Sci. USA. 71: 5765 (1978).

The DNA template can be. generated by those vectors that are either derived from bacteriophage MU3 vectors (the commerciall available M 13mp 18 and M l3mpl19 vectors are Suitable), or those vectors that contain a singie-stranded phage origin of replication as described by Viem et al. Meh Ezo-,JL3: 3 (1987). Thus, the DNA that is to be mutated may be inserted into one of thes vectors to generate single-stranded template. Production of the single-stranded template is described in Sections 4.21-4.41 of Samnbrook et supra: Alternatively, single-stranded DNA template may he generated by denaturing double-stranded plasmnid (or other) DNA using standard techniques.

For alteration of the native DNA sequence (to generate amino acid sequnce variants, for example), the oligonucleocide is hybridized to the single-stranded template under suitable bybridization conditions. A DNA polymerizing enzyme, usually the Klenow fragment of DNA polymnerase 1, is then added to synthesize the complementary swrd of the template using the oligomiceotide as a primer for synthesis. A beteroduplex molecule is thu ibrmed such that ami stad of DNA encodes the mutated form of PROM 1. and the other strandf (the original template) encodes the native. unaltered sequence of PROMl. This; heteroduiplex molecule is then transformed into a suitable host cell. usually a prokaryote such as E. coli JMIOI. After the cells amc grown, they arm plated onto agaiw plates and screened using te ofigoicldeotide primer tadiolabeled with nP to identify the bacterial colonies that ontande imitated DNA. 7Te mutated region is then removed and placed in an appropriate vector for protein production, generally an expression vector of the type typically employed (or transformation of anjppropriate host.

The method described immediately above may be modified such that a bomoduplex molecule is created wherein both strands of the pLastaid contain the imutation(s). The modifications are as follows: The single-stranded oligonucleotide is annealed to the single-stranded template as described above. A mixture of three dexyriboucleoddes, deoxyriboadenosine (dATP), deoxyriboguanosine (dGTP), and deoxyribothymidine (dMV), is combined with a modified thdeoxrocytosie called dCTP-(aS) (which can be obtained from the Amershamn Corporation). This mixture is added to the template-oigonucleoti complex. Upon addition of DNA polymerase to dhis mixture, a strand of DNA identical to the template except for the mutated bases is generated. In addition, this new standl of DNA will contain dCr-(aS) insead of dCTF, which serves to protect it firm restriction endonucleas digestion.

After the template staAnd of the double-sixanded heteroduplex is nicked with an appropriate restriction etnyme. the template strand can be ifigested with ExoMf nuclease or anothe appropriate nuclease past the region that contains the site(s) to be mutagenized. The reaction is then stopped to leave a molecule that is only partially single-satraded. A complete double-stranded DNA homoduplex is then formed using DNA polymcrase in the pitsence of all four deoxyribonacleotide triphosphates. AT?, and DNA ligase. This homoduplex molecule can then be transformed into a suitable host cell such as E. coil JMI10l. as described above.

DNA encoding PR0317 mxutants with more than one amino acid to be substituted may be generated in one of several ways. If t amino acids are located close together in the polypeptdec chain, they may be mutated simuzltaneously wing one oligonucleotide that codes for all of the desired amino acid substitutions. If. however, the amino acids are located sonme distance fromi each other (separated by more than about ten amino acids). it is more difficult ID generate a single oLigonucleotide that encodes all of the desired changes. Instead, one of two alternative methods may be employed.

In the first method, a separate oligonucleotidc is generated for each amino acid to be substituted. The oligonucleoddes are then annealed to the single-stranded template DNA simultaneously, and the second strand of DNA that is synthesized from the template will encode all of the desired amino acid substitutions.

The alternative method involves two or more rounds of niutagenesis to produce the desired mutant. The first round is as described for die single mi-ts: wiki-Myp DNA is used for the template, an oligonuclcotide encoding the first desired amino acid substitution(s) is annealed to this template, and the heteroduplex DNA molecule is then generated. The second round of mutageneuis; utilizes the mutated DNA produced in the first round of mutagenesis as the temlate. Thus, this template already contains one or more mutations. The oligonucleotdec encoding the additional desired amino acid substitution(s) is the annealed to this template, and the resulting stratnd of DNA now encodes mutations from both the first and second rounds of mutagenesas. This resultant DNA can be used as a template in a third round of niutagenesis, -and so on.

PCR wnugeraiss is also suitable for making amino acid variants of PROM 1. While the following discusion refers to DNA. it is understood that the techaique also finds application with RNA. The PCR- technique generally refers to the following procedure (see Erlich. PCR Tehnolog, (Stockton Press. NY. 1989). the chapter by R. Higuchi. p. 61-70): When small amounts of temnplate DNA are used as stauting material in a PCP, prinmers that differ slightly in sequence from die corresponding region in a template DNA can be used to gencrate relatively larp quantities or a specific DNA fragment doat differs from the template sequence only at the pq(ioons where the nmrsdiffer from the tarplate. For introduction of a autation into a plasmid DNA, one of the primers is designed wo overlap the position of the wtan and to contain the mutation; the sequence of the other primer must be idtentical to a stretch of sequence of die opposite strand of the plasmid, but this sequence can be located anywhere along the plasmid DNA. It is preferred. however, that the sequence of the second primer is located within 200 micleotides from that of the first, such thai in the end the entire amplified region of DNA bounded by the primers can be easily sequenced. PCR amplification using a primer pair l ikete one just described results in a population of DNA fragments that differ at the position of the mutation specified by the primer, and possibly at other positions, as template copying is somewhat error-prone.

Another method for preparing variants, cassette mutagenesis. is based on the technique described by Wells er at., fg, M4: 315 (1985). The starting material is the plasmid (or other vector) comprising the PR0317 DNA .tobemated. The codon(s) in the PR0317 DNA to be imutated are identified. There must be a unique restriction cadonuclease site on each side of the identified mutation site(s). If no such restriction sites exist, they may be generated using the above-described oligonucleotide-adi e imutagenesis method to introduce them at appropriate :locations in the PR0317 DNA. After the restriction sites have been introduced into the plasmid, the plasmid is cut at these sites to linearize it. A double-stranded oligonucleocide encoding the sequence of the DNA between the restriction sites but containing the desired mutation(s) i s synthesized using standard procedures. The two strands are :syntesized separately and then hybridized together using standard techniques. This double-stranded oligonucleotide is referred to as the cassette. This cassette is designed to have 3' and 5' ends that are compatible with the ends of ute linearized plasmid, such that it can be directly ligated to thc plasmid. This plasniid now contains the mutated PR0317 DNA sequence.

Covalent modifications of PR0317 are also irluded within the scope of this invention. One type of covalent modification includes reacting targeted amnin acid residues of the PRO317 with an organic derivatizing agent that is capable of reacting with selected side chains or the N- or C- terminal residues of the PROM 1. Derivatization with bifunctional agents is useful. for instance, for crosslinkig PR0317 to a water-insoluble support matrix or surface for use in the method for purifying anti-PR03 P antibodies, and vice-versa. Commonly used crosslialting agents inrlude l.1-bis(diazoaceYt2-phenylethane. glutaraldehyde, N-hydroxysuccinimide esters, for example. eaters with 4-azidosalicylic acid. hoinobifumctional inidoesters, including disuccinimidyl esters such as 3.3'-dithlobis- (succinimidyipropionate), bifunctional maleimides such as bis-N-malcimido-l .9-octane, and agents such as inethyl-3- ((p-idopheny~ditio)propioinidate.

Other modifications include deamidation of glutainyl and asparaginyl residues to the corresponding glutamyl and aspartyl residues, respectively. hydroxylation of proline and lysine, pbosphorylation of hrydroxyl groups of seryl or threonyl residues, inethyladon of the *-araino groups of lysine. arginine. andl histidine side chains CUE Creight Proteins: Structure and Molecular EPrtrne. W.H. Freemuan Co.. San Francisco. pp. 79-86 (1983)).

acatyladon at the N-terminal amine. and aridation of any C-terniinaJ carboxyl group.

Another type of covalent modification of the PR0317 polypeptide included within the scope of this invention comprises altering the native glycasylation pattern of the polypeptde. "Altring the natve glycosylation pattern* is intended for purposes herein to mean deleting one or more carbohydrate moieties found in native-sequence PRO polypeptde. and/or adding onm or more glycosyhation sites that are not present in the native-6equec PRO polypeptide. The deduced amino acid sequence of PR0317 shwn in Fgure 42 (SEQ ID NO: 114) has one predicted N-linked glycosylation site at residue 160.

Addition of glycosylation sites to the PR0317 polypeptide may be accomplished by altering the amino acid sequence. The alteration may be nude, for example. by the addition of. or substitution by, one or more serine or threonine residues to the native-sequence PR0317 (for 0-linked glycosylation sites). The PR0317 amino acd scquci may optionally be altered through changes at the DNA evel, particularly by mutating the DNA encoding the PR0317 polypeptde at preselected bases such that codons are generated that will translate into the desired amino adds.

Another man of eai ig die number of carbohydrate moieties on the PR0317 polypeptdec is by chemical or enzymatic coupling of glycosides to the polypeptide. Such methods are described in the art, in WO 97/05330 published I1I September 1987, and in Aplin and Wriston, CRC Crit Rev- Biochern.. pp. 259-306 (1981).

Remnoval of carbohydrate moieties; present on the PR0317 polypeptde may be accomplished chemically or enzymatically or by mutational substitution of codons encoding amino acid residues that serve as targets for glycosylation. Chemical deglycosylation fthnjucs ame known in the art and described, for instance, by Hakimuddin.

er al., Arch, Biochem. Bophvs. M:52 (19M7 and by Edge et al.nl ohr., JUj 131 (1981). Enzymatic cleavage of carbohydrate moieties on polypeptides can be achieved by the use of a variety of endo- and exoglycosidases as described by Thotakura a al., Meth. EnzynxlL. 131:350 (1987.

Another typ of covalent modification of PR0317 comprises linking the PRO317 polypetdecto one of a variety of nonproteinaceous polymers, polyethylene glycol. polypropytene glycol, or polyoxyalkylenes, in the manner set forth in U.S. Patent Nos. 4,640,835; 4.496.689; 4.301.144; 4.670,417; 4.791,192 or 4,179,337.

The PR0317 of the present invention may also be modified in a way to farm a chimeric molecule comprising PR0317 fused to another. heterologous polypeptide or amnino acid sequence. In one embodiment, such a chimeric molecule comprises a fusion of the PR0317 with a tag polypeptide which provides an epitope to whichi an anti-tag antibody can selectively bind. hec epitope tag is genierally placed at the amino, or carboxyl- termis of the PROM1.

The presence of such epitope-tagged forms of the PR0317 can be detected using an antibody against the tag polypeptide. Also, provision of the epitope tag enables the PRO3 17 to be readily purified by affinity purification using an anti-tag antibody or another type of affinity matrix that binds to the epitope tag. In an alternative embodiment, the chimeric molecule may comprise a fusion of the PR0317 with an immunoglobulin or a particular region of an itumnoglobulin. For a bivalemt form of the chimeric molecule, such a fusion could be to the Fc region of an lgG molecule.

Various tag polypeptides and their respective antibodies are well known in the art. ExAmples include polyhistidixt (poly-his) or poly-histidine-glycine (poly-liis-gly) tags; the flu HA tag polypeptide and its antibody 12CA5 (Field af aL. Cel Bi 1 :2159-2165 (1998)). die c-myc tag and the 81F9, 3C0, 6El0, G4, B7. and 9E10 antibodies thereto (Evan ea Molecular AMd Cellular Biologx, 5:3610-3616 (1985)); and the Herpes Simplex virus glycoprotein D (jD) gag and its antibody (Paborsky etaat.. Prti Enj rig (6):547-553 (1990)). Other tag polypeptides include the Flag-peptide (Hopp er at.. Bicanlory. fi:1204-1210 (1988)); the KT3 epitope peptide (Martin eta9!., 5~ Z: 192-194 (1992)); an '-tubulin epitope peptide (Skinner et at.. L Bil Chm 20: 15163- 15166 (1991)); and the TY genie 10 protein peptide tag (IAm-Freyermuh et al, Proc. Nail. Acad. Sci 7 USA- f2:6393- 6397 (1990)).

53. Prepuration of PRO Polypentides The description below relates primarily to production of PRO polypeptides by culturing cells transformed or transfecied with a vector containing the desired PRO polypeptide nucleic acid. It is, of course, contemplated that alternative methods. which are well known in the art, may be employed to prepare the PRO polypeptide. For ame she PRO polypeptde sec~ or portions thereof, may be produced by direct peptide synthesis using solidphase techniques (see. Stewart es al.. Solid-Phase PEpide Syndheis, W.H. Freem= San Francisco. CA (1969); Merrifield, JAm. Chem, Soc 15:2149-2154 (1963)]. In itro protein synthesis may be performed using manual techniques or by automation. Automated syuheis may be accomplished. for instance, using an Applied Biosystenms Peptide Synthesizer (Foster City. CA) using manufacturer's instructions. Various portions of dhe desired PRO polypeptide may be chemically synthesized separately and combined using chemical or enzymatic methods to produce the hAl4ength PRO polypeptide.

:*20 Isolation of DNA Encodine PRO Pollpsptides DNA encoding PRO polypeptides may be obtained from a cDNA library prepared from tissue believed to :possess die desired PRO polypeptide niRNA and to express it at a detectable level. Accordingly, human PRO polypeptde DNA can be conveniently obtained from a cDNA library prepared from human tissue, such as described in the Examples. The PRO polypeptide-encoding gene may also be obtained from a genomic library or by oligonucleotide synthesis.

Libraries can be aceried with probes (such as antibodies to the desired PRO polypeptide or oligonucleotides of at least about 20-80 bases) designed to identify die gene of interest or dhe protein encoded by it. Screening the eDNA or genomic library with the selected probe way be conducted wing standard procedures, such as described in Sambrook et al., Molculr Cloning: A LajboatoryManual (New York: Cold Spring Harbor Laboratory Press, 1989). An alternative means to isolate the gene encoding the desired PRO polypeptide is to use PCR methodology [Sambrook et al.. u=g; Dieffenbach et al., PCR PrimnirA I.aboratory Manual (Cold Spring Harbor Laboratory Press, 1995)].

The Examples below describe techniques for screening a cDNA library. The oligonicleouide sequences selected as probes should be of sufficient length and sufficiently unambiguous that false positives are minimized. Thei oligonucleotide is preferably labele such that it can be detected upon hybridization to DNA in the library being screened. Methods of labeling are well known in the art, and include the use of radiolabels like 'P-labeled AlP, biotinylation or enzyme labeling. Hybridization conditions, including moderate stringency and high stringency, are provided in Sambrook ct al.. am._ Sequences identifie *d in such library screening methods can be compared and aligned to other known sequences deposited and available in public databases such 2s GeeBank or other private sequence databases.

Sequence identiy (at either the amino add or smceoside level) within defined regions of the mlecule or across the Mlength sequence can be determined through sequence aligrnent using computer software programs such as BLAST. ALIGN, DNAstar. and INHERI which employ~ various algorithms to measure homology.

Nucleic acid having protein coding sequence may be obtained by screening selected cDNA or genomic libraries using the deduced amino acid sequence disclosed herein for the first time, and, if necessary, using* convenional prb=u extension procedures as described in Samibrook etal., RM, to detect precursors and processing intermediates or niRNA that may not have been reverse-transcribed into eDNA.

B. SelectionM and Transfonnation of Host Cells Host cells art transfected or trantsformed with expression or cloning vectors described herein for PRO polyepid production and cultund in conventional autrient media modified as appropriate for inducing promoters, selecting tr-ansformants. or amplify'ing the genes encoding the desired sequences. The culture conditions. such as media, temperature. pH and the like, can be selected by the skilled artisan without undue experimentation. In general. principles, protocols, and practical techniques for maximizing the productivity of cell cultures can be found in MamMalian Cell BiotechnolozY7 a Pracical AyMroach, M. Butler, ad. (IRL Press. 1991) and Samb~rook at al..

'llm.

Methods of transfaction are kniown to the ordinaril skilled artisan, for example, CaPO, 4 and electoporation.

Depending on the host cell used. transformation is performed using standard techniques appropriate to such cells.

The calcium treatment employing calcium chloride, as described in Sambrook et al., j= or eleetroporation is gerally used for prokaryotes or other cells that conzain substantial celt-wall barriefs. Infection with Agrobadreruan rnzefadtat is used for transformnation of certain plant cells, as described by Shaw et al.. G= 22:315 (1983) and WO 89/05859 published 29 June 1989. For mammalian cells without such cell walls, the calciumn phosphate precipitation method of Graham and van der Eb. Virology. 52:456-457 (1978) can be employed. General aspects of manmaian cell host system ttansfornions have been described in U.S. Paten No. 4,399.216. Transformations *into yeast are typicaly carried out accordfing to the method of Van Solingen et J. Bat. L :946 (1977) and Hsiao.

cc aL. Proc- Nad Aad- Sc (US 2§W39 (1979. However, other methods for introducing DNA into cells, such as by nuclear microinjection, electroporation, bacterial protoplast fusion with intact cells, or polycations. e.g..

polybruz. polyomnithine. may also be used. For various techniques for transforming mammalian cells, sea Kerwn et al., Methods in Enzymology .JU527-537 (1990) and Mansour em al.. Nkturt. 2M:348-352 (1988).

Suitable host cells for cloning or expressing the DNA in the vectors herein include prokazyote. yeast, or higher eukaryote cells. Suitable prokaryotes include but are not limited to eubacteria, such as Gram-negative or Gram-positive organism for example, Enterobacteriaceae such as E. coi. Various E. coil strains are publicly available, such as E. coil K12 strain MM294 (ATCC 31.446); E. coli X1776 (ATCC 31.537); E. coil strain W31 (ATCC 27-125) and K5 772 (ATCC 53,635). Other suitable prokaryotic host cells include Enterobacteriaceac such as Eich eridma, E. coi, Exuerobacder *Eninia, Kiebsiella, Proieus. Salmonella, Salmtonella typhimuriwn, Sa'rtuia, e-g. Serraia mwcexauw, and Sidgella. as well as Bacillf such as B. juiul and B. fichemzformis B.

licheniformis 41P disclosed in DD 266,710 published 12 April 1989). Pseudononas such as P. aeniginosa. and Sl7eponyces. Various E. call stains are publicly available, such as E. cvii K12 strain MM294 (ATCC 31.446); E.

coli X 1776 (ATCC 31,537); E. cod strain W3 110 (ATCC 27,325); and KS 772 (ATCC 53,635). These examples are iLhustr4ve rather than limiting. Strain W3110 is one particularly preferred host or parent host because it is a conmmon host strain for recombinan DNA product fermentations. Preferably, the host cell secretes suinimai.amoto of protcolytic enzymes. For example, strain W3110 many be modified to effect a genetic mutation in the genes encoding proteins endogenous to the host, with examples; of such hosts including E. coii W31 10 strin IA2. which has the complete genotype wanA E. coil W31 10 strain 9E4, which has the complete genotype tonA prr3; E. coi W3110 strain 27C7 (ATCC 55.244), which has the complete genotype fonA prrphoA EI5 (argF-lac)I69 degP WmpTka'; E. Cvii W3110 strain 37D6. which has the complete genotype IonA ptri phaA E15 (argF-lac)169 degP WopT rbs7 lvG kon' E. coi W31 10 strain 40B4, whieh is strain 37D6 with a non-kanamycin resistant degP deletion nztaionand an E. coli strain having mutant per plasmic protease disclosed in U.S. Patent No. 4,946,783 issued 7 August 1990. Alternauively, in viaro methods of cloning, PCR or other nucleic acid polymerase reactions, are suitable.

In addition to prokaryotes. eukazyotic microbes such as flamentous fungi or yeast arc suitable cloning or expression hosts for PRO polypepadceccding vecturs. Saceharomycer cere'dsiae is a conmmonly used lower eukaryotic host microorganism. Others include Scfulasaccharomyces pombe (Beach and Nurse. N~ammi 22D: 140 11981). EP 139,383 published 2 May 1985); Khuyvromtyces hosts Patent No. 4,943,529; Fleer et ai.

Bkfeinmwlg. 2: 968-975 (1991)) such as. la crs (MW984C. CBS683, CBS4574; Louvencourt et at., L BactcroL-, 737 11983D. K. fragilis (ATCC 12,424). K. bulgganczsr (ATCC 16,045), K. Wckerai (ATCC 24.178).

K. %Wui (ATCC 56.500), K. drosophilenam (ATCC 36,906; Van den Berg el ai.. Bio/Technology, 1: 135 (199)).

K. rhermorolerwus, and K. mwrxkaus yarrowla (EP 402,226): Pichiapastoris (EP 183,070; Sreekrishna et vi. *L.

Baic Mirobiol.. 265-78 [198]; CanzdidA-~ Taicoden~na reesia (EP 244.234); Nezsrospora cnusa (Case ea ai., Proc- Nat]i Acad. Sdi USA, L 259-5263 (1979]); Schwtmnniomyces such as Schwwniomyces occidenualis (EP 394,538 published 31 October 1990); and filamentous fungi such as, Neurospora. Pencidiium. Totypocladiwn (WO 91/00357 published 10 January 1991). and AspergiiW hosts such as A. nidulans (Ballance ar at., Bicen *.*Bionys Res- Comm=ii, In: 284-289([19831; Tilburn et at., fim 205-221 (19831; Yelton eat.. PMC. Nad- Acd Sc.US.l: 1470-1474 (1984]) and A. niger (Kelly and Hynes. EMBO 4: 475-479 (1995D).

Methylotropic yeasts are suitable herein and include, but arm not limited to, yeast capable of growth on methanol selected from the genera consisting of Hansenula, Candida, Kibeckera, Pichia. Saccharomyces, Torudopsis. and Rhodowrnda. A list of specific species that ame exemplary of this class of yeasts may be found in C. Anthony. 33M Biochemistry of Methvlogrnhs, 269 (1982).

Suitable host cells for the expression of glycosylated PRO polypeptides are derived from roulticellular organisms. Examples of invertebrate cells include insect cells such as Drosophila S2 and Spodoptera sf9, as well as plas cells. Examiples of useful mammalian host cell lines include Chinese hamnster ovary (CHO) and COS cells.

Mote specific eamples include monkey kidney CVl line transformed by SV40 (COS-7. ATCC CRL 1651); human embryonic kidney line (293 or 293 cells subeloned for growth in suspension culture, Graham et al., 1. n irol., M:59 (1977)); Chimes hamster ovary cefls-DHFR (CHO, Urlab and Chasin, Proc- Nati. Aead- Sco USA,. 22:4216 (1990)): use senoli cefls (Th14. Madicir, Biol Rc=W. fl:243-25I (1980)); human lung cells (W138, ATCC CCL human liver cells (Hcp G2. lID 8065); and mousc mammzary tumor (MMT 060562. ATCC CCL5 The selection of the appropriate host cell is deemed to be within the skill in the art.

C. Selection andiUse of a Replicable Vector The nuceic acid cDNA or genomic DNA) encoding a desired PRO polypeptide may be inserted into a replicable vector for cloning (amplification. of the DNA) or for expression. Various vectors are publicly available.

The vector may. for examnple, be in t form of a plasmid. cosmid. viral particle, or phage. The appropriate nucleic acid sequence may be inserted into the vector by a variety of procedures. In general. DNA is inserted into an appropr iate restriction endorocicase sit(s) using techniques known in the amt Vector components generally include, but are not limited to, one or more of a signal sequence, an origin of replication, one or more marker genes, an enhancer ena a promoter, andl a transcription termination scquw. Construction of suitable vectors containing one or more of these components; employs standard ligation techniques which are known to the skilled artisan.

The PRO polypeptide of interest may be produced recombinantly not only directly. but also as a fusion polypepid with a heterologous polypeptide, which nay be a signal sequence or other polypeptdec having a specific cleavage site at the N-terminus at the mature protein or polypeptide. In general, the signal sequence may be a component at the vector, or ft may be a pan at the PRO polypeptide DNA that is inserted into the vector. The signal :sequence may be a prokaryotic signal sequence selected, for example, from the group of the alkaline phosphatase.

penicillinase, lpp, or heat-stable enterotoxin BI leaders. For yeast secretion the signal sequence may be, the yeast invertase leader, alpha factor leader (including Sacdzaromyces and Kluyveronryces a-factor leaders, the latter described in U.S. Patent No. 5,010,182). or acid phosphatase leader, the C. albicans glucoantylase leader (EP 362.J79 published 4 April 1990), or the signal described in WO 90/13646 published 15 November 1990. In mmmalian cell expression, rmmalian signal sequncs may be used todirec sceinothpreinscasigl sequences from secreted polypcpdecs at the same or related species, as well as viral secretory leaders.

Both expression and cloning vectors contain a nucleic acid sequence that enables the vector to replicate in one or more selected host cells. Such sequences are well known for a variety of bacteria, yeast, and viruses. The origin of replication from the plasmid pBR322 is suitable for most Gram-negative bacteria, the 2 pu plasmid origin is suitable for yeast, and various viral origins (SV40, polyoma, adenovirus, VSV or BPV are useful for cloning 'vectors in mammalian cells.

Expression and cloning vectors will typically contain a selection gene, also termed a selectable marker.

Typical selection genes encode proteins that confler resistance to antibiotics or other toxins. ainpielllin.

neomycin, methotrexate, or tetracycline, complement auxotrophic deficiencies, or supply critical nutrients not available from complex media, dite gene encoding D-alanine racemase for Baifi.

An example of suitable selectable markers; for mammalian cells are those that enable the identification of cells competent to take up the PRO polypeptide nucleic acid, such as DHFR or thymidine kinase. An appropriate host cell when wild-type DI{FR is employed is the CHO cell line deficient in DHFR activity, prepared and propagated as described by Urlaub et al., Proc. Nad. Acad_ Set lISA, Zl:4216 (1980). A suitable selection gene for use in yeast is the 11PI Sene Present in the Yeast plasmid yRp7 (Stinchenaib ct al.. Ntlaiig. W.:39 (1979); Kingsnmnt al., =,2:141 (19179): Tahemper et al.. e 1D.157 (1980)). The typt gene provides a selection maxker for a utart 9train Of Yeast lacking the biliy to grow in tryptohati. for example. ATCC No. 44076 or PEPN- I [Jones, Geeds 15:12 (177)].

Expresiion and cloning vectors usually contain a promoter operably linked to the PRO polypeptide nucleic acid sequence to dimac mRNA synthesis. Prormoters recognized by a variety of potential host cells arc~veU known.

Promoters suitable for use with prokaryotic hosts include the P-Lama2e and lactose promoter systems [Chang at al., Nlature. 2U-.615 (1978); Goeddel et al. Nature, 21:544 (1979)]. alkalinc pbosphatasc, a tryptophan (trp) promoter system (Goeddel. Nuli Acd e. 1:4057 (1980); EP 36,7761, and hybrid promoters such as the tac promoter (deBocr et al., Em-c Nail Aad, Scd USA. AQ2-25 (1983)1. Promoters for use in bacteria systems also will contain a Shinc-Daigariio sequence operably linked to the DNA encoding the desired PRO polypeptide.

Examples of suitable promotintg sequences for use with yeast hosts include the promoters for 3- %:oplgycrl kinase [itreman et al.. J- igLQ Cb 255-2073 (19M]) or other Slycolytic enzymes [Hess et al., J Adv. EnzyMereu 149 (1968); Holland, BjQb~minD. 12:4900 (1978)). such as ewlasc, glyccraldehyde-3phoslute ddhydogenase, hexolwmzse pymuate decArboxylase, phosphofiructokiiiue. glucose-6-phosphate isomerase.

15 3-phosphoglYcerat: =mase, pynivate kinase, tri~pose sphatc isomerase, phosphoglucose isonierase, and glucokinase.

Other yeast promoters, which arc inducible promoters having the additional advantage of transcription controlled by growth conditions, are the promoter regions for alcohol dehydrogenase 2, isocytochromle C, acid hosphatasc. degradative enzymes associated with nitrogen metabolism, metallodioieitl. glyceahydc3-bosphat dehydrogenase. and enzymes responsible for maltose and galactose utilization. Suitable vectors and promoters for use inyeast expression arfurthr dscndin EP 73,67 PRO polypeptide transcription from vectors in mammalian host cells is controlled, for example. by promoters; obtained from the genomes of viruses such as polyomna virus, fowlpox virus (UK 2.211.504 published July 1989). adenovirus (such as Adenovitus bovine papilorna virus, avian sarcoma virus, cytomegaloviflis. a retrovirus, hepatitis-B virus and Simian Virus 40 (SV40), from heterologous mammalian promoters. the actini promoter or an inmumoglobulin promoter. and from heat-shock promoters, provided such promoters are compatible with the host Cell systems.

Transcription of a DNA encoding the desired PRO polypeptide by higher eukaryotes may be increasecd by inserting an enhancer sequence into the vector. Enhancers are cis~acting elements of DNA. usually about from to 300 bp, that act on a promoter to increase its transcription. Many enhancer sequences are now known from mammalian genes (globin. elastase. albumin. a-fectoproteia. and insulin). Typically. however, one will use an enharcer from a cukaxyotic cell virus. Examples include the SV40 enhancer on the late side of the replication origin (bp 10-70). the cytnvnegalovitiis early promoter enhancer, the polyonta enhancer on the late side of the replication origin, and adenovirus enhancers. The enhancer way be spliced into dhe vector at a position 5S or 3* to the PRO po4ypptde coding sequence, but is preferably located at a site S' from the promoter.

Expression vectors used in etakaryotic host cells (yeast, fungi, insect, plant, animnal, humn, or nucleated cells from other nnlticellu organisms) will also contain sequences necessary for the termination of transcription and for stabilizing the mRNA. Such sequences are commonl available from the 5' and, occasionally untranslated regions of eukaryotic or viral DNAs or cDNAs. These regions contain nucleocide segments transcribed as polyadenylaxed fragments in the unmrnslated portion of the mRNA cncoding PRO polypeptides.

Still other methods, vectors, and host cells suitable for adaptation to the synthesis of PRO polypeptides in recombinant vertebrate cell culture are described in Getting et al., bNtur. 2n~:620-625 (1981); Mantei et al., Nature, 2U:40-.46 (1979); EP' 117.060; and EP 117.058.

D. Detectinr Gene AmnnifictonlErnression Gene amplification and/or expression may be measured in a sample directly, for example, by conventional Southcmoblotting, Nonhern blotting to quantitate the transcription of :nRNA rlboias. Proc. NatI. Acad- Sci. USA, Th:5201-5205 (1980)]. dot blotting (DNA analysis), or in sit hybridization, using an appropriately labeled probe.

based on the sequences provided herein. Alternatively, antibodies may be emiployed that can recognize specific duplexes. inchling DNA duplexes, RNA thipexes, and DNA-RNA hybrid duplexes or DNA-protein duplexes. The antibodies in turn mnay be labeled and the assa may be carried out where the duplex is bound to a surface, so that upon the formation of duplex on the surface, the presence of antibody bound to the duplex can be detected.

Gene expression, altenatively. may be measured by iraratinlogical methods, such as inununohistocheniical staining of cells or tissue sections and assay of cell clture or body fluids, to quantitate directly the expression of gene product. Antibodies useful for iminnolistodienkal staining and/or assay of sample fluids my be either monoclonal or polyclonal, and may be prepared in any mammal. Conveniently, the antibodies may be prepared against a native sequence PRO polypeptide or against a synthetic peptide based on the DNA sequences provided herein or against exogenous sequence fused to a PRO polypeptide DNA and encoding a specific antibody epitope.

E. Purification of Polynentide Form o(PRO polypptides may be recovered from culture medium or from host cell lysates. If membranebound, it can be released from the membrane using a suitable detergent solution Triton-X 100) or by enzymatic cleavage. Cells employed in expression of PRO polypeptides can be disrupted by various physical or chemical means, such as freeze-thaw cycling, sortacation. mechanical disruption, or cell lysing agents.

It may be desired to purif PRO polypeptides from recombinant cell proteins or polypeptides. The following procedures are exemplary of suitable purification procedures: by fractionation on an ion-excng coun ethaol precipitation; reverse phase HPLC; chromatography on silica or on a cation-exchange resin such as DEAF, chromatofocusing; SDS-PAGE; amnmonium sulfate precipitation; gel filtration using, for example, Sephadex 0-75; protein A Sepharose coluas to remeve contaminants such as lgG; and metal chelating columins to bind epitopetagged forms of the PRO polypeptide. Various methds of protein purification may be employed and suach methods are inown in the art and described for example in Deutscher, Methods in Enzvrooe, 132(1990); Scopes, £wisin Purification: Princiles and Pracc, Springer-Verlag, New York (1982). T'he purification step(s) selected will depend, for example, on the nature of the production process used and the particular PRO polypeptide produced.

S4. Uses for PRO Polyneutldes Nucleodde sequences (or their complement) encoding t PRO polypeptides ot die prcsen invention have various applicatiowns at d art of molecular biology. including uses as hybridization probes. in chromosome and gene mapping and in the generation of anti-sene RNA and DNA. PRO polypcpdde-enooding nucleic acid will also be useful for the preparation of PRO polypeptides by the recombinant techniques described herein.

The full-length native sequence PRO polypeptide-enooding nuclic acid or portions therof. may be used as hybridization probes for a cDNA library to isolate die fullcngth PRO polypeptide gene or to isolate still other geme (for instance, those encoding naturally-ocazrring variants of the PRO polypeptide or PRO polypeptides from othe species) which have a desired sequence identify to the PRO polypeptide nucleic acid sequeces. Optionally.

the length of the probes will be about 20to about 50lass. Tebybridzadon probes may be derived from the micleotde sequec of any of the DNA molecules disclosed herein or from genomie sequence including promoters.

enancer elements and Wnrons of native sequnce PRO polypeptide encoding DNA. By way of example, a screening method will comprise isolatin the coding region of the PRO polypeptide gene using the known DNA sequecec to syethesize a selected probe of about 40 bases. Hybridization probes may be labeled by a variety of labels, including radionucleotides such as up or," S. or enzymatic labels such as alkalin phosphatase coupled to the probe via avidinibiotmn coupling system. Labeled probes having a sequence complementary to that of the specific PRO polypeptide geme of the present invention can be used to screen libraries of burmn cDNA. genomic DNA or mnRNA to determine which members of such libraries the probe hybridizes to. Hybridization techniques are described in further detai) in the Examples below.

The ESTs disclosed in the present application may similarly be employed as probes, using the methods r~ go.20 disclosed herein.

Thc probes may also be employed in PCR techniques to generate a pool of sequences for identification of closely related PRO polypeptide sequences.

0 Nucleotide sequences encoding a PRO polypeptide can also be used to construct hybridization probes for see. mapping the gene which encodes that PRO polypeptide and for the genetic analysis of individuals with genetic disorders. The nucleotide sequences provided herein may be mapped to a ch romosome and specific regions of a chromosome using known techniques, such as in situ hybridization, linkaige analysis against known chrormosomal markers, and hybridization screening with libraries.

The PRO polypeptidc can be used in assays to identify its ligands. Similarly. inhibitors of the rnccpror/lipr bindilng inkteior can be idecntified. Proteins involved in such binding interactions can also be wsed to screen for peptide or small mtolecule inhibitors or agonists of the binding interaction. Screening assays can be designed to find lead compounds tha mimic the biological activity of a native PRO polypeptide or a ligand for the PRO polypeptide. Such screening assays will include assays amtenable to high-throughput screening of chemnical libraries, making them particularly suitable: for identifyin small molecule drug candidates. Small molecules contemplated include synthetic organic or inorganic compounds. The assays can be performed in a variety of formats, including protein-protein binftn assays, biochemical screening assays. imunoassays and cell based assays.

which are well characterized in the art.

Nucleic acids which encode a PRO polypeptide or its modified forms can also be used to generate either transgenic animals or 'knock out" animals which, in turn. are useful in the development and sacning of therapeutically useful reagents. A transgcnic, animal a mouse or rat) isian animal having cells that contain a umangene, which transgene was intred into dhe animal or an ancestor of the animal at a prenatal, an embryonic stage. A transgene is a DNA which is itegrated into the genotne of a CeCl (nin which a transgcnic animal develops. In one embodiment, cDNA encoding a PRO potypeptide of interest can be uased to clone genomic DNA encoding the PRO polypeptide in accordance with established Wcchiquc and [be genoniic sequences used to generate rangenic animls hat contain cells which express DNA encoding the PRO polypeptide. Methods for generating transgenic anifml, particularly animals such as mice or rats, have become conventional in the arn and arc described.

for example, in U.S. Patent Nos. 4.736.866 and 4,870.009. Typically, particular cells would be targeted for PRO polypeptide ransgene incorporation with tissue-specific enhancers. Transgenic; animals that include a copy of a transgene encoding a PRO polypqxide inrduced into the germ line of the animal at an embryonic stage can be used ~to examine the effect of increased expression of DNA encodin tePRO polyetdec. Such Animals can be used as teste animals for reagents thought so confer protection from, for example, pathological conditions associated with its overexpression. In accordance with this facet of the invention, an animal is treated with the reagent and a reduced incidence of the pathological condition, compared to untreated animals bearing the trans gene, would indicate a potential therapeutic intervention for the pathological condition.

Alternatively. non-human homiologues of PRO polypeptides can be used to construct a PRO polypeptide 'knock out" animal which has a defective or altered gene encoding the PRO polypeptide of interest as a result of homologous recombination between the endogenous gene encoding the PRO polypeptide and altered genomic DNA 20 encoding the PRO polypeptide inrouced into an embriyonic cell of the animal. For example, cDNA encoding a PRO polypeptide can be used to clone genomic DNA encoding the PRO polypeptide in accordance with established techniques. A portion of the genomic DNA encoding a PRO polypeptide can be deleted or replaced with another gene, such as a gene encoding a selectable marker which can be used to monitor integration. Typically, several kilobases of unaltered flanking DNA (both at the 5' and 3' ends) are included in the vector [see Thomas and Capecclii, fmU, aM0 (1987) for a description of homiologous recomnbination vectors]. The vector is introduced into an embryonic stem cell line by electroporation) and cells in which the introduced DNA has homologously' recombined with the endlogenous DNA are selected [see Li et al.. "efl, fi2:915 (1992)]. The selected cells arm then injected into a blastocyst of an animal a mouse or rat) to form aggregation chimeras [see Bradley, in Terrvocardnomas W&d Embryonic Stem Cells: A Pracrical Approach. E. J. Robertson. ed. (IRL, Oxford, 1987)w.o 113-152]. A chimneric embryo can then be implanted into a suitable pseudopregnant female foster animal and the embryo brought to term to create a "knock out' animl. Progeny harboring the homologously recombined DNA in their germ cells can be identified by standard techniques and used to breed animals in which all cells of the animal contain the homologously recombined DNA. Knockout animals can be characterized for instance, for their ability to defend against certain pathological conditions and for their development of pathological conditions due to absence of the PRO polypeptide.

With regard to the PRO211I and PRO217 polypeptide, therapeutic indications include disorders associated with the preservation and unintennc of gastrointestinal mucosa and the repair of acute and chronic mucosal lesions euzerocolitis. Zollinger-ElLison syndrome- gastrointestinal ulceration and congenital microvifus atrophy), skin diseases associated with abnormal keratinocyte differentiation psoriasis. epithelial cancers such as lung squamous cell carcinoma, epidcriwid carcinomna of the vulva and gliomas.

Sinc the PR0232 polypeptdec and nucleic acid encodiT it possess; sequence homology to a cHl surface stem cell antigen and its encoding ucleic acid. probes based upon the PR0232 nucleotide sequence may be employed to idenify othier novel stem cell surface antigen proteins. Soluble forms of the PRO232 polypeptide way be employed as antagonists of membrane bound PRO232 activity both an vitro and w vivo. PR0232 polypeptides; may be employed in screening assays designed to identify agonists or antagonists of the native PR0232 polypeptide, wherein such assays may take the form of any conventional cell-type or biochemical binding assay. Moreover, the PR0232 polypepide may serve as a molecular markter for the tissues in which the polypeptide is specifically expressed.

With regard to the PRO187 polypeptides; disclosed herein, FOE-8 has beent implicated in cellular differentiation and embnyogeneis, including the patterning which appears during limb formation. FGF-8 aiul the PRO 187 molecules of t invention therefore are likely to have potent effects on cell growth and development.

Diseases which relate to cellular growth and differentiation are therefore suitable targets for therapeutics based on functionality similar to FGF-8. For example, discases related to growth or survival of nerve cells including Parkinson's disease, Alzhteimer's disease, ALS, neuropathies. Additionally, disease related to un~ontrolled cell growth, cancer, would also be expected therapeutic targets.

With regard to the PR0265 polypeptides disclosed herein, other methods for use with PRO265 are described in U.S. Patent 5.654.270 to Ruoslahti et al. In particular, PR0265 can be used in comparison with she fibromodulin disclosed therein to compare its effects on reducing dermal scarring and other properties of the fibroinodulin described therein including where it is located and with what it binds and does not.

The PR0219 polypeptides of the present invcntion which play a regulatory role in the blood coagulation *:cascade may be employed in vivo for therapeutic purposes as well as for in vitr purposes. Those of ordinary skill in the art will well know how to employ PR0219 polypeptides for such uses.

The PR0246 polypeptides of the present invention which serve as cell surface receptors for one or more viruses will find other uses. For example, extraceliular domains derived from these PR0246 polypeptides may be employed therapeutically in viv for lessening the'effects of viral infection. Those PR0246 polypeptides which serves as tumoer specific antigens may be exploited as therapeutic targets for anti-turnor drugs, and the lik. Those of ordinary skill in the art will well know how to employ PR0246 polypeptides for such uses.

Assays in which connective growth factor and otlher growth factors are usually used should be performed with PR0261. An assay to determine whether TOP beta hnduces PR0261. inilcating a role in canicer is performed as bwnoi in the art. Wound repair and tissue growth assays art also performed with PR0261. The results are applied accordingly.

PR0228 polypeptidcs should be used in assays in which 13MR1. CD97 and latrophilin would be used in to determine their relative activities. The reaths can be applied accordingly. For example, a competitive binding assay with PR0228 and CD97 can be performied with the ligand for CD97, Native PR0533 is a 216 amino acid polypeptide of which residues 1-22 are the signal sequence. Residues 3 to 216 have a Blast score of 509. corresponding to 53 homology to fibroblast growth factor. At the nucleotide level, DNA47412. the EST from which PCR oligos were generated to isolate dhe Mul length DNA49435-1219. has beenobserved to up tolipiS. Sequence lxxnolouyto the IlpIS locus would indicate that PR0533 may have utility in the treatmnt of Usher Syndrome or Atrophia areata.

As mentioned previously. fibroblast growth facor can act upon cells in both a mitogenic arnd non-ozitogenic marner. These aors ame zitogenic for a wide variety of normal diploid mesodermn-deuived and neural crest-derived cells, Imliring gramilosa cells. adrenal cortical cells. chrondrocytes. myoblasts, comeal and vascular endothelial cells (bovinc or human). vascular smooth muscle cells. lens, retdn and prostatic epitheliL- cells. oligodcndrocytes.

astrocytes. chrondocytes, aryoblasts and osteoblasts.

Non-mitogenic actions of fibroblast growth factors include promotion of cell migration into a wound area (diemotaxis). initiation of new blood vessel fonialation (arngomis). modulation of nerve regeneration and survival :9.10 (ncurotrophism), modulation of endocrine fitions, and stimtulation or suppresion of specific cellular protein exrsion exiracellular matrix production and cell survival. Baird, A. Bohlen. Handbook of Lip. Phrmacol.

369.418 (IM9). These properties provide a basis for using fibroblast growth factors In therapeutic approaches to accelerate wound healing, nerve: repair, collateral blood vessel formation. and the lie. For example. fibroblast growth factors, have been suggested to minimize myocardium damage in heart disease and surgery (U.S.P.

4.378,437).

Since the PR0245 polypctide andl ncleic- aci encoding it possess sequence homology to a ramcmbrane protein tyrosine kiznse protein and its emodirig micleic acid, probes based upon the PRO245 inueceotde sequence may be employed to identify other novel trantsinobrane tyrosine kinae proteins. Soluble forms of the PR0245 polypeptd& may be employed as antagonists of nzmtrbrane bound PR0245 activity both in ir and in Wyo. PR0245 polypeptides my be employed in screening assays designed to identify agonists or antagonists of the native PR0245 polypeptide, wherein such assays may take the form of any conventional cell-type or biochemical binding assay.

Moreover, the PR0245 polypcptide may serve as a molecular marker for the tissues in which the polypeptide is specifically expressed.

PR0220. PR0221 and PR0227 all have leucine rich repeats. Additionally, PR0220 and PRO221 have homology to SLIT1 and leucine rich repeat protein. Therefore. these proteins arm useful in assays described in the literature, wherein the SLIT and leucine rich repeat protein are used. Regarding the SLITl protein. PR0227 can be used in an assay to determine the affect of PR0227 on neurodegeitrative disease. Additionally' PR0227 has homology to human glycoprotein V. In the case of PRO22, this polypeptidc is used in an assay to determine its affect on bleeding, clotting, tissue repair and scarring.

Ile PR0266 polypeptide can be used in assays to determine if it has a role in neurodegenerative diseases or their reversal.

PR0269 polypeptides and portions thereof which effect the activity of thrombin may also be useful for in Wwr therapeutic purposes. as well as for various in "fav applications. I n addition, PR0269 polypeptides and portions thereof may have therapeutic use as an antithrombotic agent with reduced risk for hemorrhage as compared with heparmn. Peptides having homology to throibomodulin are particularly desirable.

PR0287 polypeptides and portions thereof which cffcct the activity of bone morphogcnic protein IBMPI */procollagen C-proteinase (PCP) may also be useful for in vow therapeutic purposes, as well as for various 112 vitro applications. In addition. PRO28 polypeptides and portlowM thereof may have therapeutic applications in wound healing and tisse repair. Peptides having homlogy to procoliageii C-protetflase enhancer protein and its precursor may also be used to induce bowe and/or cartilage formation and amt therefore of particular interest to the scientific and medical communites.

Therapeutic indications for PR0214 polypeptides include disorders associated with the preservation and maintenance of gastrointestinal mucosa and the repair of acute and chronic niucosal lesions enterocolitis, Zollinger-lion syndrome, gastrointestinal ulceration and congenital microvillus atrophy), akin diseases associated with abnormal kerauinocyte differentiation psoriasis. epithelial cancers such as lung squamous cell carcinoma.

epidermoid carcinoma of the vulva and gliomas.

Stadie on the generation and analysis of mice deficient in members of the TGF- superfamily are reported in Matzuk. Trends in Endocrinol. &W Metabol. 1 120127 (1995).

The PR0317 polypeptide, as well as PR0317-speciic antibodies, inhibitors, agonists, receptors, or their analogs, herein are useful in treating PRO3 17-associated disorders. Hence, for example, they may be employed in modulating endometrial bleeding angiogenesis, and may also have an effect on kidney tissue. Endometrial bleeding can ocnu in gynecological diseases such as endometral cancer as abnormal bleeding. Thus, the compositions herein may find use in diagnosing and treating abnormal bleeding conditions in the emalometrium, as by reducing or eliminating the need for a hysterectomy. The molecs herein may also find usc in angiogenesis applications such as anti-tumor indications for which the antibody against vascular endothelial growth factor is used, or, conversely.

iseliemic indications for which vascular endothelial growth factor is employed.

Bioactive compositions comprising PR0317 or agonists or antagonists thereof may be administered in a suitable therapeutic dose determined by any of several methodologies including clinical studies on mammalian species to determine maximal tolerable dose and on normal human subjects to determine safe dose. Additionally, the :bioactive agent may be complexed with ai variety of well estahlished compounds or compositions which enhance stability or pharmacological properties vach as half-life. It is contemplated that the therapeutic, bioactive composition may be delivered by intravenous infusion into the bloodstream or any other effective means which could be used for treating problem of the kidney, uterus, endometimm, blood vessels, or related tissue. in the heart or genital tract.

Dosages and administration of PRO317. PR0317 agonist, or PR0317 antagonist in a pharmaceutical composition may be determined by one of ordinary skill in the art of clinical pharmacology or pharmacokinsucs.

See, for example, Monlenti and Rescigno. h, cuia eerh 2:17-25 (199); Morenti at a1. nhraceltica1 Research 11:1351-1359 (1991); and Mordenti and Chappell, -The use of interspecies scaling in toxicokiwetcs* in Toxicokinetics and New Dnr Dvelopmnt Yacobi er at. (eds) (Pergamon Press: NY, 1989). pp. 42-96. An effective amount of FROM 1, PRO3 17 agonist, or PRO317 antagonist to be employed therapeutically will depend, for example, upon the therapeutic objectives, the route of administration, and the condition of the mammal.

Accordingly, it will be necessary for the therapist to titer the dosage and mnodify the route of administration as required to obtain the optimaJ therapeutc effect. A typical daily dosage might range from about 10 ngfkg to up to 100 mg/kg of the nmma's body weight or more per day, preferably about I gig/kg/day to 10 mg/kg/day. Typically, the clinician will admitmister PROM1. PR0317 agooist, or PR0317 antagonist, until a dosage is reached that achieves the desired effect for treatment of the above mentioned disorders.

PR0317 or an PR0317 agonist or PR0317 antagonist may be administered alone or in combination with another to achieve the desired pharmacological effect. PR0317 Itself, or agoists or antagonists of PR0317 can provide different effects whien administerod therapeutically. Such compounds for treatment will be formulated in a nontoxic. inert, pharmaceutically acceptable aqueous carrier medium preferably at a pH of about 5 to 8, more preferably 6 to B. although dhe pH may vary according to the characteristics of the PRO3 17. agonist, or antagonist being formulated and the condition to be treated. Characteristics of the treatment compounds include solubility of the molecule, half-life, and aniigenicityfznmmntogenicity; these and other characteristics may aid in defining an effective carrier.

PR0317 or PR0317 agonists or PR0317 antagonists may be delivered by known routes of administration including but not limited to topical mm and gels; transniucosal spray and aerosol, transdermal patch and bandage; injiectable, Intravenous, and lavage formulations; and orally administered liquids and pills. particularly forulated to resist stomach adid and enzymes. The particular formulation. exact dosage, and route of administration will be determined by the attending physician and will vary according to each specific situation.

Such determinations of administration are made by considering multiple variables such as the condition to 15 be trated, the type of mnammal to be treated. the compound to be administered, and the pharmacokinetic profile of the particular treatment compound. Additional factors which may be taken into account include disease state (e.g.

severity) of the patient, age, weight, gender, diet, sam of administration, drug combination, reaction sensitivities.

:and tolerarce/responsc to thezapy. Long-acting treatment compound formulations (such as liposomally encapsulated PR0317 or PEGylated PR0317 or PR0317 polymeric microspheres. such as polylactic acid-based microspheres) might be administered evry3to 4days. every week, or owce ever two weeks depending on half-life and clearanice :rate of the particular treatment compound.

Normal dosage amounts may vary from about 10 ng/kg to uip to 100 mg/kg of marmal body weight or more per day, preferably about I jpg/kg/day tolO m/k/dy, depending upon the route of admirtistration: Guidance as to particular dosages and methods of delivery is provided in the literature; see, for example. U.S. Pat. Nos.

25 4.657,760; 5,206344; or 5,225212. It is anticipated that different formulations will be effective for different *~*uca~compounds and different disorders, that administration targeting the uterus, for example. may necessitate delivery in a manner different from diat to another organ or tissue, such as cardiac tissue.

Where sustained-release administration of PR0317 is desired in a formulation with release characteristics suitable for the treatment of any disease or disorder requiring administration of PRO3 17, microencapsulation of PR0317 is contemplated. Microencapsulation of recombinant proteins for sustained release has been successfully performed with hrnan growth hormone (rhGH). interferon- (rhlFN- interleukin-2, and MN rgpl120. Johnson er of., kl!L Mod.. 2: 795-799 (1996); Yasuda- fiomned- 'fer 2.2:1221-1223 (199); Horsa erat., Bio/TchnoJogy. 755-758 (1990) Cleland, *Design and Production of Single Immunization Vaccines Using Polylactide Polyglycolide Microsphere Sysu=rns in Vaigzin Desigaw The Suhmlt and Aftynt Anproach Powell and Newman, cds, (Plemima Presi: New York. 1995). pp. 439-462; WO 97/03692, WO 96/40072, WO 96/07399; and U.S Pat. No. 5,654,010.

The sustained-release formulations of these proteins were developed using poly-lactic-coglycolic acid (PLGA) polymer due to its biocompatibility and wide range of biodegradable properties. The degradation products oftPLGA lactic and glycolic acids, can be cleared quickly within the biman body. Moreover the degradability of this Polymer can be adjusted from months to years depending on its molecular weight and composition. Lewis, "Controlled release of bioactivc agents from lactide/glycolide polymner.' in: M. Chauin and R. Lantger (Eds.), Biodeeradable Potymers as QiuP DehlX Systems (Marcel Dekkcer: Ncw York. 1990), pp. 1-41.

For examle. for a formulation that can provide a dosing of approximately 90 g/kglday in mammals with a snazirmm body weight of 85 kg. the largest dosing would be approximately 6.8 mig PR0317 perday. In order to achieve this dosing level, a sustained- release formulation which contains a maximumn possible protein loading (1Swlw PROW1) with the lowest possible initial burst is necessary. A continuous (zero-order) release of PRO317 from microparticles for 1-2 weeks Is also desirable. In addition, the encapsulated protein to bci released should maintain its integrity and stability over the desired releas period.

It is contemplated that conditions or diseases of the uterus, endometrial tissue, or odoer genital tissues or **cardiac tissues may precpitate damage that is treatable with PR0317 or PR0317 agonist where PR0317 expression i reduced in the diseased stae; or with antibodies to PRO3 17 or other PRO3 17 antagonists where the expression of PR0317 is increased in the diseased state. These conditions or diseases may be specifically diagnosed by the probing tests discussed above for physiologic and pathologic problem which affect the function of the organ.

The PRO317. PR0317 agonist, or PRO.317 antagonist may be administered to a mammal with another biologically active agent, either separately or in the same formulation to treat a common indication for which they arc appropriate. For example, it is contemplated that PR0317 can be administered together with EBAF-1 for those indicatin on which they demonstrate the same qualitative biologital effects. Alternatively, where they have opposite effects, EBAF-l may be administered together with an antagonist to PRO3 17. such as an anti-PR0317 antibody.

Further. PRO3 17 may be admninistered together with VEGF for coronary ischemia where such indication is warranted, or with an ant-VEGF for caner as warranted, or, conversely, an antagonist to PRO317 may be administered with VEGF for coronary ischemia or with anti-VEOF to treat cancer as warranted. These administrations would be in effective amounts for treating such disorders.

Native PRO301 (SEQ ID NO:119) has a Blast score of 246 and 30% homnology at residues 24 to 282 of :25 Figure 44 with A33 -HUMAN. an A33 antigen precursor. A33 antigen precursor, as explained in the Background is a turrir-qsecifzc antigmn and as such, is a recognized marker and therapeutic target for the diagnosis and treatment of colon cancer. The expression of tumor-specific antigens is often associated with the progression of neoplastic tissue disorders. Native PRO301 (SEQ ID NO:119) and A33-HUMAN also show a Blast score of 245 and homiology a residuas 21 to 282 of' Fig. 44with A33-HUMAN, doe variation dependent upon how spaces are inserted intecompared sequences. Native PRO301 (SEQ ID) NO:119) also has a Blast score of 165 and 29% homology at residues 60 to 255 of Fig. 44 with HS46KDA-1. a human coxsacie and adenovirus receptor protein, also known as cell surface protein HCAR. This region of PRO301 also shows a similar Blast score and hornology with HSU907161. Expression of such proteins is usually associated with viral infection and therapeutics for the prvnto of such infoction may be accordingly concived. As mentioned in the Background, the expression of viral receptors is often associated with neoplastic tumors.

Theraipeutic uses for the PR0234 polypeptitles of the invention includes treatments associated with leukocyte homing or doe interaction between leukocytes and the endot heliumi during an inflammatory response. Eaples Include asthma, rhteumnatoid arthritis, psoriasis and multiple sclerosis.

SineC the PR0231 polypeptide and nucleic acid encoding it possess sequence homology to a putative acid phosphatase and its ecmodin inuclcic acid. probes based upon the PR0231 inicotide sequenc may be employed to idetifyv othe novel phoqphatase proteins. Soluble forms of the PR023 1 polypesdoc may he employed as antagonists of inmraw bound PR0231 activity both in iro and in vai. PR0231 potypeptides may be employed in screnng assays designed on identify agoaists or antagonists of die native PR0231 poWypptde, wherein such'btsays my take the form of any conventional cell-ype or biochemical binding assay. Moreover, the PR02231 polypeptide may serve as a molecular marker for the tissues in which the polypeptide is specifically expressed.

PRC229 polypeptdac can be fused with peptides of interest to determine whether the fusion peptide has an inceased half-life over the pcptdec of interest. The PR0229 polypeprides can be used accordingly to increase the half-life: of polypeptides of interest. Portions of PR0229 which cause the increase in half-life are an embodiment of the invention herein.

PR28 a be used in assays which measure its ability to reduce substrates, including oxygen a Aceyl- CoA, and particularly, measure PROM38s ability topuc oxygen free radicals. This is done by using assays which have beeni previously described. PRCY238 can futher be used to assay for candidates which block, reduce or reverse its-reducing abilities. This is done by prformning side by side assays where candidates are added in one assay having PR0238 andi a substrate to reduce, and not added in another assay, being the same but for the lack of the presec of the candidate.

20 PR0233 polypeptitcs and portions thereof which have homology to reductase may also be useful for in vim therapeutic purposes, as welas for various other applications. Ric identification of novel redutase proteins and related molecules may be relevant to a number of human disorders such as inflammatory disease. organ failure, atherosclerosis, cardiac injury. infertility. birth defcts, premature agg AIDS, cancer, diabetic complications and nwtations in general. Given that oxygen free radicals and antioxidants appear to play important roles in a number of disease processes, the identification of new reductase proteins and reductase-likc molecules is of special imupotance in diat such proteins may serve as potential therapeutics for a variety of different human disorders. Such polypeptides ~25 may also play important roles in biotchwnlogical and medical research, as well as various industrial applications.

As a result, there is particular scientific and medical interest in new molecules, such as PR0233.

The PR0223 polypcpuides of the present invention which exhibit serine carboxypeptidease activity may be employed in vim for therapeutic purposes as well as for in Wiro purposes. Those of ordinary skil in the art will well know how to employ PR0223 polypeptides for such uses.

PR0235 polypeptides and portions thereof which may be involved in cell adesion ame also useful for in viv thcrautic purposes. as well as for various in Wo, applications. In addition, PR0235 polypeptides and portions thereof may have therapeutic applications in disease states which involve cell adhesion. Given the physiological importance of cell adhesion mechanisms in vivo efforts are currently being undler taken to identify new, native proteins which are involved in cell adhesion. Therefore. peptides having homology to plexin are of particular interest to the scientific and medical communities.

Beccause the PR0236 and PR0262 polypeptides disclosed herein are hornologous to various known IPgalactosidase proteins, the PR0236 and PR0262 polypeptides disclosed herein will find use in conjugates of monoclonal antibodies and the polypepide for specific illing of tnror cells by generation of active drug from a galctosylatd prodrug (ecg.. t geneartionof ShrrWii from dhe prodrug P-D-gahctosyl-S-fluorouuidinc). The PR0236 and PR0262 polypeptides disclosed herein may also find various use both in vima and in vitro, wherein those uses will be similar or identical to uses; for which P-galactosidase proteins are now employed. Those of ordinary skill in the art will well know how to employ PR0236 and PR0262 polypeptides for such uses.

PR0239 polypeptides and portions thereof which have homology to densin may also be useful for in vw therapeutic pwpoes as wel as for various i Wvin applications. In addition, PR0239 polypeptides and portions thereof may have therapeuc applications in disease states which involve synaptic mechanisms. regeneration or cell adhesion.

Given the physiological impotamc of synaptic processes. regeneration and cell adhesion mechanism in v, efforts are currently being wale takcen to identify new, native proteins which are involved in synaptic machinery and cell adhesion. Thcemfore, pepfides having homology to densin are of particular interest to the scientific and medical The PR0260 plypeptides described herein can be used in assays to determine their relatio to rucosidase.

In particular, the PR0260 polypeptides can be used in assays In determining their ability to remove fucose or other sugar residues from proteoglycans. The PR0260 polypeptides can be assayed to determine if they have any funcional or locational similarilies a fixosidase. The PR0260 polypeptides can then be used to regulate the systems in which they are integral.

PR0263 can be used in assays wherein CD44 antigen is generally used to determine PRO263 activity **relative to that of CD44. The results can be used accordingly.

PR0270 polypeptides. and portions thereof which efect reduction-oxidation (redox) state may also be usefu for in wvo therapeutic purposes, as well as for various in Won applications. More specifically, PRO270 polypeptides; may affect the expression of a large variety of genes thought to be involved in thre pathogenesis of AIDS, cancer, atherosclerosis. diabetic complications and in pathological conditions involving orodative stress such as stroke and inflammation. In addition, PR0270 polypptides and portions theeo may affect the expression of a genes which have a role in apoptosis. Therefore, peptides having homology to thioredoxin are partictlarly desirable to the scienitific and *25 medical communities.

PR0272 polypeptdes ani portions *tdcfwhich possess the ability to bind calcium may also have numerous In Wvo therapeutic rusm as wvell as various in vir applications. Therefore. peptides having homology to reticulocalbin are particularly desirable. Those with ordinary skill in the art will know how to emiploy PR0272 polypeptides and portions thereof for such purposes.

PR0294 polypeptides and portions theref which have homology to collagen may also be usefu for in vivo therapeutic purposes, as well as for various other applications. The identification of novel collagens and collage-like molecules may have relevance to a number of human disorders. Thus, the identification of new collagens and collage-lice molecules is of special importance in that such proteins my serve as potential therapeutics for a variety of different human disorders. Such polypeptides may also play important roles in biotechnological and medical research as well as various industrial applications. Given the large numbcr of uses for collagen, there is substantial interest in polypeptides with homology to the collagen molecule.

107 PR0295 polypeptides and portions thereof which have hoology to integrin may also be useful for in vivo therapeutic purposes, as well as for various other application. The identification of novel integrins and integrin-like nolecules may have relevance to a nunmer of human disorders such as modulating the binding or activity of cells of the immune systen.

Thus, the identification of new integrins and integrin-like-molecules is of special inportance in that such proteins may serve as potential therapeutics for a variety of different human disorders. Such polypeptides may also play inportant roles in biotechnological and medical research as well as various industrial applications. As a result, there is particular scientific and nedical interest in new molecules, such as PRO295.

S..As the PRO293 polypeptide is clearly a leucine rich repeat polypeptide hnmologue, the peptide can be used in all applications that the known NLRR-1 and NLRR-2 polypeptides are used. The activity can be cacpared between these peptides and thus applied accordingly.

nThe PRO247 polypeptides described herein can be used in assays in which densin is used to determine the activity of PR0247 relative to densin or these other proteins. The results can be used accordingly in diagnostics and/or therapeutic applications with PRO247.

20 PR0302, PR0303, PRO304, PRO307 and PRO343 polypeptides of the present invention which possess protease activity may be enployed both in vivo for therapeutic purposes and in vitro. Those of ordinary skill in the art will well know how to enploy the PR0302, PRO303, PRO304, PR0307 and PRO343 polypeptides of the present invention for such purposes.

2 5 PR0328 polypeptides and portions thereof which have honology to GLIP and CRISP may also be useful for in vivo therapeutic purposes, as well as for various other applications. The identification of novel GLIP and CRISP-like nolecules may have relevance to a number of human disorders which involve transcriptional regulation or are over expressed in human tumors. Thus, the identification of new GLIP and CRISP-like nolecules is of special importance in that such proteins may serve as potential therapeutics for a variety of different human disorders. Such polypeptides may also play inportant roles in biotechnological and medical research as well as in various industrial applications. As a result, there is particular scientific and medical interest in new nolecules, such as PRO328.

Uses for PRO335, PRO331 or PRO326 including uses in ccupetitive assays with LIG-1, ALS and decorin to determine their relative activities.

The results can be used accordingly. PR0335, PR0331 or PR0326 can also be 107a used in assays where LIG-l wold be used to determine if the same effects are incurred.

PR0332 contains GAG3 repeat (GKEK) at amino acid positions 625- 628 in Fig. 108 (SE)2 ID NO):309). Slippage in such repeats can be associated with hurman disease. Accordingly, PRO332 can use useful f or the treaunet of such disease conditions by' gene therapy, i e. by introduction of a gene containing the correct GKEX sequence notif.

other uses of PR0334 include use in assays in which fibrillin or fihulin would be used to determxine the relative activity of PR0334 to fibrillin or fibilin. In particular, PR0334 can be used in assays which require the nechanim inparted by epidenmn.l growth factor repeats.

Native PR0346 (SE)Q MD NO:319) has a Blast score of 230, Corresponding to 27% hcuology between amino acid residues 21 to 343 with residues 35 to 1040 CG46i UMAN, a carcinoerbryonic antigen cgm6 precursor.

This hcxnlogy region include nearly all but 2 N-terminal extracellular *dcmnain residues, including an inmunoglobuilin siuperfamnily harlogy at residues 148 to 339 of PR0346 in additicon to several transn nrae.

residues (340-343)- Carcinoemixyonic antigen precursor, as explained in The Background is a tumnor-specific antigen.

and as such. is a recognized markcr and therapeutic target for the diagnosis and treatmcot of colon cancr The ciqiression of tumor-specific antigens is often associated with dhe progression of neoplastic tisse disorders. Native PR0346 (SEQ ID N0:3A0 and P -W06974. a huiman cardnembryonic antigen CEA-d have a Blast score of 224 and homology of 28% between residues 2 to 343 and 67 to 342, respectively. This homology iocludes the entire extracellujar domain residues of native PR0346. minus the initiator methionime (residues 2 to 19) well as several bransmembrane residues (340-343).

PRO268 polypeptides which have protein disulfide isomerase activity will be useful for many applications where protein dsulfide isomerase activity is desirable including, for example, for use in promoting proper disulflde bond formation in recombinantly produced proteins so as to increase die yield of correctly folded protein. Those of ordinary skill tn die art will readily know how to employ such PR0268 polypeptides for such purposes.

PR0330 potypepddes of the present invention which possess biological activity related to that of the prolyl :4-hydroxylase alpha subunit protein may be employed both in vivo for therapeutic purposes and in wiro. Those of ordinary skill in the art will well know how to employ the PR0330 polypetides of dft present invention for such purposes.

Anti-PRO PolyVetitide Antibodies The present invention further provides anti-PRO polypeptide antibodies. Exemplary antibodies include :polyclozial, monoclonal. humanized, bispecific. and beteroconjugate antibodies.

A. Polvelonni Antlbolies The anni-PRO polypetde antibodies may comprise polycial antibodies. Methods of preparing polyclonal antibodies are known to the skilled artisan. Polyclonal antibodies can be raised in a mammal, for example, by one or more injections of an immunizing agent and, if desired, an adjuvant. Typically, the immunizing agent and/or adjuvant will be injected in the manunal by multriple subcanreous; or intraperitoneal injections. The immunizing agent may include the PRO polypeptide or a fussion protein thereof. It may be useful to conjugate the immunizing agent to a protein known to be immunogenic in the mammual being immunized. Examples of such immunogenic proteins iclude but ame not limited to keyhole limpet hemocyanin, serumn albumin, bovine thyroglobulin, and soybean trypsin inhibitor. Examples of adjuvants which may be employed include Freund's complete adjuvanu and MPL-TDM adjuvant (monophosphoryt Lipid A, synthetic trehalose dicorynomycolate). T'he imimnzation protocol may be selected by one skilled in the art without undue experimentation.

B. Monoclonal Antibodie T1he anni-PRO polypptder andiboies may, alternatively, be monoclonal antibodies. Monoclonal antibodies may be prepared using hybrdoea miethods, such as those: described by Kohler and Milstein, Nature, W:495 (1975).

In a bybridoma. method, a mouse, hamster, or other approprate host animal, is typically immunized with an iunninizing agent to elicit lymphocytes diat produce or are capable of producing antibodies that will specifically bind to the immunizing agent. Alternatvely, the lymphocytes may be immunized in viro.

The immunizing agent will typically inchude the PRO polypeptide of interest or a fusion protein thereof.

Generally, eithe peripheral blood lymphocytes ("PBLs') are used if cells of human origin are desired, or spleen cells or lymph node cells are used if non-human mammalia sources art desired. The lymphocytes are then fused with an immxormlized cell line using a suitable fusing agent, such as polyethylene glycol. to form a hybnidoma cell [Goding.

Monoclonal Antibodies- Principles AMd Pradic, Academic Press. (1986) pp. 5%-103). Immortaized cell lines are usually transformed manualian cells. particularly niyclonsa cells of rodezi, bovine and human origin. Usually, rat or maze myekxua cell Ines are employed. The hybridoma cells may be cultured in a suitable culture: medium that preferably contains one or morm substances that inhibit the growth or survival of the unfused, immortalized cells.

For example, if the parenta cells lack the enzyme hypoxanthie guanine phosphoribosyl transferase (HGPRT or HPRT). the culture medium for the hybridomas typically will include hypoxanthine. aininopterin. and thyrnidine ('HAT medium"). which substances prevent the growth of HGPRT-deficient cells.

Preferred immortalized cell lines are those that fuse efficiently, support stable high level expression of antibody by the selected antibody-producing cells, and art sensitive to a medium such as HAT medium. Mome preferred immortalized cell line are munine myeloma lines, which can be obtained, for instance, from the Salk Institute Cell Distribution Center, San Diego, California and dhe American Type Culture Collection, Rockville, Marylandt. Human myclama andI mosise-hImon hmtromyelosna cell lines also have been described for the production of hulman monoclonal antibodies (Kozbor, J. Inimwzol., .ln:3001 (1994); Brodeur et al., Monoclonal Antibody Production Technqueu and Applications, Marcel Dekker. Inc., New York, (1987) pp. 51463).

The culture medium in which the hybridoma cells are cultured can then be assayed for the presence of monoclonal antibodies directed against the PRO polypeptide of interest. Preferably, the binding specificity of monoclonal antibodies produced by the hybridoma cells is determined by imrnunoprecipitation or by an in %4tro :bindling assay, such as A' m oevvsay (RIA) or enzyme-linked immunoabsorbent assay (ELISA). Such techniques and assays; are known in the art. The binding affinity of the monoclonal1 antibody can, for example, be determined by the Scatchard analysis of Munson and Pollard, Anal. Biochem.. WZ:220 (1980).

After the desired hybridoma cells ame ientifled. the clams may be subcloned by Uimiting dilution procedures and grown by standard methods [Goding, IpW Suitable culture amdia for this purpose include, for example.

Dulbecco's Modified Eagle's Medium and! RPMI-1640 medium. Alternatively, the hybridoma cells may be grown in vivo as ascites in a mamml.

The monoclonal antibodies secreted by the subelones may be isolated or purified from the culture medium or ascites fluid by conventional immunoglobulin purification procedures such as, for example, protein A-Sepharose, hydroxylapatite chromatography, gel electrophoresis, dialysis, or affinity chrormatgraphy.

The monoclonal antibodies may also be made by recombinant DNA methods, such as those described in US. Patent No. 4.816,567. DNA encoding the monoclonal antibodies of the invention can be readily isolated and sequenced using conventional procedures by using oigornucleotide probes that are capable of binding specifically to geme encoding dhe heavy and light chains of murine antibodies). The hybridoma cells of the invention serve as a preferred source of such DNA. Once isolatd, the DNA may be placed into expression vectors, Which are then transfected into host cells such as simian COS cells. Chinese hamnster ovary (CHO) cells, or mycloina cells that do not otherwise produce immn~oglobutin protein, to obtain the synthesis of monoclonal antibodies in the recoatinan hosn cells. 7The DNA also may be =04led for example. by substituting the coding sequence for human heavy and Light chain constant domains in place of the homologous murine sequences Patent No. 4.816,567; Morrison et al.. GWxi or by covalently joining to the immimoglobulin coding sequence all or part of the coding sequence for a non-ifmmunoglobulin polypeptide.- Such a non-inmnomglobulin polypeptidc can be substituted for die constant domains of an antibody of the invention. or can be substituted for the variable dolmin of one antigencombining site of an antibody of the invention to create a chimeric bivalent antibody.

The antibodies may be monovalent antibodies. Methods for preparig monovalent antibodies are well known in the art. For example, one method involves recombinant expression of iznmunoglobuLin light chain and modified heavy chain. the heavy chain is truncated generally at any point in the Fe region so as to prevent heavy chain crosslinking. Alternatively, the relevant cysteine residues AMe substituted with another amino acid residue or are deleted so as to prevent cr=ssling.

In vitro methods are also suitable for preparing morxovalent antibodies. Digestion of antibodies to produce fragments thereof. particularly. Fab fragments, can be accomplished using routine techniques known in the art.

C. Humanized Antibodies The anni-PRO polypeptide antibodies of the invention may further comprise humanized antibodies or human antibodies. Humanized forms of non-human muirinc) antibodies are chimeric immunoglobulins, imuolobulin chains or fragments thereof (such as Fv, Fab. Fab'. F(abh) or other antigen-binding subsequences of antibodies) which contain minimal scquenoc derived from non-human imnoglobulin. Husmaized antibodies include human immunoglobulins (recipient antibody) in which residues from a complementary determining region (CDR) of doe recipient are replaced by residues from a CDR of a non-human species (donor antibody) such as mouse, rat or rabbit having the desired specificity, affinity and capacity. In some: instances. Fv framework residues of the human immunoglobulin are replaced by corresponding non-human residues. Humnized antibodies may also comprise residues which are found neither in the recipient antibody nor in the imported CDR or framework sequence. In general, the humanized antibody will Comprise substantially all of at least one, and typically two.

variable domains, in which all or substantially all of the CDR regions correspond to those of a non-human irmmunoglobulin and all or substantially all of the FR regions are those of a human irnmunoglobulin consensus sequencc. 'The humanized antibody optimally also will comprise at least a portion of an immunoglobulin constant region typically that of a human imnmoglobulin (Jones d al.. Nature, 2U: S227525 (1986); Riedirnan a Nature, IU323-329 (1988); and Presta. Or. Op. &,ruc. Biol. 2593-596 (I99)].

Methods for non-umman antibodics ame well known in doe arn. Generally, a humanized antibody has one or moeammo acid resklues inroduced into it from a souce which is non-human. These non-hwnan amino acid residues are often referred to as 'import" residues, which are typically taken from an 'import* variable domain.

Hur-niration can be essentially performed following doe nethod of Winrer and co-workers [Jones er al., Nature, n: 522-525 (1986); lRechmazin er aL, Nature. j2:323-327 (1988); Vcrhoeyen ef al, Science. ZU2:L534-1536 (1988)], by substituting rodent CD~s or CDR sequenees for the corresponding sequences of a human antibody. Accordingly, such 'humanized' antibodies are chimneric antibodies Patent No. 4,816,567). wherein substantially less than an intaet huma variable domain has been substituted by the orresponding sequence from a non-human species. In Practice, humanized antibodies ame typically human antibodies in which some CDR residues andl Possibly sonic FR residues are substituted by residues from analogous sites in rodent antibodies.

Huwn antibodies can also be produced using various taclmiques known in the art. including phage display libraries (Hoogenboom and Winter, Mot. Not., 2:391 (1991); Marks et al., J. Mol. BioL,f22581 (1991)]. Ile techniques of Cote ef al, and Boerner et ame also available for the preparation of human monoclonal antibodies (cote er aL, Morodsmul Anaiwdes arid Cowcer Therapy, AlaR. LIsa, p. 77 (1985) and Boerner etat J. ImmunoL, 11=16-95 (1991)].

D. Bisnecific Antibodies Bispecific antibodies are mnonoclonal, preferably human or humanized, antibodies that have binding specificities for at least two different antigens. In the present case, one of the binding specificities is for the PRO po4Vypptde the other one is for any other antigen, and preferably for a ceUI-surface protein or receptor or receptor subunit.

Methods for making bispecific antibodies are known in the art. Traditionally, the recombinant production of bispecific antibodies is based on the co-expression of two immunoglobulin heavy-chinlight-chain pairs, where 15 the two heavy chains have different specificities [Milstein and Cuello, Nature, 10:37-539 (1983)]. Because of the random assortment of immunoglobalin heavy and light chains, these hybridomnas (quadromas) produce a potential nixtuic of tco different antibody molecules. of which only one has the correct bispecific structure. The purification of the correct molecule is usually accomplished by affinity chromatography steps. Similar procedures are disclosed in WO 93/08829, published 13 May 1993, and in Traunedaer et EMBlO a3655-3659 (1991).

Antibody variable domains with the desired binding specificities (antibody-antigen combining sites) can be :fused to immnoglobulin constant domain sequences. The fusion preferably is with an immuunoglobulin heavy-chain consan domain, comprising at least part of the hinge, CM2, and CH3 regions. It is preferred to have the first heavychin constan region (CHIi) containing the site necessary for light-chain binding present in at least one of the fusions.

DNAs encoding the iznzmmoglobulin heavy-chain fusions and, if desired, the inimunoglobulin light chain, are inserted 2.5 into separate expression vectors. and are co-transfected into a suitable host organism. For further details of generating bispecific antibodies see, for example, Suresh ei at., Methods in Emymology, 121:210 (1986).

Heteraconjuate antibodies are also within the scope of the present invention. Heteroconjugate antibodies are composed of two covalendy joined antibodies. Such antibodies have, for example, beeni proposed to target imue system cells to unwanted cells Patent No. 4,676,801, and for tretment of H1V infection (WO 91/00360; WO 92/2=073; EP 03089]. It is contemplated that the antibodies may be prepared in vitro using known mecthods in synthetic protein chemistry, including those involving crossfinking agents. For example, immunotoxins may be consmucted using a disulfide exchange reaction or by forming a thioedjer bond. Examples of suitable reagents for this purpose incluh imincilate and methyl-4-mercaptobutyricidate and those disclosed, for example, in U.S.

Patent No. 4.676.980.

S6. Uses for Anti-Pro Polvuentide Antlbodies The and-PRO polypeptide antibodies of the invention have various utilities. For example, anti-PRO polypcptide antibodies may be used in diagnostic assays for a PRO polypeptide, detecting its expression in Specific cells, tissues, Or serum. Various diagnostic assay techniques known in the art may be used. such as competitive binding asays, direct or indirect sandwich assays and iminpreaipitation assays conducted in either heterogeneous or homogeneous phases [Zola. Monoclonal Antibodies: A Manual of Tchnques, tRC Press, Inc.

(1987) pp. 147-158). The antibodies used in the diagnostic assays can be labeled with a detectable moiety. The detectable moiety should be capable of producing. either directly or indirectly. a detectable signal. For example, die detectable moiety may be a radioisotope, such as 'Hi. m~ P.1 S. o?2' 1. a fluorescent or chetniluinescent compound, such as fluorescein isothiocyanate, rhodamine. or luciferin, or an enzyme, such as alkl~ine phosphatase, beta-galacosidase or horseradish peroxidase. Any method known in the ant for coniJugating the antibody to the detectable moiety may be employed, including those methods described by Hunter et at., Nature. 144:945 (1962); David ea al.. Biochemiry, 13:1014 (1974); Pain d aL, J. Immwwol. Meth.. 4:219 (1981); and Nygren, J.

llbochem. wid Cytochem., 20:407 (1982).

Anti-PRO polypeptide antibodies also arc useful for fth affinity purification of PRO polypepuade from recombinant cell culture or natural sources. In this process, the antibodies against the PRO polypepuide are immobiized on a suitable support. such a Sephadex resin or filter paper, using methods well known in the art. 1Te immobilied antibody then is contacted with a sample containing the PRO polypeptide to be purifie, and thereafter the support is washed with a suitable solvent that will remove substantially all the material in the sample excepit the PRO polypeptide, which is bound to the immobilized antibody. Finally, the support is washed with another suitable e 20 solvent that will release the PRO polypeptide from the antibody.

With regard to PRO211 and PR0217. therapeutic indications include disorders associated with the :5 preservation and mintenance of gastrointestinal mucosa and the repair of acute and chronic mucosai lesions awierocolitis, Zallinger-Hlisci syndlrome, gastrointestinal ulceration and congenital microvillus atrophy), skin diseases associated with abnormal keratinocyte differentiation psoriasis. epithelial cancers such as lung squamous cell carcinoma, epiderrnoid carcinoma of the vulva and glioias.

With regard to anti-PROlS? antibodies, FGF4 has been implicated in cellular differentiation and embryogenesis, including the patterning which appears during limb formation. FGF-S and the PRO 187 molecules of the invention therefore are likely to have potent effects on cell growth and development Diseases which relate to cellular growth and differentiation art therefore suitable targets for therapeutics based on functionality similar to FGF48. For examle, diseases related to growth or survival of nerve cells including Parkinson's disease, AIheaimer's disease, ALS, necuropathies. Additionally, disease related to uncontrolled cell growth, cancer, would also be expected therapeutic targets.

Native PR0533 is a 216 amino acid polypeptide of which residues 1-22 are the signal sequence. Residues 3 t0216 have a Blast score of 509, corresponding to 53% homology to fibroblast growth factor. At thenmaceotide level, DNA47412, the EST from which PCR oligos were generated to isolate the full length DNA49435-1219, has been observed to map to lI piS. Sequecme homology to the I lpiS locus would indicate that PR0533 may have utility in the treatment of Usher Syndromic or Atrophia areata.

As meoned previously. fibroblast growth factors can act upon cells in both a mitogenic and non-mitogenic mnne. These factors am mmuogenic for a wide variety of norml diploid miesoderm-deuived and neural crest-derived els, 6inc granelosa cells. adrenal cortAic cells. dirondracytes, myobLasts, corneal and vascular endodcdial cells (bovine or human), vascular smooth muscle cells, lens, retina and prostatic epithelial cells, oligodendrocytes, astracytes, chmondocytes, miyoblasts and osteoblasts. Antibodies to these factors can be generated to modulate such effect.

Non-mitogenic actions of fibroblast growth factors include promotion of cell migration into a wound area (diemotaxis), initiation of new blood vessel formulation (axigiogcesis), modulation of nerve regeneration and survival (neurotrophism). modulation of endocrine functions. and stimulation or suppression of specific cellular protein expression, extacllular matrix production and call survival. Baird. A. Doblen, Handbook of Exrp. Phnnacol.

221l): 369-418 (1990). These properties provide a basis for using fibroblast growth factors in therapeutic approaches to accelerate wound hecaling, nerve repair, collateral blood vessel formation, and dhe lim. For example, fibroblast growth factors, have been suggested to minimize myocardium damage In heant disease and surgery (U.S.P.

4,378,437). Antibodies to these factors can be generated to modulate such cifects.

Therapeutic indications for PR0214 polypeptides include disorders associated with the preservation and 15 maintenance of gastrointestinal mucosa and the repair of acute and chronic mucosal lesions enterocolitis, Zollinger-Efison syndiromie. gastrointestinal ulceration and congenital microvillus atrophy). skin diseases associated with abnormal keratinocyre differentiation psoriasis, epithelial cancers such as king squamous cell carcinomna.

epidermoid carcinoma of the vulva and gliomas.

0~0* Andi-PRO317 antibodies find use in anti-tumnor indications if they are angiostatic, or in coronary ischemic indications if they are angiogenic.

00 Native PRO301 (SEQ MD NO:119) has a Blast score of 246 and 30% homology at residues 24 to 282 of Fig.

44 with A33 HUMAN, an A33 antigen precursor. A33 antigen precursor, as explained in the Background is a *000 annor-spocific antigen. and as such, is a recognized marker and therapeutic target for the diagnosis and tieatmnit of colon cancer. The expression of tumor-specific antigens is often associated with the progression of neoplastic tissue disorders. Native PRO301 (SEQ ED NO:119) and A33 -HUMAN also show a Blast score of 245 and 30 homology at residues 21 to 282 of Fig. 44 with A33 HUMAN, the variation dependent upon how spaces are inserted into t comparod sequne. Native PRO301 (SEQ WD NO:119) also has a Blast score of 165 and 29% homology at residues to 255 of Fig. 44 with HS46KDA a human coxsackie and adenovirus receptor protein, also -known as cell sufac protein HCAR. This region of PRO301 also shows a similar Blast score and homology with HSU90716_1.

Expression of such proteins is usually associated with viral infection and therapeutics for the prevention of such infection may be accordingly conceived. Accordingly, antibodies to the above identified antigens and receptors have therapeutic potential as diagnostic and ftratment techniques.

Therapeutic uses for the PR0234 polypeptides of the inventon includes treatments associated with leukocyte homing or the interaction between leukocytes and the endotheliun during an inflammatory rese. Examples include asthma. rheumatoid arthritis, psoriasis and multiple sclerosis.

Cancr-associated or specific antigens permit the creation of tumor or cancer specific monoclonal antibodies (mAbs) which are specihic wo suchi tumar antigens. Such mAbs, which can distinguish between normal and cancerous cells are useful in the diagnosis. prognosis and treatmient of the disease.

Cancer specific monoclonal antibodies (niAbs) which are specific to tumnor antigens. Such mjAbs, which can distinguish between normal and cancerous cells ame useful in the diagnosis. prognosis and treatment of the disease. Particular antigens are kown to be associate tidi noopbastic diseases, such as colorctal and breast cancer.

Since colon cancer is a widespread disease, early diagnosis and treanncn is an imtportant medi cal goal. Diagnosis and trcatment of cancer can be implenmted using mnonoclonal antibodies (mAbs) specific therefore having Mlorescent, nuclear mnagnetic or radioactive tags. Rtadioactive genes, toxins and/or drug tagged mtAbs can be used for treatment in situ with moinimal patient description.

The following examples arc offered for illustrative purposes only. and are not intended to limnit the scope of the present invention in any way.

Al] patent and literature references cited in the present specification are hereby incorporated by reference in their entirety.

V060 0 EXAMPLES Commercially available reagents referred to in the examples were used according to manufacture's *000 0 a 15 instrcton unless otherwise indicated. The source of those cells identified in the following examples, andl throughout the specification, by ATCC accession mnimbers is the American Type Culture Collection, Rockyille. Maryland.

0 E: XAMPLE 1: Extracellular Domain liomology Screenrn to IdentfX Novel Polvpepddes and cDNA Encoding The extracellu domain (ECD) sequences (including the secretion signal sequence, if any) from about 950 imkown sectd proteins from die Swiss-Pint public database were used to search EST databases. The EST databases included public databases Dayboff, GenBank), and proprietary databases LIFEEQ'- Incyte Pharmaceuticals, Palo Alto. CA). The search was performed using the computer program BLAST or BEAMT (AltschulI, and Gish, Methods in EuZyMlogv M: 460-80 (1996); hitp://bast.wustltcdu/blat/README.hnnl) as a comparison of the ECD protein sequences to a 6 frame translation of the EST sequences. Those comparisons with Blast soeof 70 (or in sonic cases 90) or greater that did not encode known proteins were clustered and assembled into consensus DNA sequences with the program "phrap' (Phil Green, University of Washington. Seattle, WA; (tp/Ibozcma.ma ahingwide4,h pp.docs/phrpinl).

Using this extraceliular domain homology screen, consensus DNA sequences were assembled relative to the other identified EST sequeoces. In addition, the consensus DNA sequences obtained were often (but not always) extended using repeated cycles of BLAST and phrap to extend the consensus sequence as far as possible using the sources of EST sequences discussed above.

Based upon the consensus sequences obtaine as described above. oligonucleotides were then synthesized and used to identify by PCR a eDNA library that contained the sequence of interest and for use arprobes to isolate a clone of the full-length coding sequence for a PRO polypeptide. Forward and reverse PCR primers generally range from 20 to 30 nucleocides; and ame often designed to give a PCR product of about I00-1000 bp in length. The probe sequences are typically 40-55 bp in length. In some cases, additional oLigonueleotles are synthesized when the consensus sequence is greater than about 1-1 .Skbp. In order to scre several libaries for a ful-length clone, DNA from the libraries was screened by PCR amplification. as per Ausubel et aL. Curn Protocons in Molectila Hiolgry. with the PCR primer pair. A positive library was then used to isolate clones encoding the gene of interest using the probe oligomicleodde and one of the primer pain.

The cDNA libraries used to isolate the cDNA cdonrs were constructed by standard mehods using commerialy available reagents such as those from Invitrogen. San Diego, CA. The cDNA was primed with oligo dT containing a Nod site. linked with blunt to Sall hemikinased adaptors, cleaved with Not!, sized appropriately by gel electrophoresis. and cloned in a defined orientaton into a suitable cloning vector (such as pRICE or pRKD; pRKSB is a precursor of pRKSD3 that does not contain the Sf1l site; see, Holmes et al.. Scie 22:1278-1280 (1991)) in the unique Xbol and Not! sites.

EXAMPLE 2: Isolation of cDNA Clones Encodin, PRO21I and PRO217 Consensus DNA sequences were assembled as described in Example 1 above and were designated as DNA2873 and DNA28760. respectively. Based on these consensus sequences, oligonucleotides wecre synthesized and used to identify by PCR a cDNA library that containedl the sequences of interest and for use as probes to isolate a clone of the full-length coding sequence for the PRO21 1 and PR0217 polypeptides. The libraries used to isolate DNA32292-1 131 and DNA33094-1 131 were fetal lung libraries.

cDNA clones were sequenced in their entirety. The entire nucleotide sequences of PRO21 1 (DNA32292- 1131; LUNQ1S5) and PR0217 (UNQI9l; DNA33094-1 13 1) are shown in Figure 1 (SEQ ID NO: 1) and Figure 3 (SEQ HD NO:3). respectively. The predicted polypeptides are 353 and 379 amino acid in length, respectively, with respective molecular weights of approximately 38.190 and 41.520 daltom.

The oligonucleotide sequences used in the above procedures were the following: .:28730.p (OLI 516) (SEQ ID NO:S) 5'-AGGGAGCACGGACAGTGTGCAGATGTGGACGAGTGCTCACrAGCA-3 28730.f (OUI 517) (SEQ ID NO:6) 5'-AGAGTGTATCTCTGGCTACGC-3' 29730.r (OU 5.18) (SEQ ID NO:7) S-TAAGTCCGGCACATTACAGGTC-3* 28760.p (OUI 617) (SEQ ID NO:g) 5-CCCACGATGTATGAATGGTGGACmGTGiTGACTCCGGrCTGCATC-3' 29760.f (OLI 618) (SEQ MD NO:9) 5'-AAAGACGCATCTGCGAGTGTCC-3* 29760.r (OUI 619) (SEQ WD NO: 5S-TGCTGAmICACACTGCTCTCCC-3- EAMPLE.3: Isolation of cDNA Clones Fucodjng Humnan PRO230 A consensus DNA sequence was assembled relative to the other identified EST sequences as described in Example I above, wherein the consensus sequence is designated herein as DNA30857. An EST proprietary to Genentech was employed in the consensus assembly. The EST is designated as DNA20088 &nM has the nucleotide sequence shown in Figure 7 (SEQ ID NO: 13).

Based on the DNA30957 conenss sequence. oligonixleotides were synthesized to ientify by PCR a cDNA library that contained the sequence ot interest and fior use as probes to isolate 2 Clone of the fulfl-length coding sequence fo~r FROM3.

A pair of PCR primers (forward and reverse) were synthesized: forward PCR *ire 5'-TFCGAGGCCTCTGAGAAGTGC3CCC.3* (SEQ MD NO:14) reverse[ SC rmr 5-GGCGGTATCTCTCTGGCCTCCC-3* (SEQ ID Additionally. a synthetic oligonucleotide hybridization probe was constructed from the consensus DNA30857 sequecwe which hid the following nuclodde sequence hybridization iprobc S'-TCTCCACAGCAGCGTGGCATC)CGATCGTGTC~ATCCATCCrGGG-.3- (SEQ ID NO: 16) :In order to screen several libraries for a source of a full-length clone. DNA from the libraries was screened by PCR umpifcation with the PCR primer pair identified above. A positive library was then used to isolate clones encodiztg the PR0230 gene using the probe oligonucleodde and one of the PCR primers.

RNA for construction of the cDNA libraries was isolated from human feta lung tissue. DNAwWrzdzg of the clones isolated as described above gave the full-length DNA sequence for PR0230 (herein designated as UNQ20)4 (DNA33223-IL36)) and the derived protein sequence for PROM3.

The entire nucleotide sequence of UNQ204 (DNA33223-l 136) is shown in Figure 5 (SEQ ID NOM1).

Clam UNQ2O4 (DNA332Z3-l 136) onins a shnge open reading f-rme with an apparent translational initiation site at nucleotide positions 100-103 and ending at the stop codon aS nucleotide positions 1501-1503 (FgureS5; SEQ ID NO: 11). Thke predicted polypeptide precursor is 467 amino acids long (Figure 6).

EAMPLE 4: Isolation of cDNA Clones Encoding Hutnan PRO232 A consensus DNA sequence was assembled relative to the other identified EST sequences as described in Example I above. wherein the consensus sequec is designated herein as DNA30935. Based on the DNA30935 consensus sequence, oligonucleotides were synthesized to identify by PCR a cDNA library that contained the sequence of initerest and for use as probes to isolate a clone of the full-length coding sequence for PROM3.

A pair at PCR primers (forward and reverse) were synthesized: forward PCR priner 5'-TGCTGTGiCTACTCCTGCAAAGCCC-3' (SEQ ID NO:19) reverse PCR lorinier S'-TG;CACAAGTCGGTGTCACAGCACG-3' (SEQ ID Additionally, a synthetic oligonucleotide hybridization probe was constructed from the consensus DNA30935 sequence which had the following nucleotide sequence 5-AGCAACGAGGACGCCTGCAGGTGGAGAACT'GCACCCAGCTGGG-3' (SEQ ID NO:21) In order to scrm several libraries for a source of a full-length clone, DNA from the libraries was screened by PCR antplificationi with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PR0232 geme using the probe oligonucleotide and one of the PCR primers.

RNA for construction of the cDNA libraries was isolated from human fietal kidney tissue.

DNA sequencing of the clones isolated as described above gave die ful-length DNA sequence for PR0232 [herein designated as UNQ2D6 (DNA34435-1140)I and the derived protein sequne for PROM3.

The entire nucleodde sequence of UNQ206 (DNA34435-1140) is shown in Figure 9 (SEQ ID) NO: 17).

Clone UNq2O6 (DNA34435-I 140) contains a single open reading frame with an apparent translational initiation site at muclcolid positions 17-19 and ending at the stop codon at nucleotide positions 359-361 (Fig. 8;SEQ ID NO: 17).

The predicted polypeptide precursor is 114 amino acids long (Fig. Clone UNQ2O6 (DNA3443S-1140) has been deposited with ATCC on September 16. 1997 and is assigned ATCC deposit no. ATCC 209250.

Analysis of the am;n acid sequence of the full-length PR0232 suggests that it possesses 35 sequence identity with a stem cell surface antigen from Gallus gallus.

EAMPLE Isolation of cDNA Clones ncoding PROl 57 A proprietary expressed sequece tag (ESTl) DNA datbas (LIESEQ~ h Incyte Pharmaceuticals, Palo Alto, CA) was searched and an EST (#843 193) was 6rdentiod which showed homnology to fibroblast growth factor (FGF-8) also known as androgen-uince gtowt sactor. nRNA was isolated from human feta lung tisse using reagents and froms Invitrogmn San Diego, CA (Fast Track The cDNA libraries used to isolate the cDNA clones were constructed by swadard methiods using commercially available reagents lovitrogen. San Diego, CA. Life Technologies, Gaithersborg, MD). The eDNA was primed with oligo dT containing a Notd site, linked with blunt to Sall bemikinased adaptors, cleaved with Nod, sized appropriately by gel eloctropharesis, and cloned in a definted orientation into the cloning vector pRK5D using reagents and protocols from Wie Technologies, Gaithersburg. MD (Super Script Plasid System). The double-standled cDNA was sized to greater than 1000 bp and the SallINotl linkered cDNA was cloned into XboIINorI cleaved vector. pRK5D is a cloning vector that has an sp6 tranzscription initiation site followed by an Sfil restriction enzyme site preceding the XhoI/Notl cDNA cloning sites.

Several libraries from various tisse sources were screened by PCR amplification with the following oligowuceotide probes: IN843193.f (011315) (SEQ ID NO:24) 5-CAGTACGTGAGGGACCAGGGCGCCATGA..3' IN843193.r (OLI 317) (SEQ ID 5'-CCGGTGACCGiCACGTGCTGCCA.3' A positive library was then used to isolate clones encoding the PRO187 gene using oue of the above oigonucleocides and the following oligonucleotide probe: JN843l93.p (OLI 316) (SEQ ID NO:26) A cDNA clone was sequenced in entirety. The enfire nuclodde sequence of PROWS (DNA27864-1155) is shown in Figure 10 (SEQ ID 14022). Clone DNA27864-1 155 contains a single open readirg frame with an apparent translational initiation site at snxdeotide position I (Figue 10. SEQ ID The predicted polypeptide precursor is 205 amino acids long. Clone DNA27864-1 155 has been deposited with the ATCC (designation: DNA27864-1 155) and is assigned ATCC deposit no. ATCC 209375.

Based on a BLAST and PastA seqenc alpue analysis (using the AUIGN computer program) of the fulllength seqe .e the PRO187 polypeptide shows 74% amino ac id sequence identity (Blast score 310) to human fibroblast growth factor-8 (androgen-induced growth factor).

EXAMPL.E 6: Isolation of cDNA Clones EncodIni PRO265 A consensus DNA sequence was assembled relative to other EST sequences as descrlcd in Example 1 above using phrap. This consensus sequence is herein designated DNA33679. Based on the DNA33679 consensus sequence, oligomicleoddes were synthesized: 1) to identify by PCR a cDNA library that contained the sequence of interest. and 2) for use as probes to isolate a clone of fth fulfl4eogth coding sequence for PROW6.

PCR primers (two forward and one reverse) were synthesized: forward PCR primer A- S-CGGrCACCrGTATGGCAACC-3' (SEQ ID 80:29); forward PCR primier Be 5'-GCAGGACAACCAGATAAACCAC.3' (SEQ ID N0:30); 'evene PR Vimer 5'-ACGCAGATrGAGMGGCGTC.3* (SEQ ID 140:31) Additionally, a synthetic oligonuceotide hybridization probe was constructed from the consensus DNA33679 sequence which had die following nucleotide sequence hyridizaio~n Robe 5*7CCGCGT~ CCGT~TAG~GAACICC3 (SEQ ID 80:32) In order to screen several libraries for a source of a ful-length clone, DNA from the libraries was screened by PCR amplification with PCR primer pairs identified above. A positive library Was then used to isolate clones encoding the PRO265 geme using the probe oligonucleotide and one of die PCR primers.

20 R.NA for construction of the cdNA libraries was isolated from human a fetal brain library.

DNA sequencing of the clones isolated as described above gave the full-length DNA sequence for PR0265 (herein designated as UNQ232 (DNA36350-1 159] (SEQ ID N0:27) and the derived protein sequence for PR0265.

The entire mucleotide sequence of UNQ232 (DNA36350-11S8) is shown in Figure 12 (SEQ ID 80:27).

Clone UJNQ232 (DNA36350-1 158) contains a single open reading frame with an apparent translational initiation site at nucleotide positions 352-354 and ending at the stop codon at positions 2332-2334 (Figure 12). The predicted o polypqttde precursor is 660 amin acids long (Figure 13). Clone UNQ232 (DNA36350-1158) has been deposited wth ATCC and is assigned ATCC deposit no. ATCC 209378.

Analysis of the amino acid sequence of die ful-length PRO265 polypeptide suggests that portions of it possess significant homology to the fibromodulin and the fibromoduilin precursor, thereby indicating that PRO265 may be a novel member of the leucine rich repeat faily, particularly related to fibromodulin.

EXML 7: solation of erNA Clones Ecoding Humann PRO71 9 A consensus DNA sequence was assembled relative to other EST sequences using pdirap as described in Example 1 above. This consensus sequence is herein designated DNA28729. Based on the DNAWZ9 consensus sequmce, ollgonucleotides were synthesized: 1) to identify by PCR a cDNA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PROM 1.

A pair of PCR primers (forward and reverse) were: synthesizedi forward PCR pri=e S*-GTGACCCTGG7TGTGAATAcCtC.3* (SEQ ID reverse PCR iVriMUe 5S-ACAGCCATGGTCTATAGCTrGG-3- (SEQ [D NO:36) Additionally. a synthetic oligonucleotide hybridization probe was constructed from the consensus DNA28729 sequence which had tie following nucleotide sequence 3 (SEQ ID NO:37) In order 10 scr=t several libraries for a source of a full-length clone. DNA from the libraries was screened by PCR amplification with the PCR primer pair identified above. A positive library was then used to'isolate clones encoding the PR0219 gene using the probe oligonncleoddc and one of the 7CR primers.

RNA for construction of dfe cDNA libraries was isolated from human fetal idney tissue DNA sequcncirg of the clooms isolated as described above gave die MIl-length DNA sequcm for pR0219 [therein designated as UNQ193 (DNA32290-1164)I (SEQ ID NO:33) and die derived protein sequence for PROW1.

flie entire raulotde seqiar of UNQ193 (DNA32290-1164) is shown in Figure 14A-B (SEQ ID NO:33).

Clone UNQ 193 (DNA32290-1 164) contains a single open reading frame with an apparent translationAl initiation site .15 at naclode positions 204-206 and ending at tde stop codon at nucleotide positions 2949-2951 (Figures 14A-B). The predicted polypeptidc precursor is 915 amino acids long (Figure 15). Clone UJNQ193 (DNA32290-1 164) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 20984.

Analysis of die amino acid sequence of the full-length PR0219 polypeptide suggests that portions of it possess significant homology to the mouse and human inatrilin-2 precursor polypeptides.

EXAMPELE 8: Isolation of cDNA Clones jtcndinw Mn PROM4 A consensus DNA sequence was assembled relative to other EST sequences using phrap as described in Example I above. This consensus sequence is herein designated DNA30955. Based on the DNA30955 consensus sequence, oligonucleotides were synthesized: 1) to identify by 7CR a cDNA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PROM4.

A pair of 7CR primers (forward and reverse) were synthesized: fowr PCIRi z 5'-AGGGTCTCCAGGAGAAAGACTC.3- (SEQ ID reverse PCR primer 5S-ATI'GTGGGCCIGAGACATAGAC.3' (SEQ ID NO:41) Additionally, a synthetic oligomicleotide hybridization probe was constructed from the consensus DNA3O9S sequence which had the following hucleotide sequence ATTCGICTCGTC3(SEQ ID NO:42) In order to screen several libraries for a source of a full-length clone. DNA from the libraries was screened by 7CR amplification with the PCR primer pair identified above. A positive library was then use4 to isolate clones encoding the PR0246 gene using the probe oligormcleotide and one of the 7CR primniers.

RNA for constrution of the eDNA libraries was isolated from human fetal liver tissue. DNAnqmi of the clones isolated as described above gave the fbu-length DNA sequence for PR0246 (herein designated as UNQ220 (DNA35639-l 172)J (SEQ ID NO:38) and the derived protein sequence for PROM4.

T'he entire micleotde sequence of UNQ220 (DNA35639-1 172) is shown in Figure 16 (SEQ ID NO.38).

Clone UNQ220 (DNA35639-1 172) contains a single open reading frani witi an apparent translational initiation site at nucleotide positions 126-128 an ending at tie stop codon at nicleotide positions 1296-1298 (Figure 16). The prccicted pojypeptide precursor is 390 ano acids long (Figure 17). Clone UNQ220 (DNA35639-1 172) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209396.

Analysis of the amino acid sequene of the full-length PR0246 polypeptide suggests that it possess significant homology to tie huiman cell surface protein HCAR, thereby indicating that PR0246 may be a novel cell surface virus receptor.

EXAhiPLF 9: Isolation of cDNA Clone Fincodii Hunu~n PRQ=2 A consensus DNA sequene was assembled rlative to other EST sequences using pbrap as described in Exampule 1 above. This consesus sequence is hercin designated DNA2S75S. An EST proprietary to Genentcds was employed In the consensus assembly. This EST is shown in Figure 20 (SEQ ID NOiG0) and is herein designated as DNA2195l.

.15 Based on the DNA28758 consensus sequence. oligomicleotides were synthesized: 1) to identify by PCR a cDNA library that contained the sequence of interest. and 2) for use as probes to isolate a clone of the full1-length coding sequence for PR0228.

PCR primers (forward and reverse) were syntheized: *.:forward PCR grimer S-GGTAATGAGCTCCATACAG-3' (SEQ ID NO:51) forward PcR orisn 5'-GGAGTAGAAAGCGCATGG-3' (SEQ ID NO:52) forward PCR oRin 5'-CACCTGATACCATGAATGGCAG..3 -(SEQ ID NO:53) rrverse PCR Rrimer 5'CGAGCTCGAATrAATTCG-3* (SEQ ID NO:54) reverse PCR primer 5'-GOATCICCTGAGCTCAGG-3- (SEQ MD N0.55) reverse PCR Rri=e S'-CCTAGflGAGTGArVCMTAG.3 (SEQ ID NO:56) Additionally. a synthetic oligonizeleotide hyWdization probe was constructed from the consensus DNA29758 sequenc which had die following mcleotide sequence hybridization orobe 5S-ATGAGACCCACACCCATGCCGATCTGACACA1-mYJCAAn.

3 (SEQ ID NO:57) In order to screen several, libraries for a source of a full-length cdone, DNA from the libraries was screened by PCR amtplification with the PCR primer pairs identified above. A positive library was then used to isolate clones encoding the PR0228 gene using the probe oligonucleotide and one of the PCR primers.

RNA for construction of the eDNA libraries was isolated from human fetal Madcy tissue.

DNA sequencing of the clone isolated as described above gave the fulfl-length DNA sequence for PR0228 [hereint desizoated as UNQ202 (DNA3309-1202)] (SEQ ID NO:48) and the derived protein sequejice for PROM2S.

The entire nucleotide sequence of UNQ202 (DNA33092-1202) is shown in Figure 18 (SEQ MD NO:48).

Clone UNQ2D2 (DNA3309-1202) contains a single open reading frarne with an apparent trnslational initiation site at isicowie positions 24-26 of SEQ ED NO:48 and ending at the stop codon after mucleotide position 2093 of SEQ ID NO:48. The preicted polpeptide pncursor is 690 amino acids long (Figure 19). Clone UNQ202 (DNA33092- 1202) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209420.

Analysis of the amino acid sequence of the full-length PR0228 polypeptide suggests that portions of it Possess significant homology to the secretin-related proteins CD97 and EMRI as well as the secretn member, lairophilin. thereby indicating that PR022 ay be a new member of dhe secretin related proteins.

IAMPLE 10: Isolation of cDNA Clones ncodlw Human PRO533 The ESTsiquerice accession mmbr AF00768. a murine fibroblast growth factor (FGF-15) was used to searvarus pblic ESTdatabases Genflank, Dayhoff, etc.). The search was perfored using the computer program BLAST or BLAST2 (Altcdhul et al.. Methods in Emvmloev, 2:460480 (1996).

htp/b=ws/&~/EDEM as a comparison of' the ECD protein sequences to a 6 frame t~lation of the EST sequences. The search resulted in a hit with GenBank EST AA220994, which has been identified as stratagene NT2 neuronial precursor 937230.

Based on the Genbank EST AA220994 sequence. oligonucleotides were synthesized: 1) to identify by PCR a cDNA library that contained the sequence of interet, and 2) for use as probes to isolate a clone of' the full-length coding sequence. Forward and reverse PCR primers nay range from 20 to 30 nucleotides (typically about 24), and ame designed to give a PCR product of I00-1000 bp in length. The probe sequences are typically 40-55 bp (typically about 50) in length. In order to screen several libraries for a source of a fulfl-lengthl clone, DNA from the libraries was screened by PCR amplification, as per Asusibe er al.. Oirrew ProtocoLs in Molecular Biology, with the PCR primer pair. A positive library was then wsed to isolate clones encoding the gene of' interest using the probe 20 oligonudeotide and one of th PCP. primers.

In order to scret several libraries for a source of a fil-length clone, DNA from the libraries was screened by PCR amzplification with the PCR primer pair identified below. A positive library was then wed to isolate clones encoding the PR0533 gene using the probe oligonucleotide and one of the PCR primers.

R.NA for construction of the cDNA libraries was isolaied from hulman fetal retina. Thc cDNA Libraries used to isolated the cDNA clones were constructed by standard methods using commercially available reagents Invitrogen, San Diego, CA, Clontedi, etc.) The cDNA was primed with oligo dT containing a Nod site, linked with blunt to Sail hemnikinased adaptors, cleaved with Notd, sized appropriately by gel electrophoresis, and cloned in a defined orientation into a suitable cloning vector (such as pRCB or pRKD; pRKSB is a precursor of pRKSD that does not contain the Sf!l site; see, Holmes et al., Science. 128-1280 (1991)) in the unique XhoI and Notd sites.

A cDNA clone'was sequenced in its entirety. 'Me fall length micleoodde sequence of PR0533 is shown in Figure 21 (SEQ ID NO:58). Clone DNA49435-1219 contains a single open reading frame with an apparent translational. initiation site at nucleotide positions 459-461 (Figure 21; SEQ ID NO:58). The predicted polypeptide prectirsor is 216 amino acids long. Clone DNA47412-1219 has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209480.

Based on a BL.AST-2 and FastA sequence allgunmem analysis of the foil-length sequence, PR0533 shows ammno acid sequence identity to flbroblast growth factor The oligonueleotide sequences used in the above procedure Were dic following: FGFIS.forward: 5'-ATCCGCCCAGATGGCACAATGTGiTA-3* (SEQ ID FGF1S.probe: S*-GCCTCCCGG=cTCCTrGAGCAGTGCCAAACAGCGGCAGTGTA.3' (SEQ ID NO:6 1); FGF1S .reverse: S*-CCAG~TCGGTGACAAGCcCAAA-3 (SEQ ID NO:62).

EXAMPLE 11: Isolation of cDNA Clones Encoding Hunmin PRO245 A consensus DNA sequence was assembled relative to the other identified EST sequences as described in Example 1 above. wherein the census sequence is designated herein as DNA309S4.

Based on the DNA30954 canensus seqtmi. olioniadeotides were synthesized to identify by PCR a cDNA library that contained the sequence of interest and for use as probes to isolate a clone of the fulfl-length coding sequence for PROM4.

A pair of PCR primers (forward and reverse) were synthesized: farward PCR prime S'-ATCGflGTGAAGrAGTGCCCC-3' (SEQ MD reverse PCR 2rimer S-ACCTGCGATATCCAACAGAAITG-3' (SEQ ID NO:66) Additionally, a synthetic oligortucleotide hybridization probe was constructed from dhe consensus DNA30954 sequence which had the following nuelcotide sequence hybridaiw R 5-GGAAGAGGATACAGTCACTCTGGAAGTATTAGTGGCTCCAGCAGrC.3' (SEQ ID NO:67) In order to screen several libraries for a source of a full-length clone. DNA from the libraries was screened by PCR amplification with the PCR primer pair identified above. A positive library was then used to isolate clones 20 encoding the PRO245 gene using the probe ofigonucleotide and one of the PCR primers.

RNA for construction of the cDNA libraries was isolated from human fetal liver tissue. DNAa~mng of the clones isolated as described above gave the full-length DNA sequence for PR0245 (herein designated as UNQ219 (DNA3S63B-l 141)] and the derived protein sequence for PROW4.

The entire nuclootide sequence of UNQ219 (D)NA3S63S-1 141) is shown in Figure 23 (SEQ ID NO:63).

Clone UNQ219 (DNA3S638-1141) contains a single open reading frame with an apparent translational initiation site at nucleotide positions 89-91 and ending at the stop codon at sazeleotide positions 1025-1027 (Fig. 23; SEQ MI NO:63). The predicted polypeptide preiror is 312 amino acids long (Fig. 24). Clone UNQ219 (DNA3S638-l 141) has been deposited with ATCC on September 16.,19917 and is assigned ATCC deposit no. ATCC 209265.

Analysis of the amino acid sequence of the full-ength PR0245 suggests that a portion of it possesses 60 amino acid identity with the hurman c-myb protein and, therefore, may be a new member of the transmembrane protein receptor tyrosine kinase family.

EAMPLE 12: Isolation of eDNA Clones Encoding Human PRO22O, PR0221 and PROM2 PRO22D A cansensus DNA sequence was assembled relative to die other identified EST sequences as described in ExamoplelI above, wherein the consensus sequence is designated herein as DNA28749. Based on the DNA28749 consensus sequence. oigonucleoddes were synthesized to identify by PCR a eDNA library that contained the sequence of interest and for use as probes to isolate a clone of the full-length coding sequence for PR0220.

A pair of PCR primes (forward and reverse) were synthesized: fowr Rpie S'-TCACCTGGAGCCTATGCC-3- (SEQ WD NO:74) rmePCR Rme* 5.-ATACCAGCrATAACCAGGCT'GCG.3- (SEQ ID Additionally, a synthetic oligonucleotidc hybridization probe was constructed from the consensus DNAM849 sequence which had the following nucleotide sequence: ATCTCTCCATCTAGATCGATATr-IY- 3 (SEQ ID NO:76).

In order so screen several libraries for a soure of a full-length clone. DNA from dhe Libraries was screened by PCR amplification with the PCR primer pair identified above. A positive librury was then used to'isolate clones encoding the PR0220 gene using dhe probe aligomacleozide and one of' the PCR primers.

RNA for construction of the eDNA libraries was isolated from huma fetal lung timse. DNAqwasg of the clones isolated as described above gave the full-length DNA sequence for PR0220 (herein designared as UNQ194 (DNA32298-1 132) and die derived protein sequence for PR0220.

The entire nucleooide sequence of UNQ194 (DNA32298-1132) is shown in Figure 25 (SEQ ED NO:68).

15 Clone UNQ194 (DNA32298I 132) comiains a single open reading frame with an apparent translational initiation site at nucleodec positions 480-482 and ending at the stop codon at nucleotide positions 2604-2606 (Figure 25). The predicted potypeptide precursor is 708 amino acids long (Figure 26). Clowe UNQ194 (DNA32299-1132) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209257.

Analysis of the amino acid sequence of the full-length PR0220 shows it has homology to member of' the leucine rich repeat protein superfamrily, including the leucine rich repeat protein and the neuronal leucine-rich repeat protein 1.

PRO221 A consensus DNA sequence was assembled relative to the other identified EST sequences as described in Example 1 above, wherein the consensus sequec is designated herein as DNA28756. Based on the DNA28756 consensus sequece, oligomaucleotides were synthesized to identify by PCR a eDNA library that contained the sequence of interest and for use as probes to isolate a clone of die full-length coding sequence for PR0221.

A pair of PCR primers (forward and reverse) were synthesized: forgw PCR orimer 5'-CCATGTGTCrCCTCCTACAAAG-3' (SEQ ID NO:77) reverse PCR primer 5'-GGGAATAGATGTGATCTCATCvG3* (SEQ ID NO:78) Additionally. a synthetic oligonucleotide hybridization probe was constructed from the consensus DNA28756 sequence which had the following nucleotide sequence: hybiati ro 5*CCTTGAT;AACCAGATCTGGT7CTCG3 (SEQ ID NO:79) In order to screen several Libraries for a source of a full-length clone, DNA from the libraries was screened by PCR amplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PR0221 gene using the probe oligosaicleotide and one of the PCR primers.

RNA for consruction of the cDNA librauies was isolated from human fetal tuig tissue. DNAoqznEing of the clones isolated as described above gave the full-length DNA sequence for PR0221 (herin designated as UNQ195 (DNA.33089-1 132) and die derived protein sequence for PR0221.

The entire nucleotide sequence of LUNQ195 (DNA33OS9-1132) is shown in Figure 27 (SEQ ID Clone UNQ195 (DNA33089-1132) contins a single open reading frame with an apparent tr-anslational initiation site at nucleotide positions 179-181 and ending at the stop codon at nucleooidc positions 956-958 igure 27). The jpedjcted polypepiki precursor is 259 amino acids long (Figure 28). PR0221 is believed to have a transmembrane region at amino acids 206-225. Clone UNQ195 (DNA33089-1 132) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209262.

Analysis of the amin acid sequence of the full-length PR0221 shows it has homology to membe~r of the leucine rich repeat protein superfamily. including the SLIT protein.

PRfl227 A consensus DNA sequence was assembled relative to the other identified EST sequences as described in Example I above. wherin the consensus sequence is designated herein as DNA2S74O. Based on the DNA28740 consensus sequence, oligonucleotides were synthesized to identify by PCR a cDNA library that contained the sequence of interest and for use as probes to isolate a clone of the full-length coding sequence for PR0227.

A pair of PCR primers (forward and reverse) were synthesized: forward PrR Vi'er 5'-AGCAACCGCCTGAAGCTCATCC-3 (SEQ U) raverse PCR primer S'-AAGGCGCGGTGAAAGATGTAGACG-3' (SEQ ID NO:81) Additionally, a synthetic oligonucleotide hybridization probe was constructed from the consensus DNA28740 sequence which had the following nucleotide sequence: hridizaiign prob 5SGACTACATGTTCAGGACCGTACAACTCAG'ACTrJGAGGflGGJA..3. (SEQ ID N0.82).

In order to screen several libraries for a source of a full-length clone, DNA from the libraries was screened by PCR amplificatio with the PCR primer pair identified above. A positive library was then used to isolae clones encoding the PR0227 gene using the probe oligonucleotide aid one of the PCR primers.

RNA for construction of the eDNA libraries was isolated from human fetal lung tissue. DNAvzpurdzg of the clones isolated as described above gave the full-length DNA sequence for PR0227 (herein designated as UNQ201 (DNA33786-1132) and the derived protein sequence for PR0227.

The entire nicleotide sequence of UNQ201 (DNA33786-1 132) is shown in Figure 29 (SEQ ID NO:72).

Clone UNQ201 (DNA33786-l 132) 0Ontain a single open reading frame with an apparent translational initiation site at nucleotidc positions 117-119 and ending at the stop codon at nucleotide positions 1977-1979 (Figure 29). The predicted polypeptie precursor is 620 amrin acids long (Figure 30). PR0227 is believed to have a transmernbrane region. Clone UNQ201 (DNA33786-l 132) has been deposited with ATCC and is assigned ATCC qposit no. ATCC 209253.

Analysis of the amino acid sequence Of the full-length PR0221I shows it bas homology to member of the leucirst rich repeat protein superfamily, including the platelet glycoprotein V precursor and the human glycoprotein

V.

EXANMIS 1: Isolation of cDNA Clones Fitcadng Human PRO258 A consensus DNA sequence was assembled relative to other EST sequences using phrap as described in Example 1 above. This consensus sequence is herein designated DNA28746.

Based on the DNA28746 consensus sequence, oLigonucleotides were synthesized: 1) to identify by PCR a cDNA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PRO2SS.

PCR primers (forward and reverse) were synthesized: forward PCR miter S*-GCrAGGAATrCCACAGAAGCCC-3' (SEQ U) *reverse PCR nnimer 5*-AACCTGGAATGTCACCGAGCTG-3' (SEQ ID Na.6) reverse PCR primner S'-CCTAGCACAGTGACGAGGGACITGGC-3' (SEQ ID NO:87) Additionally, synthetic oligomacleodide hybridization probes were constructed from the consensus; DNA29740 sequence which had the following nucleotide sequence: hbhdatin groe -AAGACACAGCCACCCTAAACTTCAGTCTTCTGGGAGCAIGCCTGCAGCC.3- (SEQ ID NO:88) 5SGCCCTGGCAGACGAGGGCGAGTACACCTGCTCATTCCTATCCTJT.3* (SEQ ID NO:89) In order to sczmej several libraries for a source of a full-length clone, DNA from the Libraries was screened by PCR amnplificationi with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PR0258 gene using the probe oligonucleotide and one of the PCR primers.

RNA for construction of the cDNA libraries was isolated from human fetal lung tissue. DNAsezzmr of the clones isolated as described above gave the full-length DNA sequence for PR0258 (herein designated as UNQ225 (DNA3S918-1174)J (SEQ ID NO:83) and the derived protein sequence for PR0258.

The entire nucleodde sequence of UNQ225 (DNA3S9IS-1174) is shown in Figure 31 (SEQ ID NO:83).

.Clone UNQ225 (DNA35918-1174) contains a single open reading frame with an apparent transbtiossl initiation site at nucleotide positions 147-149 of SEQ 133 NO:83 and ending at the stop codon after nucleodde position 1340 of SEQ ID NIO:83 (Figure 3 Ile predicted polypeptdec precursor is 398 amino acids long (Figur 32). Clone UNQ225 (DNA359IS-l 174) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209402.

Analysis of the amino acid sequence of the full-length PRO2S8 polypeptide suggests that portions of it possess significant homology to the CRTAM and the poliovinus receptor and have an Ig domain. thereby indicating that PR0258 is a new member of the Ig superfamily.

EXAMPLE 4: Isolation of cDNA Clons Enoda Human PRO26 An expressed sequence tag database was searched for ESTs having homology to SUT. resulting in the identification of a single EST sequence designated herein as T73996. Based on the 7P3996 EST sequence, oligonucleotides were synthesized: 1) to idena*f by PCR a cDNA library that contained the sequence of interest, and 2) for use as probes to isolate cldone of the full-length coding sequence for PROM6.

A pair of PCR primers (forward andl reverse) were synthesized: forward PCR prime S'-GTrGGATCTGGGCAACAATAAC-3* (SEQ ID NO:92) revere FPCR Rrime S--AUTGTMGCAGGCTGAG7-ITAAG-3V (SEQ ED NO:93) Additionally, a synthetic oligonucleotide hybridization probe was constructed which had the following nucleotidc sequence S'-GGTGGCTATACATGGATAGCAA1TACCTG;GACACGCTGTCCCGGG-3 (SEQ ID NO:94) In order to screen sever-al librarics for a source of a fuW-lengt clone, DNA from the libraries was screened by PCR aroplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PR0266 gene using the probe oligonucleotidc and one of the PCR primers.

RNA for constuion of the cDNA libraries was isolated from buman feta brain tissue. DNAseqxnrg of the clones isolated as described above gave the fuUl-cngth DNA sequence for PR0266 [herein designated as UNQ233 (DNA371SO-l l7S)I (SEQ ID NO:90) and the derived protein sequence for PR0266.

The entire nucleodde sequence of UNQ233 (DNA37150-1178) is shown in Figure 33 (SEQ ID Clone UINQ233 (DNA37LSO-1178) contains a single open reading frame with an apparent translational initiation site at siucleotide positions 167-169 and ending at te stop codon after nucleotidc position 2254 of SEQ ID NO:90. The predfictod polypeptide precursor is 696 amino acids long (Figure 34). Clone UNQ233 (DNA37lSO-1 178) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209401.

Analysis of the amino acid sequence of the ful-length PR0266 polypeptide suggests that portions of it possess; significant homology to the SLIT protein, thereby indicating that PR0266 may be a novel leucine rich repeat protein.

FXAMIFl. .13: Isolation of cDNA Clones Encoding Human PRO269 A consensus DNA sequence was assembled relative to other EST sequences using phrap as described in Exanple 1 above. This consensus sequence is herein designated DNA35705. Based on the DNA35705 consensus *sequeici oligonucleotides were synthesized.~ 1) to identify by PCR a cDNA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0269.

Forward and reverse PCR primers were synthesized: forward PCR grimer (Mf) S'-TGGAAGGAGATGCGATGCCACCTG -V (SEQ ID NO:97 forward PCR swinier (12) 5'-TGACCAGTGGGGAAGGACAG-3' (SEQ ID NO:98) forward PCR primer S'-ACAGAGCAGAGGGTGCCrfG-3' (SEQ ID NO:99) reverse PrR lzrimer S'-TCAGGGACAAGTGGTGTCTCTCCC-3* (SEQ ID NO: 100) reverse PCR primer 5'-TCAGGGMAGGAGTGTGCAGTTCTG-3' (SEQ ID NO: 101) Additionally. a synthetic oligonticleocde hybridization probe was constructed from the consensus DNA35705 sequence which had the following nucleofide sequence: -ACAGCTCCCGATCTCA0rrACT GCATCGCGGACGAAT7CGGCGCTCGCTJ-3 (SEQ ID NO: 102) in order to screen several libraries for a source of a full-leogth clonec. DNA from the librarie s was screened by PCR amnplification with the PCR primar pairs identified above. A positive library was then used to isolate clos=s encoding the PR0269 gene using the probe oligonucleotide and one of the PCR primers.

RNA for consticstion of the cDNA libraries was isolated from human fetal kidney tissue.

DNA sequencing of the clones isolated as described above gave the full-length DNA sequence for OR0269 Pierein designated as UNQ236 (DNA3S26O-1 180)) (SEQ ID NO:95) and the derived protein sequence for PRCY269.

The entire nucleooide sequence of UNQ236 (DNA38260-1 180) is shown in Figure 35 (SEQ ID k40:95).

Clone UNQ236 (DNA38260- ISQ) contains a single open reading fme with an apparent translational initiation site at nucleonde positions 314-316 and ending at the stop codon at nueetide positions 1784-1786 (Fig. 35; SEQ ID 71c predicted polypcptide precuror is 490 amino acids long (Fig. 36). Clone UNQ236 (DNA38260-1 180) has been deposited with ATCC and is assigned AT7= deposit no. ATCC 209397.

Analysis of the *rm acid sequence of the full-length PR0269 suggests that portions of it possess significant homology to the human thoinbomdulin proteins, thereby indicating that PRCY269 may possess one or more thromboniodulin-like domains.

EXAMPLE..M~ Isolation of eDNA Clones ncodine Human PRO297 20 A consensus DNA sequem encoding PRO297 was assembled relative to the other identified EST sequences as described in Example 1 above. wberen the consensus sequence is designated herein as DNA28728. Based on the :DNA28728 consensus sequa3=, oligomiclentides were synthesized to identify by PCR a cDNA library that contained sequence of interest and for use as probes to isolatte a clone of the Mil-length coding sequence for PR0287.

A pair of PCR primers (forward and reverse) were synthesized: forward PcR siriter S'-CCGA1TCATAGACCrCGAGAGF.3- (SEQ ID NO: 105) *reverse PCR primle 5- 4TCAAGGAGTCCTCCACAATAC-3- (SEQ ID NO: 106) Additionally, a synthetic oligonucleotide hybridization probe was constructed from the consensus DNA29M2 sequence which had the following emleotide sequence hyrdaio pob 5S-GTGTACAATGGCCATGCCAAGGCCAGcGcATGGCCG{~rcGT.3' (SEQ ID NO: 107) In order to screwn several libraries for a source of a full-length clone, DNA frm the libraries was screened by PCR amplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding te PR0297 gene using the probe oligonucleotide and one of die PCR primers.- RNA for construction of the cDNA libraries was isolated from human fetal kidney tissue.

DNA sequencing of the clones isolated as described above gave the full-length DNA sequence for PR0287 (herein designated as UNiQ250 (DNA39969-1 185), SEQ ID NO: 103) and the derived protein sequence for PROW.

Thie entire nuclonde sequence of UNQ250 (DNA39969-1 185) is shown in Figure 37 (SEQ MD NO: 103).

Clone UNQ2S (DNA39969-I 185) contains a single open reading frame with an apparent translational initiation site at nucleatide positions 307-309 and ending at the stop codon at nucleotide positions t552-1554 (Fig. 37; SEQ MD NO:103). The predicted polyetd preaoor is 45 a o acids long (Fig. 38). Clone UNQ250 (DNA399-9118s) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209400.

Analysis of thc amin acid sequence of the fll-length PR0287 suggests that it may possess one or more procolLagen C-Mrteinase enhancer protein precursor or procoilagen C-proteinase enhancer protein-like domains.

Based on a BLAST and FastA sequence aligninent analysis of the ful-kngth sequenc, PROM shows ,mcleic acid sequence identity to procoflgcn C-proteinase enhance protein precursor and procollagen C-protemnase enhancer protein (47 and 54%, respectively).

LXAMPLIX 1 Isolation of cDNA Clones Encoding Hnman PRO214 A consensus DNA sequec was assembled using phrap as described in Example I above. Tibs consensus :9 DNA sequence is designated herein as DNA29744. Based on this consensus sequence. oligonjicleotides were syntheszd: 1) to identify by PCR a cDNA library thai contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-lngth coding sequence.

9 9 In order to screeni several ibiaries; for a source of a full-length clone, DNA from the libraries was screened by PCR amnplification with the PCR prim pair identifled below. A positive library was then used to isolate clones encoding the PR0214 gene using the probe oligonuclootidec and one of the PCR primers.

*.:RNA for construction of the eDNA libraries was isolated from human fetal lung tissue. AcDNAdcone was sequenced in its entirety. The full length nucleotideseqxene of DNA32286-1191 is shown in Figure 39 (SEQ ID NO: 108). DNA32286-1191 contaiins a single open reading frame with an apparent translational initiation site at .9 *nucleotide position 103 (Fig. 39; SEQ ID NO: 108). The predicted polypeptide precursor is 420 amin acids long 9.9.(SEQ ID NO: 109).

Based on a BLAST and PastA sequece~ aligamem analysis of the full-length sequence, PRO2 14 polypeptide shows amino acid sequence identity to NIT protein andlor Fibulin (49 and 38 respectively).

Tnc ouigoniacawoe seune used. in the above proceidure were the followig: 287 4 4.p (01.1555) 5S-CCGGCTATCAGCAGGTGGGCCCAAGTGTCCAGAATAGYJXA.3' (SEQ ID NO: 28744.f (0L1556) s,-ATFcrGCGTGAACAMTAGGGC..3* (SEQ ID NO: I11) 28744.r (OLISS?) 5*-ATCTGCTTGTAGCCCTCGGCAC-3 (SEQ ID NO:112) EXAMPLEIS: Isolation of eDNA Clones ncodingr HuMan PR0317 A consensus DNA sewne was assembled using phrap as described in Example I above, wherein the conisensus sequence is herein designated as DflA2872. Based on this consensuts sequence. oligonuclcotides were synthesized:- 1) to identify by PCR a eDNA library that contained the sequence of interest, and 2) for use as probes go isolate a clone of the fizll4euigth coding sequence. 7be forward and reverse PCR primers, respectively. synthesized fo is purpose were: S*-AGGACTGCCATAACTTGCCT'G (0U489) (SEQ ID NO: 11IS) and S'-ATAGGAG7TGAAGCAGCGCTGC (011490) (SEQ ID NO:116).

The probe synthesized for this purpose was: 5*TTTGCTGCATCCc~cc~rCACCG (0U1488) (SEQ ID NO: 117) mRNA for construction of the cDNA libraries was isolated from humnan fetal idney tissue.

In order to sceen several lilrares for a source of a full-length clone, DNA from the libraries was screened by PCR amplification, as per Ausubel et at.. Curremr Protocols in Moleevia, BioIgY (1989). with the PCR primer pair identified above. A positive library was then used to isolate clones containing the PRO317 genie using die probe oligonuclcotide idetified above and one of the PCR primers.

V066 A cDNA clone was sequenced in its entirety. Mme endre: nucleooide sequence of DNA33461-1199 (encoding 0O PR0317) is shown in Figure 41 (SEQ ID NO: 113). Clone DNA33461-1199 contairis a single open reading frame so with anappauetiitrnslational initiation site at nucleotide Positions 68.70 (Fig. 41; SEQ ID NO:113). The predicted polypeptide precursor is 366 amino acids long. The preicted signal sequence is 2-in acids 1-18 of Figur 42 (SEQ 15 ID NO:114). There is one predicted N-linked glycosylation site at amino acid residue 160. Clone DNA33461-1 199 ses has been deposited with ATCC and is assigned ATCC deposit no. ATCC 20967.

Based on BLASTru and FastA~h sequence alignment analysis (using dhe ALIGN'" computer program) of the ful-length PRO317sequence. PR0317 shows the most amino acid sequence identity to EBAF-l The 0@ results also demonstrate a significant homology between human PR0317 and mouse LEFTY protein. The C-terminal end of the PR0317 protein conains many conserved sequences consistent with the pattern expected of a member of the TGP- superfamily.

I sifu expression analysis in human tissues performed as described below evidence that there is distinctly strong expression of the PR0317 polypeptide in pancreatic tissue.

EXAMPLE..Il: Isolation of eDNA clones Encodin,! Human PRO301 A consensus DNA sequence designated herein as DNA35936 was assembled using phrap as described in Example I above. Based on this consensus sequence. oligonucleotides were synthesized: I) to identify by PCR a eDNA library that cotainied the sequence of interest and 2) for use as probes to isolate a clone of the full-length coding sequence.

In order to screen several libraries for a source of a full-length done, DNA from die libraries was screened by FCR amplification with the PCR primer pair identified below. A positive library was then used to isolate clone encoding the PRO301 gene using the probe oligomicdeotide and one of the PCR primers.

RNA for construction of the eDNA libraries was isolated from human fetal kidney.

A cDNA clone was sequeniced in its entirety. The full length rauclotide sequence of native sequence PRO301 is shown in Figure 43 (SEQ ID NO: 118). Clone DNA40628-1216 conains a single open reading frame with an apparent transl1ational aination site at nucleotide positions 52-54 (Fig. 43; SEQ ID NO: 118). The predicted polypeptide precursor is 299 amino acids long with a prdcted molecular. weight of 32,583 daltons and p1 of 8.29.

Clone DNA40628-1216 has been deposited with ATCC and is assigned ATCC deposit No. ATCC 209432.

Based on a BLAST and PastA sequence alignmnent analysis of tie full-length sequence. PRO301 shows amioad seqnce identiy to A33 antigen premrmo and coxsackie and adenovirus receptor protein The oligonucleotide sequences used in the above procedure were the following: 0L12162 (3593611) 5*-TCGCGGAGCTGTGTFCTGMrCCC-31 (SEQ ID NO:120) 0L12163 3 59 36 .pl) 5S-TGATCCGATGGGACAAAGGCGCAAGCrCGAGAGCGrGTCC.J.3(SEQ ID, NO: 121) 0L12164 (35936.12) 5.-ACACCTGGTCAAAGATGGG-3- (SEQ ID NO: 122) 0L12165 (35936.rl) 5'-TAGGAAGAGTGCTGAAGGCACGG..3 (SEQ ID NO: 123) 0112166 (35936.13) 5'-TCCTFACTCAGGTGCTAC-3' (SEQ ED NO-:124) 0112167 (35936.r2) D 5,-AcTcAGCAGTGGTAGGAAAG-3, (SEQ ]EXAMPLE Isolation of c;DNA Clones Encodlnj Human PRO224 A consensus DNA sequence assembled relative to dhe otlher identified EST sequences as described in Exampile 1, wherein the consensus sequence is designated herein as DNA30845. Based on the DNA30845 consensus sequence. oligonucceotides were synthesized to identify by PCR a cDNA library diat contained the sequence of interest and for use as probes to isolate a clone of the fullngth coding sequence for PR0224.

A pair of PCR primers (forward and reverse) were synthesized: :forward PCR Vnne S'-AAGTrCCAGTGCCGCACCAGT)GGC-3' (SEQ ID NO:128) reverse IPCR primer 5*-FOGrCCACAGCCGAGCTCGTCG-3' (SEQ ID NO: 129 Additionally. a synthetic oligonucleotide hybridization probe was constructed from the consensus DNA30845 :sequence which had the following nucleotide sequence hybridization vrobe S.-GAGGAGGATGCAGGATGAGCCATGTACCCAAGGCAJATGCCACC.3.(SEQ ID NO:130) In order to screen several ibrauies for a source of a full-length clone, DNA from die libraries was screened by PCR amplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding dhe PR0224 gene using the probe oligonucleotide and one of the PCR primers.

RNA for construction of the eDNA libraries was isolated from human fetal liver tissue. Dt4Asecr of the clones isolated as described above gave the full-length DNA sequence for PR0224 [herein designated as UNQ198 (DNA33221-1133)J and dhe dcrived protein sequence for PROM2.

The entire nuicleotide sequence of UNQ198 (DNA33221-1133) is shown in Figure 45 (SEQ ID NO: 126).

Clone UNQ198 (DNA33221-1133) contains a single open reading fr-ame with an apparent translational initiation site at nucleotide positions 96-98 and ending at the stop codon at nucleotide positions 942-944 (Figure 45; SEQ ID NO: 126). The start of a trnnembrane region begins at nueleotide position 777. The predIcted polypeptide precursor is 282 amino acids long (Figure 46). Clone LUNQ199 (DNA33221-l 133) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 2092,63.

Analysis of the amino acid sequence of the fulfl-length PR0224 suggests that it has homology to very low-' density lipoprotrin receptors. apolipoprotein E receptor and chicken oocyte receptors P95. Based on a BLAST and FastA sequence aligntnent analysis of the MtI-length sequence, PR0224 has amin acid identity to portions of these proteins in the range from 28 to 45%. and overall identity with these proteints in the range from 33 to 39%.

EXAMPLEB21: Isolation of eDNA Clones Eagading Human PRO=2 A consensus DNA sequence was assembled relative to the other identified EiST sequences as descuibed in Example 1 aborve, wherein the consensus scquec is designated herein as DNA28771t. Based on the DNA28771I consensus sequence, oligonucleotides were synthesized to identif by PCR a cDNA library that contained the sequence of interest and for use as probes to isolate a clone of the full-liength coding sequence for PR0222.

A pair of PCR primers (forward and reverse) were synthesized: forward FCR primer S'-ATCTCCrATCGCTGCITrCCCGG-3' (SEQ ID NO:133) reverse PCR primer 5*-AGCCAGGATCGCAGTA4CTCC.3' (SEQ ID NO:134) Additionally. a synthetic oligotrucleotide hybridization probie was constructed from the consensus DNA28771 sequence which had the following nucleodde sequence: b~ridizain probe 5,-ArrAAAcTrGATGGGTcTGCGTATcOAGTGCrACAAAACCnATCT.3- (SEQ ID NO: 135) In order to screen seveal libraries for a source of a full-lengtli clone, DNA from the libraries was screened by PCR arrplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PR0222 gene using the probe oligonucleotide andi one of the PCR primers.

:RNA for construction of the eDNA libraries was isolated from humnan fetal kidney tissue.

DNA sequencing of the clones isolated as descibed above gave the fuill-length DNA sequence for PRO22 (herein designated as UNQ 196 (DNA33 107-1135)] anth de derived protein sequence for PR0222.

Cone The entire nucicotide sequenc of UJNQ196 (DNA33107-1 135) is shown in Figure 47 (SEQ ID NO: 131).

Cln UNQ196 (DNA3310Y7-1135) contains a single open reading framei with an apparent translational initiation site at nuclcotide positions 159-161 and ending at the stop codon at mucleortide positions 1629-1631 (Fig. 47; SEQ ID NO:131). The predicted polyp*&tidpreursoris 490 an acids long (Fig. 48). Clone UNQ196 (DNA33107-1135) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209251.

Based on a BLAST and FastA sequence alignment analysis of the full-length sequence, PR0222 shows amino acid sequence Identity to mouse: compicroem factor h precursor conipletrent receptor mouse complement C3b recptor typ 2 long formt precursor C25-47 and human hypothetical protein kiaaO27 ]EXAMPLE 22: Isolation of cDNA clones Encodin PR0734 A consensus DNA sequence was assembled (DNA30926) using phrap as described in Example I above.

Based on this consensus sequence, oligomicleoddcs; were syilhesized: 1) to identify by PCR a eDNA library tha contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence.

RNA for the construction of the cDNA libraries was isolated using standard isolation protocols. e.g..

Ausubel et at.. Current Protocols In Molecular Biology. from tisue or cell line sources or it was purchased from comamercial sources alonte). The cDNA libraries used to isolate the cDNA clones were constructed by nandard methods Ausubel et at.) using commnercially available reagents Invitrogen). This library was derived from 22 week old fetal brain tissue.

A cDNA clone was sequenced in is entirety. The enire nucicotide sequence of PR0234 is shown in Figure 49 (SEQ ID-NO: 136). The predicted polypeptide premursor is 382 amino acids long and has a calcutlated molecular weight of approximately 43.1 kDa.

Th7e oligoaucleotide sequences used in the above proccdurc were the following: 30926.p (0U826) (SEQ ID NO: 138): 5'-G1CATrGAAAACcTCTGCCATCT OATGGT*GACTTCTGGATTGGGCTCA-3* 3096.f (0U827) (SEQ ID NO:139): 5-AAGCCAAAGAAGCCTGCAGGAGGG-3' 30926.r (0U828) (SEQ ID NO: 140): 5-CAGTCCMGCATAAAGGTCCTGGC-3' EXAMPLE 23: Isolation of cDNA Clones Encodin, Human==R23 A consensus DNA sequence was assembled relative to the other identified EST sequececs as described in Example I above. wherein the consensus sequence was designated herein as DNA30933. Based on the DNA30933 .15 consensus sequence. oligonucleocides were synthesized to identify by PCR a cDNA library that contained the sequence of interest and for use as probes to isolate a clone of the ful-lngth coding sequenc for PR023 1.

Three PCR primers (tw~o forward and one reverse) were synthesized: forward PCR primner I 5*-CCAACTACCAAAGCTGCTGGAGCC-3* (SEQ DN14:143) *forward PCR grimer 2 S'-GCAGCTCTATACCACGGGAAGGA-3- (SEQ ID NO:144) reverse PCR primer 5Y-TCC1TCCCGTGyGTAATAGAGCTGC-V' (SEQ ID 140:145) Additionally. a synthetic oligonucleodde hybridization probe was constructed from the consensus DNA30933 sequence which had the following nucicoddc sequence hybridization probe 5*-GGCAGAGAACCAGAGGCCGGAGGAGACTGCCTCTIACAGCCAGG-3' (SEQ ID 140:146) In order to scree several libraries for a sourcne of a full-length clone. DNA from the libraries was screened by PCR amplifiation with t PCR primer pairs identified above. A positive library was then used to isolate clones encoding the PR0231 genc using the probe oligomidleotide and one of the PCR primers.

RNA for construction of the cDNA libraries was isolated from hun, fetal liver tisse. DNAsqnm of the clones isolated as described above gave the full-length DNA sequence for PR0231 (herein designated as UNQ205 (DNA34434-1139)J and the derived protein sequence for PRO23 1.

The entire nuclootide sequence of UNQ205 (DNA34434-1l139) is shown in Figure 5S1 (SEQ ID NO: 14 1).

Clone UNQ20S (DNA34434-1139) contains a single open reading frame with an apparent translational initiation site at nucleotide positions 173.175 and ending at the stop codon at nucleocide positions 1457-1459 (Fig. 51; SEQ ID NO: 141). The predicted polyptde precursor is 428 amino acids long (Fig. 52). Clone UNQ205 (DNA34434-l 139) has been deposited with ATCC on September 16, 1997 and is assigned ATCC deposit no. ATCC 209252.

Analysis of the amin acid sequence of the full-lengt PR0231 suggests that it possesses 30% and 31 amino acid identity with the human and rat prostatic acdd phosphatase precursor proteins, respectively.

£XAMPLEJA: Isolation of cDNA Clones Encodiny HuMan pR029 A consensus DNA sequence was assembled *relative to other EST sequences using phrap as described in Example I above. This consensus soequence is herein designated DNA28762. Based on the DNA28762 consensus sequence. oLigomicleotides were synthesized: 1) to identify by PCR a cDNA Library that contained the sequence of Ineret, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0229.

A pair of PCR primers (forward and reverse) were synthesized: fbrwardPCR Rdmci 5'-1TCAGCTCATcAccTrcAccTGCC-3* (SEQ ID NO: 149) reverse SCR ottmer 5*-GGCTCATACAAAATACCACFAGGG.3' (SQ MD Additionally, a synthetic oigonucleouide hybridization probe was constructed from the consensus DNA29762 sequence which had the following nucleotide sequence 5'-GGCCTCCACCGT-GTGAAGGGCOGGGTGGAGGTGGACGAAA(JCCAG1r- 3 (SEQ ID NO:151) In order to screew several libraries for a source of a full-length clone. DNA from the libraries was screened by PCR anplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PR0229 gene using the probe oligorrucleotide and one of the PCR primers.

RN4A for construction of the cDNA libraries was isolated from humann fetal liver tissue. lX(Aqming of the clones isolated as described above gave the full-length DNA sequence for PRO229 [herein designated as UNQ203 (DNA33lOO-1159)J (SEQ ID NO: 147) and the derived protein sequence for PR0229.

The entire nucleotide sequence of UNQ203 (DNA331OO-1 159) is shown in Figure 53 (SEQ ID NO: 147).

Clone UNQ2O (DNA33 IOD-1 L59) coitazins a single open reading frame with an apparent tanslational initiation site at nucleotide positions 98-100 and ending at the stop codon at nucleotide positions 1139-1141 (Figure 53). The predicted polypeptide precursor is 347 amino acids long (Figure 54). Clone UNQ203 (DNA33100-1159) has been deposited with ATCC and is assigned ATCC deposit no.ATCC 209377 Analysis of the amino acid sequence of the full-lnt PR29 lpI esget thatpronofi possess significant homology to antigen wcl. 1, M 130 antigen and CD6.

EXME i25: Isolation of cDN Clanes Enodine Hurran PRrYYIR A consensus DNA sequenc was asserrbled relmive to other EST sequences using phrap as described above in Example 1. This conseinu sequence is herein designated DNA30908. Based on the DNA30908 consensus sequesice, olig onucleotides were synthesized: 1) to identify by PCR a cDNA library that contained the sequence of interest, and 2) for use as probes to isolate a cdone of the full-length coding sequence for PROM3.

PCR primers (forward and reverse) were synthesized: forward PCR primer I 5-GGTG0FAAACTGTGCrCTGTGJC3' (SEQ ID NO: 154) forward PQR Udmner 2 5'-CAGGGCAAGATGAGCA17CC.3' (SEQ ID NO: 155) reverse PCR prime 5-TCATACrGTCCATCrCGGACG,>3' (SEQ ID NO: 156) Additionally, a synthetic oligomicleotide hybridization probe was constructed from the consensus DNA30908 sequence which had the following ncleotide sequence hb~ridization probe S'-AATGGTGGGGCCCTAOAAGAGCTCATCAGAGAACTCACCGCTTCTCATGC..3' (SEQ ID NO; 157) In order to screcat several libraries for a source of a fufl-lngdi clone, DNA from the libraries was scre ened by PCR amplification with the PCR primr pair identified above. A positive library was then used to isolate clones arnding dhe PR0238 gene using the probe oligonucleotide and one of the PCR primers.

RNA for constructon of the cDNA libraries was isolated from humnan fetal liver tissue. DNAsbqxnriqg of die clones isolated as described above gave the full-length DNA sequence for PR0239 and the derived protein sequence for PROM3.

The entire nucleotide sequence of DNA35600-1162 is shown in Figure 55 (SEQ ID NO: 152). Clone DNA356OD-1 162 contains a single open reading frame with an apparenit translational initiation site at micleotide positions 134-136 and ending prior to the stop codon at nrucleotide positions 1064-1066 (Figure 55). The predicted polypeptdec precusor is 3 10 amino acids long (Figure 56). clone DNA35600-1162 has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209370.

Analysis of the amino acdd sequence of the full-length PR0238 polypeptide suggests that portions of it possess significant homology to reductae, particularly oxidoreductase, thereby indicating that PR0238 may be a novel reductase.

EXAMPLEl2i: Isolation of cDNA Clones FEodinh Human PRO233 The extracellular domain =EC) sequences (including the secretion signal if any) of from about 950 known 20 seced proteins from the Swiss-Prot public protein database were used to search expressed sequence Lag (EST) databases. 7be EST databases included public EST databases GanBank) and a proprietary EST DNA database

:(UIFESEQ

T

Incyte Pharmaceuticals, Palo Alto, CA). The search was performed using the computer progrm BLAST or BLAST2 (Altshul et al., MeZ i D= 6g :460-4R (1996)) as a comparison of the ECD protein siequences to a 6 frame translation of the EST sequence. Those comparisons resulting in a BLAST score of 70 (or in sorne cases 90) or greater that did not encode known proteins were clustered and assembled into consensus DNA *sequences with the program Ophrap" (Phil Green, University of Washington, Seattle. Washington; hup'Jibozemanint.washington.eduphrap.docs/phrap.liml).

An expressed sequme tag (ES'I) was Ideiied by the EST database search and a consensus DNA sequence was assembled relative to other EST sequences using phrap. This consensus sequence is herein designated DNA30945. Based on the DNA30945 consensus seqene, oligonucleotides were synthesized: 1) to identfy by PCR a cDNA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0233.

Forward and reverse PCR primers were synthesized: forward PCR pdivnr S'-GGTGAAGGCAGAAATrGGAGATG-3' (SEQ ID NO:160) reverse PCR mimer 5'-ATCCCATGCATCAGCCTGTTTACC-3' (SEQ ID NO: 161) Additionally, a synthetic oligonucleotide hybridization probe was constructed fhom the consensus DNA30945 sequence which had the following micleotide sequence 5S-GCTGGTGTAGTCTATACATCAGA1TG7rFCTACACAAGATCCTCAG.3' (SEQ ID NO: 162) In order to screcti several libraries for a source of a full-ength clone, DNA from the libraries was screened by PCR amnplification withr the PCR primer pair identified aborve. A positive library was then used to isolate clones encding the PR0233 gene using the piobc oligoicleotide..

RNA for construction of the cDNA libraries was isolated from huma-n fetal brain tissue. DNAaporig of the clones isolated as described above gave the fuill-length DNA sequence for PR0233 (herein designated as UJNQ207 (DNA34436-1238)] (SEQ MD NO:158) and the derived protein sequence for PR0233. The entir nucceotide sequence of UNQ2O7 (DNA34436-1238) Is shown in Figre 57 (SEQ ID NO: 158).

Clowe UNQ207 (DNA34436-Mg3) cotains a single open readig frame with an apparent tramnlational initiation site at nucleotide positions 101-103 and ending at the stop codon at micleotide positions 1001-1003 (Figure 57). The predicted polypeptide precursor is 300 amino acids long (Figure 58). The full-length PR0233 protein shown in Figure 58 has an estimated mbolecular weight of about 32.964 daltons and a pi of about 9.52. Clone UNQ2O7 (DNA34436-1238) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209523.

Analysis of the amno acid sequence of the ful4cngth PR0233 polypeptide suggests that potions of it posses significant homology to reductase proteins, thereby indicating that PR0233 may be a novel reductasc.

:EAMLEi.2 Isolation of cDNA Clonesi Encodine Human PRO223 A consensus DNA sequence was assembled relative to other EST sequcences using phrap as described in Example I above. This consensus sequence is herein designated DNA30836. Based on the DNA30836 consensus sequence. oligontrcleotides were synthesized: 1) to identify by PCR a cDNA library that contained the sequec of intres, nd or seas probes to isolate aclone of the full-length coding sequence for PRO223.

PCR primer pairs (one forward and two reverse) were synthesized: forward PcIR ptrimer 5'-TrCCATGCCACCTAAGOGAGACTC-3- (SEQ ID NO:165) reverse PCR primier 1 5*-TGGATGAGGTGTGCAATGGCTGGC-3' (SEQ MD NO: 166) reerse PCR grirner 2 5*-AGCTCrCAGAGGCGGTCATAG3c-3* (SEQ ID NO:167) Additionally, a synthetic oligonuclootidc hybridization probe was constructed from the consensus. DNA30836 sequence which had the following nucleotide sequence S*-GTCGGCCCmCCCAGGACTGAACATGAAGAGrATGCCGCrmCCTSAC-3 (SEQ ID NO: 168) In order to screeni sevealA libraries for a source of a full-length clone, DNA from the libraries was screened by PCR amplification with the PCR primer pair identified above. A positive library was then used to isolate lonies encoding the PR0223 gene using the probe oligonueleotide and one of the PCR priners.

RNA for construction of the eDNA libraries was isolated from humin fetal liver tissue. DNAseqznciqg of the clones isolated as described above gave the full-length DNA sequence for PR0223 (heein designated as UNQ197 (DNA33206-I 165)) (SEQ ID NO: 163) and the derived protein sequence for PR0223.

The entire nucleode sequence of UNQI 97 (DNA33206-1 165) i shown in Figure 59 (SEQ MD NO: 163).

Clonc UNQ197 (DNA33206-1165) contains a singl open reading frame with an apparent translational initiation site at nucleotide positions 97-99 and ending at the stop codon at micleotide positions 1525-1527 (Figure 59). The piedicte~d polypeptide precursor is 476 amino acids long (Figure 60). Clone UJNQ197 (DNA3 3206-1165) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209372.

Analysis of the amino acid sequence of die full-length PR0223 polypeptide suggests that it possesses significant homology to various serine carboxypeptidase proteins, thereby indlicating that PR0223 ay be a novel serine carboxypeptidase.

EXAMPLE 2: hnlatiin of enNA Clones Encoftn Human PRC235 A consensus DNA sequec was assembled relative to other EST sequences wing phrap as described in' Example 1 above. Tis consennits seq i s herein designated IDNA30927'. Based on the DNA30927 coaenu *.sequiene. oligormleotides were synthesized: 1) to identify by PCR a cDNA library that contained the sequenc or interest, and 2) for use as probs to isolate adneof the fMJleh codingsequence for PROM3.

A pair of PCR primers (forward and reverse) were synthesized: forward PCR primer S'-TGGAATACCGCCTCCTGCAG-3' (SEQ ID NO: 171) reverse PCR primer 5'-CTCTGCCC riTF GAGAAGATGGC-3' (SEQ ID NO: 172) Additionally, a synthetic ofigonuiceotidc hybridization probe was constructed from the consensus DNA30927 sequence which had the following nucleotide sequence hybridization vrobe 5'-GGACrCACTG~CCCAGGCCrcAATATcAcCAGCCAGGACGAT-3* (SEQ ID NO: 173) In order to screen several libraries for a source of a full-length clone, DNA from the libraries was screened by PCI amplification with the PCR primer pair identified above. A positive libary was then used to isolate clones enoigtePR0235 gene using the probe oligonuckootide and one of the PCI primers.

RNA for construction of the cDNA libraries was isolated from human fetal liver tisse. DNAa~qxsrig of the clones isolated as described above gave the full-length DNA sequence for PR0235 (herein designated as UNQ209 (DNA3SSS8-1167)] (SEQ ID NO:169) and the derived protein sequence for PR0235.

The entire nucleotide sequence of UNQ209 (DNA3SSSS-1167) is shown in Figure 61 (SEQ ID NO.:169).

Clone UNQ2D9 (DNA35559-1167) contains a single open reading frame with an apparent translational initiation site at nucleotide positions 667-669 and ending at the stop odon at nucleotide positions 2323-2325 (Figure 6 The predicted polypeptide precursor is 552 amno acids long (Fgur 62). Clone UNQ209 (DNA35558-1167) has been deposited with ATCC and is assigned ATCC deposit no. 209374.

Analysis of the amnin acid sequence of the full-length PR0235 polypeptide suggests that portions of it possess significan homology to the humian, mouse and Xenwptu plexin protein. thereby indicating that PR0235 may be a novel plexin protein.

EXAMPIR 79: Isolation of enNA Clones Encoding Human PRCY236 and Human PR026 Consensus DNA sequences were assembled relative to o ther EST sequePnfces using phrap as described in Example I above. Thetse consenus sequences are herein designated DNA30901 and DNA30847. Based an the DNA30901 and DNA30947 consensus sequerces, otigemcleotides were synthesized: 1) to identify by PCR a cDNA library that-conzained the sequence of interest., and 2) for use as probes to isolate a clone of the full-length coding for PR0236 and PR0262, respectively.

Based upon the DNA30901 consensus sequence, a pair of PCR primers (forward and reverse) were synthesized: forwvard PCR Prime S'-TGGCTACTCCAAUACCCTGGCATG.3- (SQID NO:l78) rerePCR nrimer 5'-TGGACAAATCCCCFTTGCTCAGCCC-3- (SQID NO: 179) Additionally. a synthetic oigonucleotide hybridization probe was constructed from the consensus DNA30901 sequence which had the following micleotide sequer- 5S-OGGCTCACCGAAGCAGTGGACCTAT~rACCACCTGATCCAGGG-3 (SEQ MD NO: 180) Based upon the DNA30847 consensus seguence, a pair of PCR primers (forward and revers) were 15 synthesized: forward PCR primer S'-CCAGCTATGACTATGATGCACC-3' (SEQ ID 140:181) reverse PCR primer 5'-TGGCACCCAGAATGGTGI700CTC-3' (SEQ ID NO: 192) Additionally. a synthetic oligonucleotidc hybridization probe was constructed from the consensus DNA30847 sequence which had the following nucleotide sequence hrybridization orobe 5-CGAGATGTCATCAGCAAG1rCCAGGAAGTCI-nJGGAC1--ACCT)CC.3' (SEQ ID NO: 183) In order to scre several libraries for a source of full-length clones. DNA from the libraries was screened **by PCR amplifiation with the PCR primer pairs identified above. Positive libraries were then used to isolate clons encoding the PR0236 and PR0262 genes using the probe oligonucleooddes and one of the PCR primers.

RNA for construction of the cDNA libraries was isolated from humnan fetal lung tissue for PR0236 and uman fetal liver tissue for PR0262.

DNA sequmeing of the clos isolated as described above gave the full-length DNA sequence for PR0236 [herein designated as LTNQ210 (DNA35S99-l 168)] (SEQ ID NO: 174). the derived proteint sequence for PR0236.

the Fuf-kzigth DNA sequee for PR0262 (herrin designated as UNQ229 (DNA36992-1168)] (SEQ MD NO: 176) and the derived protein sequenc for PR0262.

The entire nucleotide sequence of UJNQ210 (DNA35599-l 168) is shown in Figure 63 (SEQ ID 140:174).

Clon UNQ210 (DNA3SS99-l 168) contains a single open reading frame with an apparent translational initiation site at nucleotide positions 69-71 and ending at the sto codon at nucleotide positions 1977-1979 (Figure 63). Te predicted polypeptide precursor is 636 amo acids long (Figure 64). Clone UNQ210 (DNA35S99- 168) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209373.

The entire nucleocide sequence of UNQ229 (DNA36992-l 168) is shown in Figure 65 (SEQ ID N0:176).

Clone UNQ229 (DNA36992-1 168) contains a single open reading frmne with an apparn translational initiation site at nucleouide positions 240-242 and ending at the stop codon at inucleotide positions 22D2-2204 (Figure 65). The predicted polypeptide precursor is 654 amino acids long (Figure 66). Clone UNQ229 (DNA36M921168) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209382.

Analysis of the amino acid sequence of the fungt PR0236 and PRO26 polypepides suggests that portions of those po~peptdcs possess significant homology to A-galactosidase proteins derived from various sources, thereby indicating that PR0236 and PRO26 may be novel V-galactosidas'e honiologs.

EX&MELF I: Isoation of rTDNA Clones Encoding Hinnan PRO239 A consensus DNA sequence was assembled relative to other EST sequences using phrap as described in Example I above. This consensus sequence is herein designated DNA30909. Based an the DNA30909 consensu sequence, oligonucleoddes were synthesized: 1) to identify by PCR a cDNA library that contaied the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0239.

A pair of PCR primers (forward and reverse) were syntlesized.

*forward PCR mimer 5'-CCTCCCTCTATTACCCATGTC-.3' (SEQ ID NO:186) reverse PCR pdmuer S'-GACCAAC7TCTCTGGGjArrcAGcJ3' (SEQ ID NO: 187) Additionally. a synthetic oligonucleotide hybridization probe was constructed from the consenus'DNA30909 sequence which had the following nucleotide sequence 20 ~~(SEQ ID NO: 188)asore afi-eghcoe frmtelbreswscend ~In order to screen several libraries for a oreof a ullnt lnDNAfrmtelbaiswscend by PCR amplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PR0239 gene using the probe oligonucleodde and one of the PCR primers.

RNA for construction of the cDNA Libraries was isolated from huiman fetal lung tissue. DNAgcrg of the clones isolated as described above gave the ful-length DNA sequence for PR0239 (herein designated as UNQ213 (DNA34407-1 169)] (SEQ ID NO: 184) and the derived protein sequence for PROM3.

The entire nucleotide sequem of UNQ213 (DNA34407-1 169) is shown in Figure 67 (SEQ ID NO. 184).

Clone UNQ213 (DNA34407-1169) contins a single open reading frame with an apparent translational initiation site at macleodde positions 72-74 and ending at the stop codon at mucleotide positions 1575-1577 (Figure 67). The predicted polypeptide precursor is 501 amin acids long (Figure 68). Clone UNQ213 (DNA34407-1 169) has been deposited with ATCC and is assigned ATCC deposit no.ATCC! 209383.

Analysis of the amino acid sequence of the full-lngt PR0239 polypeptide suggests that portions of it possess significant homology to the densin protein.. thereby indicating that PRO239 nray be a novel molecule in the densin family.

EXAMPE 31: Isolation of cDNA Clonies Encoding Human A consensus DNA sequence was assembled relative to other EST sequences using phrap as described in Example I abovc. This consensus sequence is herein designated DNA28731. Based on the DNA28731 consensus sequence. oligomzcleotides were synthesized: 1) to identify by PCR a cDNA library dtha contained the sequence of iteret, and 2) for use as probes to isolate a cdone of the ful-lngth coding sequence for A pair of PCR primers; (forward and reverse) were synthesized: forward PCP. or_2e= S-TCTCTATCCAAACTGTOGCG..3' (SEQ ID NO:191) reverse PCR Vrme S'-rOATGACGATCGAAGGTGG.3- (SEQ ID NO:192) Additionally. a synthetic oligonucleotide hybridization probe was constructed from the consensuis DNA28731 sequence which had the following raicleoude sequence AC3(SEQ ID NO:193) In order to screen several liraries for a source of a full-length clone. DNA from the libraries was scroened by PCR am~plification with the PCd primer pair identified above. A positive library was then used to isolate clo=e :44. ecoding the PRO2S7 ge= using the probe oligomicleotide and one of the PCR primers.

RNA for construction of the cDNA libraries was isolated from human fetal kidney tissue.

DNA sequencing of the clones isolated as described above gave the full-lengffh DNA sequence for PRO2W [hri as UNQ24A (DNA35841-1 173) (SEQ ID NO: 189) and the derived protein sequence for 4***15 .me entire zaicleotide sequence of UNQ224 (DNA3S841-l 173) is shown in Figure 69 (SEQ ID NO:189).

Clone UNQ224 (DNA35841-l 173) contains a single open reading frame with an apparent translationxal initiation site at nucleotide positions 964-966 and ending at the stop codon at niucleotide positions Z785-2787 (Figure 69). The predicted polypeptide precuror is 607 amino acids long (Figure 70). Clone UNQ224 (DNA35841-l 173) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209403.

Analysis of the amzin acid sequence of the full-length PR0257 polypeptide suggests that portions of it possess; significant homology to the ebnerin protein, thereby indicating that PR0257 may be a novel protein member related to the ebnerin protein.

EXAMPLE 32: Isolation of cDNA Clones Encoding Human PRO260 A *consensus DNA seqence was assembled relative to other EST sequences using phrap as described in Examplel1above. This consensus sequence is herein designated DNA30834. Based on the DNA30834 consensus sequence, oligomacleotides were synthesized: 1) to identify by PCR a cDNA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0260.

PCR primers (forward and two reverse) were synthesized: formard PCR primce S'-TGG1TrrGACCAGGCCAAG=rGG-3- (SEQ ID NO: 196); reverse Pr imerkA 5'-GGA7TCATCCTCAAGGAAGAGCGG-3 (SEQ ID NO:197); and reverse PCR primer B# 5AACTTUCAGCATCAGCCACTCTGC..3* (SEQ ID NO: 198) Additionally, a synthetic oligornrcleotide hybridization probe was constructed from the consensus DNA30834 sequence which had the following nucleoude sequence hvybridbmzjp robe- 5'-TrCCGTGCCCAGCTTCGGTAGCGAGTGGTGGGTA-IYJGJA-3 (SEQ ID NO:199) In order to Sa=r several libraries for a source of a full-length clon, DNA from the libraries was screened by PCR amplification with the PCR primer pair identified above. A positive library was then used ro isolate clones encoding tie PR0260 gmn using the probe oligonucleotide and one of the PCR prme,.

RNA for construction of the cDNA libraries was isolated from human fetal kidney tissue.

DNA sequencin oftlhe ckincs isolated as described above gave the fukll-lengt DNA sequence for PR0260 [herein designated as UNQZ (DNA33470-117S) (SEQ ID NO: 194) and the derived protein sequece= for PR0260.

The entire nucleotde sequence of UNQ227 (DNA33470-l 175) is shown in Figure 71 (SEQ MD NO: 194).

Clone UNQ2 (D5NA33470-1 175) aomins a single open reading frame with an apparent translational initiation site at nucleotdec positions 67-69 aind ending at dhe stop codon 1468-1470 (see Figure 71). The predicted polypeptide precrsor Is 467 amino acids long (Figure 72). Clown UNQ227 (DNA33470-1 175) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209398.

Analysis of the anti acid sequence of the fuflengt MR0260 polypeptide suggests that portions- of it possess significant homology to the alpha-1-fueosidasc precursor. thereby indicating that PR0260 may be a novel fuicosidase.

EXMPE 33f: lsolation %If eDNA Clones Farodinp Hluman PRO263 A consensus DNA sequence was assembled relative to other EST sequences using phra as described in Example I above. T1his consensus sequence is herein designated DNA30914. Based on the DNA30914 consensus sequence, ortgomicleotides were synthesized: 1) to identify by PCR a eDNA library that contained the sequence of interest. and 2) for use as probes to isolate a clone of the full-length coding sequence for PROM6.

PCR primers (tow forward and one reverse) were synthesized: forward PCR orimer I: 5'-GAGCITI'CCATCCAGGTGTCATGC-3' (SEQ ID NO:202)-.

:forward PCR primer 2 S'-GTC-AGTGACAGTACCTACTCGG-3* (SEQ ID NO:203); reverse PCR tzrijmr: S'-TGC1AGCAGGAGOAGTAGTACTAGG-3- (SEQ ID NO:204) Additionally, a synthetic oligomicleodde hybridization probe was constructed from the consensus DNA30914 sequence which had the following nucleotide sequence: -AGGAGGCCTGTAGGCrGCTGGGACrAAG'rrrGGCCGGCAAGGACCAAGT-3' (SEQ ID NO:205) In order to screen several libraries for a source of a fullength clone, DNA from the libraries was screened by PCR amplification with the PCR primer pair Identified above. A positive library was then used to isolate clones encoding the PR0263 gene using the probe oligomaicleotide and one of the PCR primers.

RNA for construction of the cDNA libraries was isolated from humann fetal liver tissue. l2NAewnxr of the clones isolated as described above gave the full-length DNA sequence for PR0263 (herein designated as UNQ230 (DNA3443 1-1177)1 (SEQ ID NO:200) and the derived protein, sequence for PROM6.

The entire nuclootide sequence of UNQ230 (DNA34431-l 177) is shown in Figure 73 (SEQ ID NO:200).

Clow UNQ23O (DNA34431-l 177 contains a single open reading framne with an apparent translationial initiation site at sarcicotide positions 160-162 of SEQ ID, N0200 and ending at the stop codon after the nucleotide at position 1126- 1128 of SEQ ID NO:200 (Figure 73). The predicted polypeptde precursor is 322 amino acids long (Figure 74).

Clone UNQ230 (DNA34431-1 1Th has been deposited with ATCC and is assigned ATCC deposit o. ATCC 209399.

Analysis of the amino acid sequence Of the ful-length PR0263 polypeptde suggests that portions of it Possess s6goificant ho ogy to CD44 antigen. thereby indicating that PR0263 my be a novel cell surface adhesion molecule.

EAMP1L 3: Isolation of cDMA Clones Encoding Humnim PRCZt A consensus DNA sequence was assembled relative to the other identified EST sequences as described in Ezanple 1 ahow, wherein the consensus sequence was designated herein as DNA3S7 12. Based an the DNA35712 consensus sequence. ofigortucadeoti were: synthesized: 1) to identify by PCR a cDNA library that contained the sequence of interest, and 2) for use as probes-to isolate a clocc of the full-length coding sequence for PROM7.

Forward and reverse PCR primers were synthesized: forward PCR nrmV S*-OCTTGATATcGCATGGGccTAc-3, (SEQ ID NO:208) **forward PCR primer (12) S'-TGGAGACAATATOCCTGAGJ.3* (SEQ ID NO:2D9) revrse PCR Drinir (-rn 5'-AACAGTI'GGCCACAGCATGGcACG.3* (SEQID NO:210) Additionally. a synthetic aligonucleodde hybridlization probe was constructed from the consensus DNA35712 sequence which had the following nucleotide sequence CJGATGTGAG-3 (SEQ ID N:21 1) In order to screen several libraries for a source of a full-length done. DNA from the libraries was screened *20 by PCR amnplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PR0270 gene using the probe ofigormcleotide and one of the PCR primers.

:RNA for construction of the eDNA libraries was isolated from human fetal lung tissue. DNAemuwing of the clones isolated as described above gave the full-length DNA sequence for PR0270 (herein designated as UNQ237. DNA39510-1 181] (SEQ ID NO-.206) and the derived protein sequence for PROM7.

The entire micleotide sequence of UNQ237, DNA39510-1 181 is shown in Figure 75 (SEQ ID 140206).

Coe UNQ237 (DNA395 10-1181) contains a single open reading fram with an apparent translational initiationi site at rerleooite Positions 3-5 and ending at the stop codon at nucleotide positions 891-893 (Fig. 75:- SEQ ID NO:206).

The precd polpeptideprcosoris 296 ansi acids long (Fig. 76). Clone UNQ237 (DNA39510-1181) has been deposited with ATCC and is assigned ATCC deposit no.. ATCC 209392.

Analysis of the amino acid sequecwe of the full-legth PRCO suggests tha portions of it possess significant homology to the thioredoxin-protein thereby indicating that the PR0270 protein may be a novel niember of the thioredoxin family.

EXAMPLE 35: Isolation of cDNA Clones Ena&lUU Human PRO271 A consensus DNA sequence was assembled relative to other EST sequences using phrap as described in Example I above. This consensus sequence is herein designated DNA35737. Based on the DNA35737 consensus sequence. ofigornucleotides were synthesized: 1) to identify by PCR a cDNA library that contained the sequence of interest, and 2) for usc as probes to isolate a clone of the full-lengh coding sequence for PROWL1 Forward and rcverse PCR primers were synthesized: forwa2rd PCR primner I S'-TGCITCGCTACTGCCCTC-3- (SEQ ID NO.214) foward PCR n1rimer 2 5*-TTCCCTGTGGGrYGGAG-3' (SEQ ID N0:21S) forward PCR orimer 3 S'-AGGGCTGGAAGCCA(3fl. (SEQ ID NO:26) reverse PCR onniner I S'-AGCCAGTGAGGAAATGCG-3' (SEQ tD NO-t1l7) reverse PCR onimer 2 5S.TGTCCAA&GTACACACA(YCrJAG3 (SEQ ID, NO:218) Additionally. a synthetic oligomzclotide hybridization probe was constructed from the consensus DNA35737 sequence which had the following nucleotide sequence S OGATCCACATCGC AT A CAGCTCr GCCGCTGAAG-3, (SEQ 11) N0:219) In order to scrcen several libraries for a source of a full-length clone. DNA from the libraries was screened by PCR amnplification with the PCR primer pair identified above. A positive library was then used to, isolate clone cwxigtePRO271 gem using the probe oligonucleode and one of the PCR primers.

RNA for construction of the cDNA libraries was isolated from human fetal brain tissue. DNAseqzimig of the clones isolated as described above gave the full-length DNA sequee for PR0271 [herein designated as UNQ238 (DNA39423-1182)J (SEQ ID NO:212) and the derived protein sequenc for PRO271.

The entire micleotidc sequence of UNQ238 (DNA39423-1182) is shown in Figure 77 (SEQ ID NO:212).

Cone UNQ239 (DNA39423-l 182 contains a single open reading fram with an apparent translational initiation site 0* at nuecotide positions 101-103 and ending at the stop codon at nucleotide positions 1181-1183 (Figure 77). The predicted polypeptide precursor is 360 amino acids long (Figure 78). Clone UNQ238 (DNA39423-1 182) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209387.

Analysis of the amino acid sequence of the full-length PR0271 polypeptide suggests that it possess significant homology to the proteoglycan link protein. thereby indicatig that PRO271 may be a link protein homolog.

EAMPE 36I: Isolation of cDNA Clones Eeodny Human ER0272 A consensus DNA sequence was assembled relative to other EST sequences using phrap as described in ExaMle I above. This consensus sequence is herein designated DNA36460. Based on the DNA36460 consensus sequence. oigomicleotides were synthesized: 1) to identify by PMR a cDNA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PROM7.

Forward and reverse PCR primers were synthesized: forward PCR piner (1f1) 5'-CGCAGGCCCTCATGGCCAGG..31 (SEQ ID NO-:222) forward PCR Vime (M1) S'-OAAATCCTGGGTAATrTcj.3' (SEQ ID NO:223) reverse PCR mimer 5S-GTGGCGOTGCTCACAGC-rCATC.3' (SEQ ID NO.224) Additionally, a synthetic oligonucleotide hybridization probe was constructed from the consensus DNA36460 sequence which had the following nucleodde sequence

CCGUACT-

3 (SEQ ID NO:225) In order to samen several libraries for a source of a ful-1rngth clone. DNA from the libraries was screened by PCR amplikazion with ft PCR pr&n= pairs identified above. A positive library was then used to isolate clones encoding the PR0272 gene using the probe oligoauleotide and one of the PCR primers.

RN4A for construction of the cDNA libraries was isolatd from human fetal lung tissue. DN9pmrg of the clones isolated as described above gave the full-length DNA sequence for PR0272 [herein designated as UNQ239 (DNA40620-l 183)J (SEQ IM NO:220) and the derived protein sequence for PROM7.

The entire nuclcotide sequce of UNQ239 (DNA40620-1 183) is shown in Figure 79 (SEQ ID NO:220).

Clone UNQ239 (DNA4062O-1 183) contains a singec open reading firme with an apparent ranslational initiation site at nuecleotide positions 35-37 and ending at the stop codon at micleotide positions 1019-1021 (Figure 79). The predicted polypeptide precursor is 328 amino acids long (Figure 80). Clone UNQ239 (DNA40620-1183) has been deposited with ATCC and is assigned ATCC deposit no. ATCC 209388.

~Analysis of the a-in acid sequece of the ful-engt PR0272 polypeptde suggests that portions of it possess; significant homology to the human and mouse reticulocalbin proteins, repetvely, theeby indicating that PR0272 may be a novel reticulocalbin protein.

EXML 3:Iolation of cDNA Clones Encodin Humnan PRO294 Tseunsuigphpasdcrbdn A cosenss DN scqcwcwas assembled relative to other S eurc sn ha sdsrbdi Example 1 above. This consensuls sequence is herein designated DNA35731. Based on the DNA35731 consensuis sequence. oligamcleotides were synthesized: 1) to identify by ICR a cdNA library that contained the sequence of :interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0294.

20 Forward and reverse ICR primers were synthesized: favard PCR primier 5'-TGGTCTCGCACACCGAT)C-3' (SEQ ID N0228) fo9*d~ fitc '-CrGCGTCCACAGGGGAG-3' (SEQ ID NO:229) fowadPC S'-CCTFGAAGCATACTGCrC-3- (SEQ ID NO:230) forward PCR nritneu 1.4) S-GAGATAGCAAlCCGCC-3' (SEQ ID NO:23 1) rxevesePR pfw (xl) Y-TrCCTCAAGAGGGCAGCC-3' (SEQ 11) NO:232) reere CRvrinr (42) S'-CTTGACCAATGTCCGAGATMT-3r (SEQ ID NO:233) Additionally. a synthetic oligonucleotde hybridization probe was constructed from the consensus DNA35731 sequence which had the following nicleodde sequence bybridization globe 5SOCTCTGAGGAAGGTGACGCGCGGGGCCT'CCGAACCCrrGGCC-FG-3' (SEQ ID NO:234) In order to screen several libraries for a source of a fufllength clone, DNA from the libraries was screened by PCR amplification with de PCR primer pairs identified above. A positive library was then usedt to isolate clones encoding the PR0294 gene using the probe oligonuceotde and one of the PCR primer.

RNA for consmiction of the cDNA libraries was isolated from human fetal brain tissue. DNAsecqg of the clones isolated as described above gave the full-length DNA sequence for PR0294 (herein designated as 144 UN2257 (NAW40604-1187) (SE* ID NO:226) and the derive protein sequence for PR0294.

The entire nucleotide sequence of UQ257 (IA40604-1187) is shaown in Figure 81 (S2EQ ID N0:226) Clone URM257 (ENA40604-1187) contains a single open reading frane with an apparent translational initiation site at nucleotide positions 396-398 and ending at the stop codonat nucleotide positions 2046-2048 (Figure 81). The predicted polypeptide precursor is 550 amino acids long (Figure 82). Clone UNQ257 (NA40604-1187) has been deposited with AVCC and is assigned AC deposit no. 209394.

Analysis of the amino acid sequence of the full-length PRO294 polypeptide suggests that portions of it possess significant hcmology to portions of various collagen proteins, thereby indicating that PR0294 may be collagen-like nolecule.

E=MPI 38: Isolation of cENX Clnes Encodiing uman PRO295 S...i A consensus IUA sequence was asserbled relative to other EST sequences using phrap as described in Ecanple 1 above. This consensus sequence is herein designated EIA35814. Based on the ENA35814 consensus sequence, oligarnucleotides were synthesized: 1) to identify by PCR a cDNA 20 library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PRO295.

Forward and reverse PCR primers were synthesized: forward PRprir 5'-GCAGAGOGG AGA IUCAGOXCIG-3' (SEQ ID ND:237) forward PR priner(.f2) 5' -CCCAGCAEIACTGCCAG-3' (SEQ ID N:238) forward PCR prinmer(.f3) 5'-TI3GCArIrATGGAG-3' (SED ID ND:239) forward R primer(.f4) 5'-C CGCAAAAAGCAAC-3' (SED ID ND:240) reverse P*R priner 5' -C'tCT'I3OZ Ct-3' (SEB ID NOD:241) Additionally, a synthetic oligonucleotide hybridization probe was constructed from the consensus IA35814 sequence which had the following nucleotide sequence.

hybridization probe 'AGG3' (SEB ID ID:242) In order to screen several libraries for a source of a fulllength clone, EA fran the libraries mas screened by PCR anplification with the PCR primer pairs identified above. A positive library was then used to isolate clones encoding the PRO295 gene using the probe oligonucleotide and one of the PCR prinmers.

RA for construction of the cZA libraries was isolated froman hunan fetal lung tissue. NA sequencing of the clones isolated as 145 described above gave the full-length fla sequence for PRO295 [herein designated as tL2258 (tl38268-1188)] (SEQ ID N:235) and the derived protein sequence for PRO)295.

The entire nucleotide sequence of t1N258 (tUW38268-1188) is shown in Figure 83 (SED ID ND:235). Clone UQ258 (ON38268-1188) contains a single open reading frane with an apparent -translational initiation site at nucleotide positions 153-155 and ending at the stop codan at nucleotide positions 1202-1204 (Figure 83). The predicted polypeptide precursor is 350 amino acids long (Figure 84). Clone k2P258 (INA38268-1188) has been deposited with A'=X and is assigned A'IC deposit no. 209421.

Analysis of the amino acid sequence of the full-length PRO295 polypeptide suggests that portions of it possess significant hmoology to ~.the integrin proteins, thereby indicating that PRO295 may be a novel integrin.

EAMPLE 39: Isolation of dREI Clones Ecoding Ihman PR293 The extracellular darain (EMD) sequences (including the secretion signal, if any) of from about 950 knon secreted proteins fron the Swiss-Prot public protein database were used to search expressed 20 sequence tag (EST) databases. The EST databases included public EST databases GenBank) and a proprietary EST ENA database (LIFESEB", Incyte Pharanceuticals, Pal Alto, CA). The search was performed using the computer program BLAST or BIASM2 (Altshul et al., Methods in Enzymology 266:460-480 (1996)) as a ccrparison of the ED protein sequences to a 6 frame translation of the EST sequence. Those carparisons resulting in a ~BLASr score of 70 (or in sane cases 90) or greater that did not encode known proteins were clustered and assembled into consensus I sequences with the program "phrapo (Phil Green, University of Washington, Seattle, Washington; http://bozenn.nrbt .washington.edu/phrap.docs/phrap.htmnl).

Based on an expression tag sequence designated herein as T08294 identified in the above analysis, oligonucleotides were synthesized: 1) to identify by PCR a crA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0293.

A pair of PCR prinmers (forward and reverse) were synthesized: forward PCR prier 5 -AACAAorAAGAO'ICCATCCIG-3' (SEQ ID ND:245) reverse PC priner 5' -AAACII A AC-3' (SE2 ID N3:246) Additionally, a synthetic oligonucleotide hybridization probe was constructed froman the expression sequence tag Ahich had the following 146 nucleotide sequence.

hybridization probe suCl m C C (SEQ ID ND:247) In order to screen several libraries for a source of a fulllength clone, D fran the libraries was screened by PCR.anplification with the PCR priner pair identified above. A positive library was then used to isolate clones encoding the PRO293 gene using the probe oligonucleotide and one of the ECR primers.

RNA for construction of the cIMA libraries was isolated from human fetal brain tissue. EA sequencing of the clones isolated as described above gave the full-length UlA sequence for PRO293 (herein designated as UNQ256 (ENA37151-1193)] (SE) ID ND:243) and the derived protein sequence for PRO293.

The entire nucleotide sequence of UN256 (DA37151-1193) is shown in Figures 85A-B (SE2 ID 10:243). Clone UN2256 (CNA37151-1193) contains a single open reading fraire with an apparent translational .initiation site at nucleotide positions 881-883 and ending at the stop codan after nucleotide position 3019 of SE; ID N: 243, Figures 85A-B). The predicted polypeptide precursor is 713 amino acids long (Figure 86). Clone UNQ256 (ENA37151-1193) has been deposited with ACC and is assigned A'ICC deposit no. A'ICC 209393.

Analysis of the amino acid sequence of the full-length PRO293 :polypeptide suggests that portions of it possess significant honology to the NLRR proteins, thereby indicating that PRO293 may be a novel NlRR protein.

EXMPIE 40: Isolation of cMA Clanes Encoding man PRD247 A consensus MA sequence was assaerbled relative to other EST sequences using phrap as described in Exanple 1 above. This consensus sequence is herein designated L133480. Based on the ENA33480 consensus sequence, oligonucleotides were synthesized: 1) to identify by PCR a cEA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PRO247.

A pair of PCR priners (forward and reverse) were synthesized: forward PCR priner 5' -CAAANIAGGGCACCAAC-3' (SE2 ID NO:250) reverse =fR primer 5' TAl'IDAW1f3' (SED ID N3:251) Additionally, a synthetic oligonucleotide hybridization probe was constructed fran the EWA33480 expression sequence tag which had the following nucleotide sequence.

147 hybridization probe -CACCII-3' (SED ID N:252) In order to screen several libraries for a source of a fulllength clone, EM fran the libraries was screened by PCR anrrplification with the PCR prinrer pair identified above. A positive library was then used to isolate clones encoding the PR0247 gene using the probe oligonucleotide and one of the ECR priers.

RNA for construction of the cE libraries was isolated fran human fetal brain tissue. UM sequencing of the clones isolated as described above gave the full-length IA sequence for PRO247 (herein designated as U?221 (IA35673-1201) (SEQ ID ND:248) and the derived protein sequence for PRO247.

The entire nucleotide sequence of U1N221 (EUN35673-1201) is shown in Figures 89A-B (SEQ ID ND:248). Clone UNQI221 (ID235673-1201) *15 contains a single open reading frane with an apparent translational initiation site at nucleotide positions 80-82 of SED ID N:248 and ending at the stop codon after nucleotide position 1717 of SED ID ND:249 (Figures 89A-B). The predicted polypeptide precursor is 546 amino acids long (Figure 88). Clone UNQ221 (11NA35673-1201) has been deposited with A'I and is assigned A'ItCC deposit no. 209418.

Analysis of the amino acid sequence of the full-length PR0247 polypeptide suggests that portions of it possess significant homology to the densin molecule and KIAAO231, thereby indicating that PRO247 may be a novel leucine rich repeat protein.

CEAMPIE 41: Isolation of cENP Clones Eroodinm thman PED302, PRO303, PRD304, PRO307 and PRO343 Consensus IMA sequences were assabled relative to other EST sequences using phrap as described in Exanple 1 above. These consensus sequences are herein designated I1M35953, INA35955, ENA35958, 1NA37160 and INA30895. Based on the IA35953 consensus sequence, oligonucleotides were synthesized: 1) to identify by FCR a clIA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PRO302.

PCR primrers (fonzard and reverse) were synthesized: forward PCR priner 1 5' -G-T3GAtArGCTKCAIPIC3' (SEQ ID ND:263) forward PCR priner 2 5' -GCAGAGGIVICrAAGGTI-3' (SED ID N:264) reverse PCR priarer 5' -AGCICTAGACCAAI CACI-3' (SED ID ND:265) Also, a synthetic oligcnucleotide hybridization probe was constructed fran 148 the consensus DNA35953 sequence which had the following nucleotide sequence hybridization probe 3' (SEQ ID N:266) In order to screen several libraries for a source of a fulllength clone, Ck from the libraries was screened by PCR anplification with the PCR primer pairs identified above. A positive library was then used to isolate clones encoding the PR0302 gene using the probe oligonucleotide and one of the PCR primers.

RNA for construction of the cMNA libraries was isolated frao human fetal kidney tissue (LIB228) [A sequencing of the clones isolated as described above gave the full-length IA sequence for PRO302 [herein designated as tUN265 (IMA40370-1217) (SEQ ID N3:253) and the derived protein sequence for PRO302.

o *15 The entire nucleotide sequence of UNQ265 (ENA40370-1217) is shown in Figure 89 (SEQ ID NO:253) Clone UrQ265 (IEA40370-1217) contains *:oo a single open reading frane with an apparent translational initiation site at nucleotide positions 34-36 and ending at the stop codon at nucleotide positions 1390-1392 (Figure 89). The predicted polypeptide precursor is 20 452 amino acids long (Figure 90). Various unique aspects of the PRO302 protein are shown in Figure 90. Clone ulM265 (ONA40370-1217) has been deposited with the ATCC on Novenber 21, 1997 and is assigned ATOC deposit no. ATCC 209485.

Based on the INA35955 consensus sequence, oligonucleotides were synthesized: 1) to identify by PCR a cENA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0303.

A pair of PCR priers (forward and reverse) were synthesized: forward PCR primer 5' *-G3A CAOCCTAAGACATIGOC-3' (SEQ ID N3:267) reverse PCR priner 5' -GAAT~CIGAAAGCICAACIU-3' (SE ID N:268) Additionally, a synthetic oligonucleotide hybridization probe was constructed fran the consensus INA35955 sequence which had the following nucleotide sequence: hybridization probe 5' 3 AA (SEQ ID N3:269) In order to screen several libraries for a source of a fulllength clone, MA from the libraries was screened by PCR anplification with the PCR priner pairs identified above. A positive library was then used to isolate clones encoding the PRO303 gene using the probe oligonucleotide and 149 one of the PCR priners.

RNA for construction of the cNA libraries was isolated fran hutman fetal lung tissue [A sequencing of the clones isolated as described above gave the full-length DA sequence for PR0303 [herein designated as UNQ266 (MA42551-1217) (SEQ ID ND:255) and the derived protein sequence for PR0303.

The entire nucleotide sequence of UNC266 (MA42551-1217) is shown in Figure 91 (SE2 ID ND:255). Clone UNQ266 (CA42551-1217) contains a single open reading frane with an apparent translational initiation site at nucleotide positions 20-22 and ending at the stop codon at nucleotide f* positions 962-964 (Figure 91). The predicted polypeptide precursor is 314 amino acids long (Figure 92). Various unique aspects of the PRO303 protein are shown in Figure 92. Clone UNQ266 (ENA42551-1217) has been deposited on Novenber 21, 1997 with the A= and is assigned =tC deposit no. At'C 209483.

Based on the IDAN35958 consensus sequence, olignucleotides were synthesized: 1) to identify by [CR a cINA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PRO304.

Pairs of [CR priners (forward and reverse) were synthesized: forward PCR primrer 1 5' -0'IGAA1CAGAAI GWdACITCCAAG-3' (SE) ID NO:270) forward PCR primer 2 5' -CAGCCCTACMGIG-3' (SED ID NO:271) forward CR primer 3 5'-TAcIGGwI'ICAiCAA-3' (SE) ID NO:272) reverse PC prier 5' -G3'AAGCA GQGAAOXX -3 (SEQ ID NO:273) Additionally, a synthetic olignucleotide hybridization probe was constructed fran the consensus t35958 sequence which had the following nucleotide sequence hybridization probe 5'GC A -CIC r---GCAAO CAk ACAGCCCOGAGOCCGAC-3' (SEQ ID 1ND:274) In order to screen several libraries for a source of a fulllength clone, DNA fran the libraries was screened by PCR anplification with the PCR primer pairs identified above. A positive library was then used to isolate clones encoding the PRO304 gene using the probe olignucleotide and one of the PCR prinrers.

RNA for construction of the cEA libraries was isolated fran 22 week human fetal brain tissue (LIB153).

EM sequencing of the clones isolated as described above gave the full-length MA sequence for PRD3-4 [herein designated as UNQ267 150 (rA39520-1217) (SED ID ND:257) and the derived protein sequence for PRO304.

The entire nucleotide sequence of IM267 (A39520-1217) is shown in Figure 93 (SEQ ID Nl:257). Clone UNQ267 (ENA39520-1217) contains S a single open reading franre with an apparent translational initiation site at nucleotide positions 34-36 and ending at the stop codon at nucleotide positions 1702-1704 (Figure 93). The predicted polypeptide precursor is 556 amino acids long (Figure 94. Various unique aspects of the PR0304 protein are shown in Figure 94. Clone UM267 (DINA39520-1217) has been deposited with AWtC on November 21, 1997 and is assigned AI deposit no.

AWCC 209482..

Based on the ENA37160 consensus sequence, olignucleotides ware synthesized: 1) to identify by PR a cA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the ,15 full-length coding sequence for PRO307.

Pairs of PCR priners (forward and reverse) were synthesized: forward ICR priner 1 5' -OOCAG=ATICtAGOGCIC-3' (SE ID NO:275) forward PCR prinrer 2 5' -GrraMc AC 3ClAGGI-3' (SEQ ID N10:276) forward PCR.priner 3 5' -'IrACAIGAACGACAG33' (SEQ ID NO:277) reverse PCR priner 5' -CATOCATICIJWTAGAGCAAG-3' (SEQ ID N:278) Additionally, a synthetic olignucleotide hybridization probe was constructed fran the consensus CNA37160 sequence which had the following nucleotide sequence hybridization probe 5'-'TMCAGM (SEQ ID ND:279) ~In order to screen several libraries for a source of a fulllength clone, [A fran the libraries was screened by C anplification with the PCR pairs identified above. A positive library was then used to isolate clones encoding the PRO)307 gene using the probe olignucleotide and one of the PCR priners.

RA for construction of the cElA libraries was isolated fran human fetal liver tissue (LIB229).

IA sequencing of the clones isolated as described above gave the full-length IA sequence for PRO307 (herein designated as UND270 (rNA41225-1217)) (SEQ ID NO:259) and the derived protein sequence for PR0307.

The entire nucleotide sequence of UN2270 (r141225-1217) is shown in Figure 95 (SEB ID ND:259). Clone UN270 (IA41225-121 7 contains a single open reading frane with an apparent translational initiation site 151 at nucleotide positions 92-94 and ending at the stop codon at nucleotide positions 1241-1243 (Figure 95). The predicted polypeptide precursor is 383 amino acids long (Figure 96). Various unique aspects of the PRO307 protein are shown in Figure 96. Clone UNQ270 (INA41225-1217) has been deposited with A'TC on Novenber 21, 1997 and is assigned ATCC deposit no.

ATCC 209491.

Based on the ENA30895 consensus sequence, olignucleotides were synthesized: 1) to identify by PCR a cINA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0343.

A pair of PCR priners (forward and reverse) were synthesized: forward PCR priner 5' -TACAG I'aOCCCA-3' (SEQ ID NO:280) i. reverse PCR priner 5' -'OaG CX GGATLi CTCA'IGICCAGGIGGA-3' (SEQ ID N0:281) Additionally, a synthetic olignucleotide hybridization probe was constructed from the consensus IENA30895 sequence which had the following nucleotide sequence hybridization probe

TCATXAC

20 CGGC CCA GIOXCCCTACT-3' (SEQ ID NO:282) In order to screen several libraries for a source of a fulllength clone, E1A from the libraries was screened by PCR anplification with the CR prier pairs identified above. A positive library was then used to isolate clones encoding the PRO343 gene using the probe olignucleotide and one of the PCR priners.

RNA for construction of the cENA libraries was isolated from human fetal lung tissue (LIB26).

CIA sequencing of the clones isolated as described above gave the full-length ENA sequence for PR0343 (herein designated as UNQ302 (EMlA43318-1217) is shown in Figure 97 (SEQ ID N0:261) and derived protein sequence for PR0343.

The entire nucleotide sequence of UNQ302 (ENA43318-1217) is shown in Figure 97 (SEQ ID NO:261). Clone tUN302 (ENA43318-1217) contains a single open reading frame with an apparent translational -initiation site at nucleotide positions 53-55 and ending at the stop codon at nucleotide positions 1004-1006 (Figure 97). The predicted polypeptide precursor is 317 amino acids long (Figure 98). Various unique aspects of the PRO343 protein are shown in Figure 98. Clone UNQ302 (EIA43318-1217) has been 152 deposited with ATCC on November 21. 1997 and is assigned A'CXC deposit no.

A'IC 209481.

EIXRMPIE 42: Isolation of clNF ClOnes Mwoding tman PIR328 A consensus [A sequence was assarbled relative to other EST sequences using phrap as described in Exanple 1 above. This consensus sequence is herein designated ENA35615. Based an the INA 35615 consensus sequence, olignucleotides were synthesized: 1) to identify by PCR a cIEA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequace for PRO328.

Forward and reverse PCR priners were synthesized: forward PCR priner 5'-ICCIOCAGrICCTIGA7IG-3' (SEQ ID NO:285) *reverse PCR_ 5' -CCATATIGCACACCAGrAA7ITr-3' (SEBQ ID 3:286) Additionally, a synthetic olignucleotide hybridization probe was constructed fran the consensus ENA35615 sequence which had the following nucleotide sequence.

hybridization probe (SEQ ID NO:287) In order to screen several libraries for a source of a full- 20 length clone, IA fran the libraries was screened by ICR anplification with the PCR prirner pair identified above. A positive library was then used to isolate clones encoding the PR0328 gene using the probe olignucleotide and one of the PCR prinmers.

RNA for construction of the cINA libraries was isolated froman hunan fetal kidney tissue.

IA sequencing of the clones isolated as described above gave the full-length IDI sequence for PR0328 (herein designated as UNQ289 (IA40587-12131) (SEQ ID N:284) and the derived protein sequence for PRO)328.

The entire nucleotide sequence of U1Q3289 (DWA40587-1231) is shown in Figure 99 (SB ID ND:283). Clone UN.289 (EM40587-1231) contains a single open reading frane with an apparent translational initiation site at nucleotide positions 15-17 and ending at the stop codan at nucleotide positions 1404-1406 (Figure 99). The predicted polypeptide precursor is 463 amino acids long (Figure 100). Clone 1U2289 (EN140587-1231) has been deposited with AMCC and is assigned AC deposit no. AIC 209438.

Analysis of the amino acid sequence of the full-length PR0328 polypeptide suggests that portions of it possess significantly homology to the human glioblastarna protein and to the cysteine rich secretory protein 153 thereby indicating that Pi328 ny be a novel glioblastare protein or cysteine rich secretory protein.

EXWMT 43: Iaolatin of cIta Clones Encoding amun P35, PRO331 or PR0326 A consensus MA sequence was assembled relativeto other EST sequences using phrap as described in Exarple 1 above. This consensus sequence is herein designated E?936685. Based on the EM36685 consensus sequence, and Incyte EST sequence no. 2228990, oligonucleotides wre synthesized: 1) to identify by PCR a cIN library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0335, PROJ331 or PR0326.

Forward and reverse PCR printers wre synthesized for the determination of PRO335: forwrd P. prirer 5' -GGAACO3AACIr'P4GTA-3' (SE2 ID 294) forard I priuue 5' -CCTAAAtk'GACAIA MA-3' (SE2 ID :295) forward PCR printer 51 -G1OC7GAGACAIXGAACT-3' (SEQ ID ND):296) forward PCR prirer 5' -ACAGI'InCAAGC1CAGAAAA=-3 (SE2 ID NO:297) reverse PQC primer 5' -CATicAGrATAAAAa'rc-3' (SEQ ID ND:298) reverse 1~ primer 5' -G09IITUGGIGAAM (SF2 ID lN:299) reverse ICR primer 5' 3 (SE2 ID NJ:3 00) Additionally, a synthetic oligonucleotide hybridization probe was *i constructed for the determination of PR0335 whAich had the following nucleotide sequence.

hybridization probe 5AC-3' (SD2 ID ND:301) Forward and reverse PCR prinrs were synthesized for the determination of PR0331: forward IC printe S' 31'I1KAAAt ITCA(IC1AU33' (SEQ ID NO: 302) reverse PCR prinue 5' -3CC1C IttC'ImrtCC-' (SF22 ID NDD:303) Additionally, a synthetic oligonucleotide hybrization probe was constructed for the determination of HO331 which had the following nucleotide sequence hybridization probe 3 (SX22 ID NO:304) Forward and reverse PCR primers were synthesiied for the determination of PR0326: forward primr 5' -AC-3CAPDIAA13MTI'IC3' (S D ID p13:305) reverse PC. primer 5' -Tr~aCAO mra~as~x~CAA C (SE2 ID ND:306) Additionally, a synthetic olignucleotide hybrizaticon probe was constructed 154 for the detennination of PRO331 which had the following nucleotide sequence hybridization probe (SEQ ID NO: 307) In order to screen several libraries for a source of a fulllength clone, IE frman the libraries was screened by PCR anplification with the PCR primer pairs identified above. A positive library was then used to isolate clones encoding the PR0335, PRO331 or PRO326 gene using the probe olignucleotide and one of the ICR prinmers.

MIA for construction of the clIA libraries was isolated froman buman fetal kidney tissue (PR0335 and PR0326) and hunan fetal brain (PR0331).

1NA sequencing of the clones isolated as described above gave the full-length [NA sequence for PRO335, PRO331 or PRO326 (herein designated as SEQ ID NOS:288, 290 and 292, respectively; see Figure 103A-B, 105 and 107, respectively), and the derived protein sequence for PRO335, PR0331 or PR)326 (see figures 104, 106 and 108, respectively; and the derived protein sequence for PRO335, PRO331 or PRO326 (see Figures 104, 106 and 108, respectively; SEQ ID NOS:289, 291 and 293, respectively).

entire nucleotide sequences are shown in Figures 103A-B, 20 105 and 107, deposited with the A'IC on June 2, 1998, Novenber 7, 1997 and .e 0 Novenber 21, 1997, respectively.

Analysis of the amino acid sequence of the full-length PRO335, 00 0.

0 PR0331 or PRO326 polypeptide suggests that portions of it possess significant horology to the LIG-1 protein, thereby indicating that PRO335, 0 25 PR0331 and PRO326 may be a novel LIG-1-related protein.

ME E 44: Isolationa of CEN& Clanes Ekooding Banman PR1332 S Based upon an ED hamlogy search perforned as described in Exanple 1 above, a consensus (WA sequence designated herein as ENA36688 was 30 assaebled. Based on the IUW36688 consensus sequence, oligonucleotides were .0 synthesized to identify by PCR a cIMa library that contained the sequence of interest and for use as probes to isolate a clone of the full-length coding sequence for PRO332.

A pair of PCR priners (forward and reverse) were synthesized: 5' -oUTnGO03AGxACrFrC-3' (SEQ ID NO:310) -O3)ZCAC3I'CCT1O3AAATG-3' (SEQ ID NR:311) A probe uas also synthesized: -,It c'AIxCrc ACAGODCACn-3 (SED ID NO:312) In order to screen several libraries for a source of a full- .155 length clonie, ota frcmi the libraries was screened by .FM anplification with the pc priser pair identified above. A positive library was then used to isolate clones encoding the PR0332 gene using the probe olignucleotide and one of the FCR pril~rs.

RNA for construction of the cWA libraries was isolated frcin a human fetal liver library (LIB229).

ENA sequencing of the clones isolated as described above gave the full-length EN. sequence for I1M40982-1235 and the derived protein sequence for PR03 32.

The entire nucleotide sequence of MJU4982-1235 is shown in Figures 109A-B (SE)Q ID Nfl:308). Clonie EN40982-1235 contains a single open reading frarm (with an apparent -translational initiation site at nucleotide positions 342-344, as indicated in Figures 109A-B). 7he predicted polypeptide precursor is 642 amxino acids long, and. has a calculated nolecular weight of 72,067 (pI:6.60). Clone L1M40982-1235 has been deposited with ICC and is assigned A~TC deposit no. A=X 209433.

Based on a BLAST and FastA sequence aligmnt analysis of the full-length sequence, PR0332 shows about 3 0-40% amino acid sequence identity with a series of ko proteoglycan sequences, includiing, for exanple, fibrznodulin and f ibrcmod-auin precursor sequence of various species (F4)DBOVIhT, FMODC1HICK, FVMRAT, E'FVlDMOUSE, FK)DJUM@N, PJR36773), osteacridin sequences (ABOO1llC1, AB007848_1), decorin sequences (CFU83141-1, OCU033941, P..R42266, P _R42267, P R42260, P..R89439), keratan sulfate proteoglycan~s (B'1U4836..L1, AF02289...l), corneal proteoglycan (AF022256...l), and bone/cartilage proteoglycans and proteoglycane precursors (PGs1.ROfl, PS2XSE, F=SJUtMAN).

EWVME 45: Isolation of cEN& Cainesn coding Iamn PRO334 A consensus EML7 sequence uns asserrbled relative to other EST sequences using phrap as described in Eoanple 1 above. Based on the consensus seqence, oligonucleotides were synthesized: 1) to identify by PR~ a ct1M library that contained thie sequence of interest, and 2) for use as probes isolate a clone of the full-length coding sequence for PR0D334.

Forward and reverse PC prizvrs were synthesized for the determination of PRO334: forward P pRiiL 5, -GA'1omTc~raT0A~GMCXX'-3' (SEXQ ID ND: 315) reverse pCRir 5' -TIrC ICaCc~AL~mcr-3' (SE ID ND: 316) Additionally, a synthetic olignucleotide hybridization probe was constructed for the detenmination of PR~O334 which had the following 156 nucleotide sequence.

hybridization probe -CGGAI AG TCAGT-3' (SEQ ID N0:317) In order to screen several libraries for a source of a fulllength clone, ENA fron the libraries was screened by PCR anplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PR0334 gene using the probe oligonucleotide and one of the PCR primers.

Human fetal kidney cEA libraries used to isolate the cENA clones were constructed by standard methods using cannercially available reagents such as those fran Invitrogen, San Diego, CA.

ENA sequencing of the clones isolated as described above gave the full-length MNA sequence for PR0334 (herein designated as MA41379- 1236] (SEQ ID ND:313) and the derived protein sequence for PRO334.

The entire nucleotide sequence of INA41379-1236 (also referred to as UNQ295) is shown in Figure 109 (SEQ ID N0:313). Clone ENA41379-1236 contains a single open reading frane with an apparent translational initiation site at nucleotide positions 203-205 and ending at the stop codon at nucleotide positions 1730-1732 (Figure 109). The predicted 20 polypeptide precursor is 509 amino acids long (Figure 110) Clone ENA41379-1236 has been deposited with A'TC and is assigned ATCC deposit no.

ATCC 209488.

Analysis of the amino acid sequence of the full-length PR0334 polypeptide suggests that portions of it possess significant hcnology to the fibulin and fibrillin proteins, thereby indicating that PRO334 may be a novel nmenber of the EBF protein family.

EXW.M LE 46: Isolation of cEA Clones Enooding Hanan PR0346 A consensus EMA sequence was identified using phrap as described in Exanple 1 above. Specifically, this consensus sequence is herein designated ENA38240. Based on the ENA38240 consensus sequence, olignucleotides were synthesized: 1) to identify by PCR a cENA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length PR0346 coding sequence.

RNA for construction of the cUMA libraries was isolated fran human fetal liver. The cEA libraries used to isolate the cEA clones were constructed by standard methods using camercially available reagents Invirtogen, San Diego, CA; Clontech, etc). The cMA was primed with oligo dr containing a NotI site, linked with blunt to SalI hemikinased 157 adaptors, cleaved with NotI, sized appropriately by gel electrophoresis, and cloned in a defined orientation into a suitable cloning vector (such as pRKB or pRKD; pRKB is a precursor of pRK5D that does not contain the SfiI site; see, Hones et al., Science,253:1278-12 8 0 (1991)) in the unique XhoI and NotI sites.

A cVA clone was sequenced in entirety. The entire nucleotide sequence of EI44167-1243 is shown in Figure 111 (SEQ ID NO:318). Clone MA44167-1243 contains a single open reading frame with an apparent translational initiation site at nucleotide positions 64-66 (Fig. 113; SEQ ID ND:318). The predicted polypeptide precursor is 450 amino acids long.

Clone IA44167-1243 has been deposited with AME and is assigned ACC deposit no. AITCC 209434 (designation MA44167-1243).

Based on a BLAST, BALST-2 and FastA sequence alignrent analysis (using the ALIGN carputer program) of the full-length sequence, PR0346 shows amino acid sequence identity to carcinoenbryonic antigen The oligonucleotide sequences used in the above procedure were the following: 0LI2691 (38240.fl) -GATOI7AAAOA M (SEQ ID NO:320) OLI2693 (38240.rl) 3' (SEQ ID N3:321) OLI2692 (38240.pl) 5' -CICCI DAGA (SEQ ID ND:322) EMPLE 47: Isolation of dIbA Clones Encoding ramn PRO268 A cansensus MlA sequence was asserbled relative to other EST sequences using phrap as described in Exanple 1 above. This consensus sequence is herein designated EIA35698. Based on the ENA35698 consensus sequence, olignucleotides were synthesized: 1) to identify by PCR a cIMA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0268.

Forward and reverse PCR primers were synthesized: forward PCR priner 1 5' -33A(a.'1t3cAAOOXXAAA'IG-3' (SEQ ID NO:325) forward PCR prner 2 5 -TAIIGGATAGGACGIUC-3' (SEQ ID N1:326) forward PCR primer 3 5' -'3IAGT1tNICAGITAGA.T1-3' (SED3 ID INO:327) reverse PR pri er .5'-TrGAAG cAAA AATfl AC-3' (SE) ID NO:328) Additionally, a synthetic olignucleotide hybridization probe was constructed frztan the consensus iEA35698 sequence which had the following nucleotide sequence 157a hybridization probe 3' (SEQ ID E:329) In order to screen several libraries for a source of a fulllength clone, IiA fran the libraries was screened by PCR anplification with the PCR priner pair identified above. A positive library was then used to isolate clones encoding the PR0268 gene using the probe olignucleotide and one of the PCR printers.

RNA for construction of the cUMA libraries was isolated fromn human fetal lung tissue. INA sequencing of the clones isolated as described above gave the full-length INA sequence for PR0268 (herein designated as UNQ235 (ENt39427-1179)] (SEQ ID N:323) and the derived protein sequence for PRO268.

The entire nucleotide sequence of UlN235 (IUA39427-1179) is shown in Figure 113 (SEQ ID 13:323). Clone tTM235 (RNA39427-1179) contains 15 a single open reading frame with an apparent translational initiation site gat nucleotide positions 13-15 and ending at the stop codon at nucleotide positions 853-855 (Figure 113). The predicted polypeptide precursor is 280 amino acids long (Figure 114). Clone UNQ235 (IA39427-1179) has been deposited with A'ICC and is assigned ATCC deposit no. A'ICC 209395.

20 Analysis of the amino acid sequence of the full-length PR0268 polypeptide suggests that it possess significant hanology to protein disulfide iscnprase, thereby indicating that PlR268 may be a novel protein disulfide isonerase.

EKAMPME 48: Isolation of c2[M Clones Enooding anzn PE330 A consensus IMR sequence was assembled relative to other EST sequences using phrap as described in Exanple 1 above. This consensus sequence is herein designated ElA35730. Based on the IA35730 consensus sequence, olignucleotides were synthesized: 1) to identify by PCR a cINA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PR0330.

Forward and reverse PCR priners were synthesized: forward PCR priner 1 5'-CCAG3CAATrITCAGA-3' (SD ID W:332) forward PCR priner 2 5' -GOCrT- 3' (SE ID N:333) forward PCR priner 3 5' -G7ICICAAGAACTI CWIC-3' (SEQ ID D: 334) reverse ICR priner 5' -ACACIAaCATIO CItTarACG-3' (SEQ ID N:335) Additionally, a synthetic olignucleotide hybridization probe was constructed fron the ccnsensus which had the following nucleotide sequence 157b hybridization probe -3'ACAAICIA (SEQ ID Ni:336) In order to screen several libraries for a source of a fulllength clone, EA fran the libraries %as screened by PCR anplification with the PCR primer pair identified above. A positive library was then used to isolate clones encoding the PRO330 gene using the probe olignucleotide and one of the PCR priners.

IM for construction of the cNA libraries was isolated from human fetal lung tissue. IMIA sequencing of the clones isolated as described above gave the full-length INA sequence for PR0330 (herein designated as UNQ290 (IA40603-1232) (SEQ ID 10:330) and the derived protein sequence for PRO330.

IThe entire nucleotide sequence of U1290 (EN40603-1232) is shown in Figure 115 (SEQ ID N0:330). Clone MQ290 (IIA40603-1232) contains a single open reading frame with an apparent translational initiation site at nucleotide positions 167-169 and ending at the stop codon at nucleotide positions 1766-1768 (Figure 115). The predicted polypeptide precursor is 533 amino acids long (Figure 116). Clone UNQ290 (DIA40603-1232) has been deposited with ATr and is assigned AIC deposit no. ATCC 209486 on Novenber 21, 1997.

Analysis of the amino acid sequence of the full-length PR0330 polypeptide suggests that portions of it possess significant homnology to nouse prolyl 4-hydroxylase alpha subunit protein, thereby indicating that PRO330 may be a novel prolyl 4-hydroxylase alpha subunit polypeptide.

.E AMPIE 49: Isolation of cEM Clones raooding Sman PRO310 A consensus ImA sequence was assentled relative to other EST sequences using phrap as described in Ehaple 1 above. This consensus sequence is herein designated IE40553. Based on the IA40553 consensus sequence, olignucleotides were sythesized: 1) to identify by PCR a cENA library that contained the sequence of interest, and 2) for use as probes to isolate a clone of the full-length coding sequence for PRO310.

Forward and reverse PCR priners were synthesized: forward PCR priner 1 5' -IOCAAGXTPI!CTAGACGOOG-3' (SEQ ID ND:341) forward PCR priner 2 5' -C3IIrI'-IT 'JC02 3' (SEQ ID ND:342) reverse PCR prier 5' -CAAATAAGGCOGTATACCCC-3' (SE ID E:343) Additionally, a synthetic olignucleotide hybridization probe was constructed fran the consensus which had the following nucleotide sequence 157c hybridization probe GIGnIC ACA-3 (SEQ ID ND: 344) In order to screen several libraries for a source of a fulllength clone, IMA from the libraries was screened by PCR anplification with the ICR priimer pair identified above. A positive library was then used to isolate clones encoding the PRO310 gene using the probe olignucleotide and one of the PCR primers.

RA for construction of the cEMA libraries was isolated from human fetal liver tissue. NA sequencing of the clones isolated as described above gave the full-length EIA sequence for PRO310 (herein designated as (EA43046-1225)] (SEQ2 ID ND:339) and the derived protein i* sequence for PRO310 (SEQ ID NO:340).

The entire nucleotide sequence of (EM43046-1225) is shcwn in Figure 119 (SEQ ID ND:339). Clone (EA43046-1225) contains a single open reading frame with an apparent translational initiation site at nucleotide positions 81-83 and ending at .the stop codon at nucleotide positions 1035- 1037 (Figure 119) The predicted polypeptide precursor is 318 amino acids long (Figure 120) and has a calculated wolecular weight of approximately 36,382 daltons. Clone (EVA43046-1225) has been deposited with AIC and is assigned A'ICC deposit no. A'ITCC 209484.

Analysis of the amino acid sequence of the full-length PRO310 polypeptide suggests that portions of it possess harology to C. elegans ]proteins and to fringe, thereby indicating that PRO310 may be involved in developrent.

ECAMPIE 50: Isolation of dIa clones mcodwing am P1O339

C

An expressed sequence tag (EST) tIA database (LIFESEr'

M

Incyte Pharmaceuticals, Palo Alto, CA) was searched and ESTs were identified. An assaubly of Incyte clones and a consensus sequence was formed using phrap as described in Enple 1 above.

Forward and revers PCR primers were synthesized based upon the assanbly-created consensus sequence: forward PCR prirer 1 5'-G CAIOCAGUIMIGICICA'Ima3-3' (SEQ ID NO:345) forward PCR primer 2 5'-(C'CCMXWIJCGCCG3I-3' (SED ID NO:346) forward PCR primer 3 5' -GIGIGACACAGCGI'G=-3' (SEQ ID N1:43) forward PCR prinmer 4 5'-GACtCAGT3tIt ln-3' (SEQ ID ND:44) reverse PCR primer 1 5' 3' (SEB ID reverse PCR prinrer 2 5' -CIX331GC-3' (SE ID M3:46) Additionally, a synthetic olignucleotide hybridization probe was M: 1Bkrot%\Kepis*cL931786*2.dc 1tS10400 157d OS @0 0 00 OS 0

S

0@ 00 0 0 0000 0 @0 0 5* 0000 constructed fran the consensus sequence which had the following nucleotide sequence.

hybridization probe -CALA I'ID ICI-3 (SEQ ID N: 47) In order to screen several libraries for a source of a fulllength clone, WA from the libraries was screened by PCR anplification with the CR priner pairs identified above. A positive library was then used to isolate clones encoding the PCR339 gene using the probe oliganucleotide and one of the PCR priners.

RNA for constructing of the cJA libraries was isolated fran huaman fetal liver tissue. A cI1E clone was sequenced in entirety. The entire nucleotide sequence of EM43466-1225 is shown in Figure 117 (SED ID NO:337). Clone Ir43466-1225 contains a single open reading frane with an apparent translational initiation site at nucleotide positions 333-335 and 15 ending at the -stop codon foind at nucleotide positions 2649-2651 (Figure 117; SEQ ID N:337). The predicted polypeptide precursor is 772 amino acids long and has a calculated nolecular weight of appraxinately 86,226 daltons. Clone EN43466-1225 has been deposited with AICC and is assigned A'C deposit no. A'IC 209490.

20 Based on a BLAST and FastA sequence alignrment analysis (using the ALIGN carputer program) of the full-length sequence, PR0339 has hanology to C. elegans proteins and collagen-like polymer sequences as well as to fringe, thereby indicating that PR0339 nay be involved in develorpnent or tissue growth.

EAMPIE 51: Isolation of cEA Clnes IEncoding thman PRO244 A consensus IR sequence was assewbled relative to other EST sequences using phrap as described in Eanple 1 above. Based on this consensus sequence, oligonucleotides were synthesized to identify by fCR a ct library that contained the sequence of interest and for use as probes to isolate a clone of the full-length coding sequence for PR0244.

A pair of PCR priners (forward and reverse) were synthesized: TirOIGAn3*3 (30923.fl) (SEQ ID NO:377) 5'-T3ATCnI.CcATI'TACAAAaUX--3' (30923.rl) (SEQ ID N:378) A probe was also synthesized: TrrQ-3' (30923.pl) (SED ID NO: 379) In order to screen several libraries for a source of a full-length clone, Ia fran the libraries was screened by PCR anplification with the ECR 157e priner pair identified above. A positive library was thena used to isolate clones encoding the PRO244 gene using the probe olignucleotide and one of the PCR priners.

RNA for construction of the cENA libraries was isolated from human fetal kidney library. ENA sequencing of the clones isolated as described above gave the full-length MA sequence and the derived protein sequence for PRO244.

The entire nucleotide sequence of PRO244 is shown in Figure 121 (SEQ ID ND:375). Clone NA35668-1171 contains a single open reading frame with an apparent translational initiation site at nucleotide positions 106- 108 (Fig. 121). The predicted polypeptide precursor is 219 amino acids long. Clone ENA35668-1171 has been deposited with ATOC (designated as o NA35663-1171) and is assigned ATC deposit no. ATOC 209371. The protein has a cytoplasmic dcmain (aa 1-20), a transmenbrane domain (aa 21-46), and an extracellular domain (aa 47-219), with *o S* a C-lectin domain at aa 55-206.

Based on a BLAST and FastA sequence alignment analysis of the AMI-length sequence, PR0244 shows notable amle add sequence identity to bepatic lectin gallus; gallus HIC hplZO-binding C-type lectin macrophagc lectin 2 (HiUMHML2-l, 41 and sequence PR32188 (44 EXML Use of PRO Polvoepde-Encodinc Nucleic Acid as Hybridization Probes The following method describes use of a nuldetde sequence encoding a PRO polypeptide as a hybridization probe.

DNA comprising the coding sequence of of a PRO polypeptide of icemst as disclosed herein may be employed as a probe or used as a basis from which to prepare probes to screen for homologous DNAs (such as those encoding naturally-occuring variants of the PRO polypeptide) in human tissue cDNA libraries or huiman tissue genomic libraries.

Hybridization and washing of filters containing either library DNAs bs performed under die following high stringency conditions. Hybridization of radiolabeled PRO polypeptide-encoding nuclcic acid-derived probe to the filters is performed in a solution of 50% formamide. 5x SSC. 0.1 SDS. 0.1 sodium pyrophosphate, 50 mM sodiumnphospate, pH 6.8, 2x Denhard's solution, and 10% dextran sulfate a4rC for 20bours. Washing of the *filters is performed in an aqueous solution of0.lIx SSC and 0.1 SDS at 420C.

DNAs having a desired sequence identity with the DNA encoding fll-length native sequence PRO polypeptide can then be identified using standard techiques known in the art.

EXAMPF ,53- Expression of PRO Polvoentides in E. coli This example illustrates preparation of an uu~lcosylated form of a desired PRO polypeptide by recombinant expression in E. coil.

The DNA sequee encoding the desired PRO polypeptde is initially amplified using selected PCR primers.

Thep pri r should cormain restriction enzyme sites which correspond to the restriction enzym sites on the selected expression vector. A variety of expression vectors may be employed. An example of a suitable vector is pBR322 (derived from E. coil; see Bolivar et al.. C&M 2:95 (1977) which contains genes for ampicillin and tetracycline resistance. The vector is digesed with restriction enzyme and depttosphorylated. The PCR amplified sequences are then ligated into the vector. The vector will preferably include sequences which encode for an antibiotic resistance gene, a trp promoter, a polyhis leader (including fte first six ST1X codons, polyhis sequence, and enterokinase cleavage site). the specific PRO polypeptide coding region, lambda transcriptional terminator, and an argli gene.

The ligation mixture is then used to tra1frm a selected E. cod strain using the methods described in Samnbrook et al., a=p. Transformants are identified by their ability to grow on LB plates and antibiotic resistant colonies ame then selected. Plasmid DNA can be isolatd anti confirmed by restriction analysis and DNA sequencing Selected clones can be grown overnight in liquid culture medium such as LB broth supplemented with antibiotics. The overnight culture may subsequently be used to irxoculate a Lurger scale culture. The cells are then grown to a desired optical density, during which the expression promoter is turned on.

After culturing the cells for several moere howa. at cells can be harvested by centrifugation. The ccli pellet obtained by the centrifuigation can be solubilized using various agents known in the amt and dhe sohibilized PRO polypeptide cun then be purified using a metal chelating colum under conditions that allow tight binding of the protein.

PROI87, PROW.17 PR0301. PR0224 and PR0238 were successfully expressed in k. coli in a poly-His taged frm using the follow*n procedure. The DNA enoding PROW,7 PROM.7 PRO3OI. PR0224 or PR0238 was initially am~plified using selected PCR primers. ilic pruners contained restrictiont enzyme sites which correspond to the restriction enzyme sites on the selected expression vector, and other useful sequncs providing for efficient and reliable translation iitation, rapid purification on a metal chelation column. and proteolyic removal with enterokinase. The PCR-amplified. poly-His tagged seqzincs were then ligated into an expression vector, whichwas used to transform an E. catf host based on strain 52 (W31 10 fuhA(tonA) Ion galE rpoHts~brpRzs) dpP(laclq).

Tnansforiams were first grown in L.B containing 50 tug/ni carbenicilliui at 30*C with shaking until an O.D.600 of' was achad. Qiltures were then diluted 50-100 told into CRAP media (prepared by mixing 3.57 9 (NH~zSO4, 0.71 g sodium citrate-2H20, 1.07 g KCI. 5.36 g Difco yeast extract 5.36 g Sheffield hycase SF in 500 tL water, as well as 110 mM MPOS, pH 7.3, 0-55 glucose and 7 mM MgSO 4 and grown for approximately 20-30' hours at 30*C with shaking. Samples were removed to verify expression by SDS-PAGE analysis. and the bulk: clture is centrifuged to pellet the cells. Cell pellets were frozen until purification and refolding.

E. coli paste from 0.5 to I L fermentations (6-10 g pellets) was resuspended in 10 volumes in 7 M guanidine. 20 mM Tris, pH 8 buffer. Solid sodium sulfite and sodium tetrathionate is added to make final *:concentrations of 0.IM and 0.02 M. respectively, and tie solution was stirred overnight at 4*C. This step results in a denatured protein with all cysteine residues blocked by sulfitolization. The solution was centrifuged at 40,000 rpn in a Beckman Ultracenaige for 30 mini. The supernatant was diluted with 3-5 volumes of metal chelate column buffer (6 M guanidine. 20 mM Tris, p1H 7.4) and filtered through 0.22 micron filters to clarify. Depending the clarified extract was loaded onto a 5 ml Qiagen Ni-NTA metal chelate column equilibrated in the metal chelate column buffer. The column was washed with additional buffer containing 50 mM imidazole (Calbiochem, Utrol grade), pH 7.4. The protein was cluted with buffer contining 250 mM imidazote. Fractions containing the desired protein were pooled and stored at VC. Protein concntration was estimated by its absorbance at 280 n using the calculated extintion coefficient based on its a2-o acid sequence.

Mce proteins were refolded by diluting sample slowly into freshly prepared refolding buffer consisting of: mM Tris, pH 8.6. 0.3 M NaCi, 2.5 M urca, 5 mM cysteine. 20 mM glycine and I mM EDTA. Refolding volumies were chosen so that the final protein concentation was between 50 to 100 naicrogranis/uil. The refolding solution was stirred gently at VC for 12-36 hour. The refolding reaction was quenched by the addition of TFA to a final concentration of 0.4% (pH1 of approximately Before further purification of the protein, the solution was filted througha 0.22 micron filter and acetonitrile was added to 2-10% final concentration. The refolded protein was duonaograplied on a Pbros RIMH reversed phase column using a mobile buffer of 0. 1% TFA -with elution with a gradient of actonitrilc from 10 to 80%. Aliquots of fractions with A290 absorbance were analyzed on SDS polyactylamide gels and fractions containing homogeneous refolded protein were pooled. Generally. the properly refolded species of most proteins are eluted at the lowet concentrations of actonitrile since those species are the most comp~act with their hydrophobic interior shielded from interaction with the reversed phase resin. Aggregated species ame usually eluted at higher acetonitie conenstrations. In addition to resolving mis folded forms of proteins from the desired form, the reversed phase step also rernovs endotoxin from the saMles.

Fractions containing the desired folded 1R0187. PROW 1. PR0301, PR0224 and PR0238 proteins.

respectively, were pooled and the aectonitrile removed using a gentle stream of nitrogen dire cted at the solution.

Proteinswere formulated into 20 mM Hepes, pH 6.9 with 0.14 M sodium chloide and 4% mannitol by dialysis or by gel filtration using G25 Superfine (Pharmacia) resins equilibrated in the formulation buffer and sterile filtered.

EXAMPEiM 54- Emss ion of PRO Pobvpepides in Mammnalian Cells This example illustrates preparation of a glycoylared form of a desired PRO polypeptide.by recombinant expression in mam-lan cells.

The vector, pRKS (see EP 307,247. published March 15, 1989). is employed as the expression vector.

~:Optionally, the PRO polypeptide-encoding DNA is ligated into pRKS with selected restriction enzymes to allow insertion of the PRO polypeptide DNA using ligation mecthods such as described in Sambrook et al., M The resulting vector is called pRX5-PRO polypeptide.

In one embodiment. the selected host cells may be 293 cells. Humtn 293 cells (ATCC CCL 1573) are grown to confluence in tissue culture pltes in medium such as DMEM supplemented with fetal -calf serum and optionally, nuitrient components and/or antibiotics. About 10 p~g pRKS-PRO polypeptide DNA is mixed with about 1 pg DNA encoding the VA RNA gem (himnmappaya et al., C&l. U:543 (1982)) and dissolved in 500 p1 of I M .:Tnis-HCI. 0. 1 m.M EDTA. 0.227 M CaCl, To this nixture is added, dropwise, 500 pl of 50 mM HEPES (pH 7.35), 20 280 mM ?4aCl, 1.5 m.M NaPO 4 and a precipitate is allowed to form for 10 minutes at 259C. The precipitate is suspended and added to the 293 cells and allowed to setle for about four howrs at 37*C. The culture mnedium is ,:aspiraed offand2ml of 20% glycerol inPBS is added for 30 seconds. The 293 cells aret(hen washed with serum free medium, fresh medium is added and the cells are incubated for about S days.

Approximately 24 hours after the transfections, the culture medium is removed and replaced with cultare mediimi (alone) or culture medium containing 200 jCi/mJ uS-cysteine and 200 pCilmI "S..methionine. After a 12 hour izubauion, the conditioned medium is collected, concentrated on a spin filter, and loaded onto a 15 SDS gel.

The processed gel m~ay be dried and exposed to film for a selected period of tirm to reveal the presence of PRO polypeptide. The cultures containing transfiected cells may undergo further incubation (n serum free mtediumli) and the medium is tested in selected bioassays.

In an alternative technique, PRO polyepide may be introduced into 293 cells transiently using the dextra sulfate method described by Somparyrac et al., Proc. Nall Acad. Set., 12:7575 (1981). 293 cells are grown to maximal density in a spinner flask aid 700jpg pRK5-PRO polypeptid DNA is added. The cells are first concentrated fromt the spinner flask by centrifugation aid washed with PBS. The DNA-dextrant precipitate is incubated on the cell pellet for four hours. The cells are treated with 20% glycerol for 90 seconds, washed with tissue culture mnedium, and re-introduced into the spinner flask containing tissue culture medium., 5 ug/nil bovine insulin and 0.1 fmnl bovine transferrin. After about four days, the conditioned mnedia is centrifuged and filtered to remove cells and debris. T1he sample containing expressed PRO polypeptide can then be concentrated and purified by any selected method. such as dialysis and/or column chromatography.

In another embodiment. PRO polypeptides can be expressed in CHO cells. The pRXS-PRO polypeptide can be transfocted into CHO cells using known reagents such as CaPO, or DEAE-dextran. As described above, the cell culture can be incubated. and the mediumn replaced with culture medium (alone) or medlium containing a radiolabel such as "S-methionine. After determining the presence of PRO polypcptide. the culture medium may be relaced with san free medium Preferably, die cultume ane incubated for about 6 days. and then the conditioned medium bs harvested. The medium containing the expressed PRO polypeptide can then be concentrated and purified by any selected method.

Epitope-tagged PRO polypeptide may also be expressed in host C10 cells. The PRO polypeptide may be subclonei out of die pRKS vector. The subclone insert can undiergo PCR to fuse in frame with a selected epitope tag such as a poly-his tag into a Baculovinis expression vector. The poly-his tagged PRO polypeptide insert can the :.be subelone d int a SV40 driven vector containing a selection marker such as DHFR for selection of stable clones.

Finally, the CHO cells can be transfecred (as describe above) wth eSOdievctrLblngmye perfirurd as describe above, to verify expression. The culture medium containing the expressed poly-His tagged PRO polypeptide can then be concentrated and purified by any selected method, such as by Nil-cfrlate affinity chromatography.

PRO21 1. PR0217. PR0230. PRO219. PR0245. PROfl1. PRO258, PRO301, PRO224. PRO2, PR0234.

PR0229, PR0223, PR0328 and PR0332 were successfully expressed in CHO cells by both a tansient and a stable expression procedure. In addition, PR0232, PR0265. PR0246, PR0228, PR0227, PR0220, PR0266. PR0269, PR0287. PR0214, PR0231. PR0233. PR0238. PR0244, PRO235. PR0236, PR0262. PR0239. PR0257.

PR0260, PR0263, PR0270, PR0271, PR0272, PRO294, PR0295, PR0293, PR0247. PR0303 and PR0268 were successfully transiently expressed in CHO cells.

Stable expression in CH0 cells was pezrformd using the following procedure. The proteins were expressed as an IgG construct (immunoadhesin). in which the coding sequence for tie soluble forms extraceiUlar domains) of the respective proteints were fused to an IgG I constant region sequence containing the hinge. CH2 and CH2 dormans and/or is a poly-His tagged form.

Following PCR amplification. the respective DNAs were subeloned in a CHO expression vector using standard tehniques as described n Ausubel et al.. Current Protocols ofMolecular Biology, Unit 3.16, John Wiley and Sons (1997). CHO expression vectors ar constructed to have compatible restriction sites 5' and 3' of the DNA of interest to allow the convenient shuttling of cIDNA's. The vector used expression in CHO cells is as describedin Lucas el al.. ?Jucl. Adds Res. 9 (1774-1779 (1996), and uses the SV40 early promnoter/enhancr to drive expression of the cDNA of interest and dihydrofolate reductase (DHFR). DHFR expression permits selection for stable maintenance of the plasmid following transfection.

Twelve micrograms of the desired plasmid DNA were introduced into approximately 10 million CHO cells using coamtialY available trnfleedon reagents Superfiect (Quiagen), Dosper' or Pugene' (Boebringr Mannheim).

Tee cells were grown and described in 11- ae al. supra. Approximately 3 x ltO" cells are frozen in an ampule for fiurthr growth and production as described below.

The ampules containing the plassnid DNA were thawed by placement into water bath and mixed by vorexing. The contents were pipettod itto a centrifuge tube conting 10 wi-s of yvdia and centrifuged at 1000 rpm for 5 minutes. The superuatant was aspirated and the cells were resuspended in 10 nML of selective media (0.2 pm filtered PS20 with 5%S 0.2 Pm dlallltered fecal bovine serum). The cells were then aliquoged into a 100 niL spinner containing 90 niL of selctive media. After 1-2 days. the cells were transferred into a 250 niL spinner filled with 150 nLadcant growth medium and incubated at 37*C. After another 2-3 days. a 250 niL, 500 niL and 2000 niL spinners wee seeded with 3 x lOcells/niL. The cell media was exchanged with fresh media by centnfugation and resuspension in production medium. Although any suitable CHO media may be employed, a production nmedim described in US Patent No. 5,122,469. issued June 16. 1992 was actually used. 3L production spinner is seeded at 1.2 x 10' cells/niL On day 0, the cell numiber PH1 were determined. On day 1. the spin=~ was sampled and sprging with filtered air was wrnmucd. On day 2, the spinner was sampled. the teniperature shifted to 33 C. and mL of 500 gIL glucose and 0.6 niL of 10% antifoarn 35 polydiothyhloxane emulsion. Dow Corning 365 Medical Grade Emulsion). Throughout the production. pH was adjuted as necessary to keep at around 7.

After 10 days, or until viability dropped below 70%, the cell culture was harvested by centrifugion and filtering through a 0.22 pm filter. The filtrate was either stored at 4*C or immediately loaded onto columns for purification.

IS For the poly-His tagged constructs, the proteins were purified using a Ni-NTA column (Qiagen). Before purification, imidazole was added to the conditioned media to a concentration of 5 mM. The conditioned mnedia was pumped onto a 6 ml Na-NTA cobumn equilibrated in 20 mM Hepes. pH 7.4, buffer containing 0.3 M NaCI and 5 mM imidazole at a flow rate of 4-5 mi/mmn. at 4*C. After loading, dhe column was washed with additional equilibration baffer and the protein ehired with equilibration buffer conitaining 0.25 Md imidazole. The highly purified protein was subsequntly dsatd into astorage buffer containing 10 mM Hepes, 0.14 Md NaCl and 4% niannitol, pH 6.8, with a 25 ml G25 Superfine (Pharmacia) column and stored at :Immunoadhesin (Fc containing) constructs of were purified from the conditioned media as follows. The conditioned medium was pumped onto aS5 ml Protein A column (Pharmacia) which had been equilibrated in 20 mM Na phosphate buffer, PH 6.8. After loading, the column was washed extensively with equilibration buffer before elution with 100 mM citric acid, pH 3-5. The eluted protein was immediately neuttalized by collecting 1 ml fractions into tubes containing 275 pLL of I M Tris buffer, pH 9. Thbe highly purified protein was subsequently detalted into storage buffer as described above for the poly-His tagged proteins. The homogeneity was assessed by SDS polyacrylamiide gels and by N-terminal amino acid sequencing by Edmnan degradation.

PRO211. PROW,7 PR0230. PR0232. PROIS7, PROW6, PR0219, PRO246, PR0228. PR0533, PR0245.

PR0221. PR0227. PR0220. PR0258, PR0266, PR0269, PROW9, PR0214, PR0317. PRO301, PR0224.

PR0222, PR0234. PR0231, PR0229, PR0233, PR0238, 1110223. PROW.5 PR0236. PR0262, PR0239.

PR0257. PR0260, PROW6. PR0270. PR0271, PR0272, PR0294, PR0295, PR0293. PROW4, PR0304.

PR0302, PR0307. PR0303. PROW4. PR0328. PR0326, PR0331, PR0332. PR0334, PR0346, PR0268.

PR0330, PRO310 and PR0339 were also successfully transiently expressed in COS cells.

EXAMPEL.Ei: Piplessiml of PRO Polvperidet in Yeasi The following method describes recombinant expression of a desired PRO polypeptide in yeast.

lFirst, yeast expression vectors are constucated for intracellular production or secetion of PRO polypeptides from the ADH2IGAPDH pro moter. DNA encoding a desired PRO potypeptide, a selected signal peptide and the promoter isinsisetd into suitable restriction enzyme sites in the selected plasmid to direct intracelluar expression of' the PRO polypeptide. For secretion. DNA encoding the PRO polypeptid& can be cloned into the silected plasmid, together with DNA encoding the ADH2IGAPDH promoter. the yeast alpha-factor secretory signal/leader sequence.

and linker sequeccs (if needed) for expression of die PRO polypeptide.

Yeast cells, such as yeast strain ABD110. can then be transfonmed. with the expression plasruids described above and cultured in selected fermentation media. The transformed yeast supernatants can be analyzed by precipitation with 10% trichloroacetic, acid and separation by SDS-PAGE. followed by staining -of the gels with Coomassie Blue stain.

Recombinan PRO polypeptide can subsequently be isolated and purified by removing the yeast cells from the fertnezuation medium by centrifugation and then concentrating the medium using selected cartridge filters. The coetrate containing die PRO polypeptide may further be purified using selected column chromatography resins.

EXAMPLL 5: Exprerssion of PRO Pobvpotdes in Biculovirus-Infected Insec Cells T7he following mdbod describes recombinant expresion of PRO polypeptides in Baculovirus-infected insect cells.

fle desired PRO polypeptide is futsed upstream of an epitope tag containe with a baculovinu expression *20 vector. Such epitope tags include poly-his tags and imnennoglobulin tags (like Fe regions of IgG). A variety of plasmids may be employed, including plasmids derived from oommerialy available plasmids such as pVL1393 :(Novagen). Briefly. the PRO polypeptide or the desired portion of the PRO polypeptide (such as the sequence enoigdexacellular domain of a transzcmbrne protein) is amplified by 7CR with primers complementary t the 5' and 3' regions. The 5' primer may incorporate flanking (selected) restriction enzyme sites. The product is then digested with those selected restriction enzymes and subeloned into the expression vector.

Recombinant baculovimus is generated by co-transfecting the above plasmid and BaculoGold'u virus DNA (Pharmingeri) into Spodopterafliegiperda (*Sf9) cells (ATCC CR1. 1711) using lipofectin (comnmercially available from GIBCO-BRL). After 4-5 days of incubation at W8C, the released viruses are harvested and used for further amplifications. Viral infction and protein expression is performed as described by O'Reilley et al., Baculovirus expression vectors: A laboratory Manual, Oxford: Oxford University Press (1994).

Expressed poly-his tagged PRO polypeptide can then be purified, for example, by Ni 2 -chclate affinity chromatography as follows. Extracts are prepared from recombinant virus-infected Sf9 cells as described by Rupert er al.. Ndure, M:175-179 (1993). Briefly. Sf9 cells are washed, resuspended in sonication buffer (25 niL Ilepes, PH 7.9; 12.5 mM MgCl,; 0.1 mM EDTA; 10% Glycerol; 0.1 S NP-40; 0.4 M KCI), and sonicated twice for seconds on ice. The sonicates are cleared by centifugation. and the supernatant is diluted 50-fold in loading buffer mM phosphate, 300 mM NaCI. 10% Glycerol, pH 7.8) and filtered through a 0.45 jum filter. A Nil-NTA agarose column (commercially available from Qiagen) is prepared with a bed volume of 5 mL. washed with 25 mL of water and equilibrated with 25 mL of loading bufe. The filtered cell extract is loaded onto the column at 0.5 mL per minge. The column is washed to baseline A2, with loading buffer, at which point fraction collection is started.

Next. the column is washed with a secondary wash buffer (S0 mM phosphate; 300 mM NaCI, 10% Glycerol. pH which elutes nonspecifically bound protein. After reaching Aobaselit again, the column is develope with a 0 to 500 mM Irnidazole gradient -in the secondary wash buffer. One roL fractions are collected and analyzed by SDS-PAGE and silver staining or western blot with Ni'*NTA-conjgatrd to alkaline phospfitae (Qiagen).

Fractions containing die eluted His4-tagged PRO polypeptide are poole and dialyzed against loadirg buffer.

Alternatively. purifiation of the ISg tagged (or Pc tagged) PRO polypeptide cant be performed using known chromatography techniques, including for instance. Protein A or protein G column chromatography.

PRO21I1, PR0217. PROM3. PRO0187. PROM6, PROM4, PR0228, PR0533. PROM4. P10221. PR0220.

PRO258, PROM,6 PR0269, PR0287, PR0214. PRO3OI, PR0224, PR0222. PR0234, PR0231, PR0229, PROM3. PROM3. PROM,7 PROM.2 PROM9. PR0295. PR0328. PR0326. PROM3, PR0334. PR0346 and PR03 10 were successfully expressed in baculovints infected M1 or highS insect cells. While die cxprtssion was actually performed in a 0.5-2 L scale, it can be readily scaled up for larger 8 L) preparations. The proteins were expressed as an IgG construct C miaffisin), in which te protein extracellular region was fursed to an IgGl constant region sequence containing the hinge. C112 and CH3 domains and/or in poly-His tagged forms.

Following PCI amplifcation, dhe respective coding sequences were subcloned into a baculovirus expression vector (pb.PH~gG for IgG fusions and pb.PH.His.c for poly-His tagged proteins), and the vector and Baculogold* baculovirus DNA (Pharmingen) were co-transfected into 105 Spodoptcrfrugiperda cells (ATCC CRL 1711). using Lipofectin (Gibco BRL). pb.PH.lgG and pb.PH.His are modifications of the cornercially available baculovirus expression vector pVL1393 (Ptrarrningen). with modified poIylinker regions to include the His or Fc tag sequences. The cells were grown in link's TNM-FH niediurn supplemented with 10% FBS (Hyclone). Cells were :incubated tor 5 days at 2WC. The supernatant was harvested and subsequently used for the first viral amplification by infecting Sf9 cells in Hink's TNM-FH medium supplemented with 10% PBS at an approximate inultiplicity of infection (MOI) of 10. Cells were incubated for 3 days at W8C. The supernatant was harvested and the expression of die constructs in the baculovirus expresion vector was determined by batch binding of 1 ml of supernatant to mL. of Ni-NTA beads (QIAGEN) for histidine tagged proteins or Protein-A Sepharose CL-AD beads (Pliarmacia) for IgG tagged proteins followed by SDS-PAGE analysis comnpauing to a known concentration of protein standard by Coomassie blue staining.

77he first viral amplification supernatant was wsed to infect a spinne culture (500 ml) of Sf9 cells grown in ESF-921 medium (Expression Systems UPC at an approximate MO! of 0.1. Cells were incubated for 3 days at 290C. The supernatant was harvested arnd filtered. Batch binding and SDS-PAGE analysis was repeated, as necessary, until expression of the spinner culture was confirmed.

The conditioned mediumn from the transfected cells (0.5 to 3 L) was harvested by centrifugation to remove the cells and filtered through 0.22 micron filters. For fte poly-His tagged conucts. the protein construct were purified usin a Ni-NTA cohumun (Qiagen). Before purification, imidazole: was added to the conditioned media to a concentration oif 5 mM. The conditioned media were pumped onto a 6 ml Ni-NTA column equilibrated in 20 mM Hepes. pH 7.4, buffer containing 0.3 M NaCI and 5 mM inidazole at a flow rate of(4-S mitmin, at 4*C. After loading. the cohm was was~hed with additional equilibration buffer and the protein chated 'with equilibration buffr cotaining 0.25 M iniidazole. The highly purified protein was subsequendy desalted into a storage buffer containing mM Hqpea. 0. 14 M NaCI and 4 mannutol. pH 6.8. with a 25 ml G25 Superfine (Pharmacia) column and stored at IIinmadhin (Fc contag) constnucts of proteins were purified from the conditioned media as follows.

lT conditioned media were pumped onto a 5 ml Protein A column (Phantiaia) which had been equiilibrated in mM Na phosphate buffer, p116.8. After loading, dhe cohinin was washed extensively with equilibration buffer before elution with 100 mM citric acid. pH 3.5. 71= eluted protein was immediately neutralized by collecting* I ml fractions int tubes containing 275 mL of I M Tris buffer. pH 9. The highly purified protein was subsequently desalted into storage buffer as described above for the poly-His tagged proteins. The homogeneity of dhe proteins Was verified by SDS polyactylamide gel (PEG) eleetrophotesis and N-terminal amino acid sequencing by Edinan degradation.

EXAMPLE57. Prenaration of Antibodies that fijil to PRO Polvnetides This example illustrates preparation of monoclonal antibodies which can specifically bind to a PRO polypeptide.

15 Techniques for producing the monoclonal antibodies arc known in the art and arc described. for instance.

in Goding. Imniunogens that may be employed inclue purified PRO polypeptide. fusion proteins containing the PRO polypeptide. and cells expressing rccoiznm PRO polypetide on the cell surface. Selection of the immunogen can be made by the skilled artisan withot undue experimentation.

Mice. such as Balb/c, ame immnized with the PRO polypeptide immunogen eimulsified in complete Fretnd's adjuant and injected subcutaneously or itraperitoneally in an amount from 1-100 micrograms. Alternatively, the tmuunogen is emulsified in MPL-DM adjuvant (Ribi Inmmnocheniical Research. Hamilton, MI) and- injected into .thde animal's hind foot pads. The immuinized mice are then boosted 10 to 12 days later with additional immunogen emulsified in the selected adjuvans. Thereafter, for several weeks, the mice may also be boosted with additional imuian injctions. Serum samples may be periodically obtained from the mice by retro-orbital bleeding for testing in ELISA assays to detect anti-PRO polypeptide antibodies.

After a suitable antiody titer has been dtce, the animaals 'pstve o antibodies cnbe injected with a final inrave, us injection of PRO polypeptide. Thre iD four days later, the nice are sacrificed and the spleen cells are harvested. The spleen cells ame then fused (using 35 polyethylene glycol) to a selected niurine mycloina cell liac such as P3X63AgU.l. available from ATCC, No. CRL 1597. The fusions generate hybridona. cells which can then be plated in 96 well tissue culture plates; containing HAT (hypoxanhhine. amimopterin. and thyrnidine) mediumn to inhibit proliferation of non-fused cells, mycloma hybrids, and spleen cell hybrids.

The hybridoma cells will be screened in an ELISA for reactivity against the PRO polypeptide.

Determination of "positive Iybrdoma cells secreting the desired monoclonal antibodies against the PRO polypeptide is within the skill in dhe ant.

The positive hybridama cells can be injeted intraperitoncatly into syngcneic l~b/c mice to produce ascites conaiin the anti-PRO polypeptide monoclorWa antibodies. Alternatively. the hybridorna cells can be grown in tissue culture flasks or roller bottles. purification of the monoclonal antibodies produced in the ascites can be accomplished using ammonium sulfate precipitation. followed by gel exclusion chromatography. Alternatively, affinity chromatography based upon binding of antibody to protein A or protein G can be employed.

EXAMPLP 59: Chimeric PRO Polvnenddes PRO polypeptides may be expressed as clhimrnic proteins with one or more additional polypeptide domains added to facilitate protein purification. Such purification faciltating domains include, but are notjintited to, mnetal cheating peptides such as histidine-tryptophan modules that allow purification on immobilized metals, protein A domains that allow purification on immobilized inmmoglobulin, and the domain utilized in the FLAGS'h extensioniaffinity prificaton system (Irnttnex Corp.. Seattle Wash.). The inclusion of a cleavable linker sequence such as Factor XA or enterokinase (Invicrogen, San Diego Calif.) between the purification domain and the PRO polypeptide seqzei may be useful to facilitate expression of DNA encoding the PRO polypeptide.

::XAP 59MEE.: Purification of PRO Pal3'nenides Using SR~fic Anidies Native or recombinant PRO polypeptides may be purified by a variety of standard sechniques in the art of protein purification. For example, pro-PRO polypeptide, manire PRO polypeptide. or pre-PRO polypeptide is purified by immznoaffnity chromntography using antibodies specific for the PRO polypeptide of interest. In general.

an immumoaffinity column is constructed by covalently coupling the anti-PRO polypeptide antibody to an activated chromatographic resin.

Ptolyclonal imnmoglobulins are prepared from inmme sera either by precipitation with amamonium sulfate or by purification on immobilized Protein A (Pharmacia LJCB Biotechnology, Piscataway, Liewise, monoclonal antibodies are prepared from mouse ascites fluid by anmmonium sulfate precipitation or chromatography on immobilized Protein A. Partially purified immumoglobulin is covalently attached to a chromatographic resin such as Cn~r-activazEd SEPHAROSEa4 (Pharmnacia LKB Biotechnology). The antibody is coupled to the resin, the resin is blocked, and the derivative resin is washed according to die manufacturer's instructions.

Such an immunoaffinity column is utilized in the purification of PRO polypeptidr.b rprn a fraction from cells containing PRO polypeptide in a soluble form. This preparation is derived by solubilization of the whole cell or of a subelular fraction obtained via differential centrifugation by the addition of detergent or by other methods well known in the art. Alternatively, soluble PRO polypeptide containing a signal sequence may be secreted in useful quantty into the meditum in which the cells are grown.

A soluble PRO plypeptide-containing preparation is passed over the irnmunoafflnity column. and the column is washed under conditions that allow the preferential absorbance of PRO polypeptide high ionic strength buffers in the presence of detergent). Thlen, the column is cfted under conditions that disrupt andbody/PRO polypeptde binding a low pH buffr such a proimately pH 2-3, or a high concentration of a clmnotrope such as urea or thiocyanate ion), and PRO polypeptide is collected.

EXAMLE60: DrgSrenn This invcntion is particularly useful for screening compounds by using PRO polypeptides or binding fragment thereof in any of a variety of drug screening techniques. The PRO polypeptide or fragment employed in such a oeut may cither be free in sohidon affixod to a solid support, borne on a cell surface, or located inracliularty.

One method of drug screening utilizes eukazyotic or prokaryotic host cells which ame stably tramsformed with recombi-nu cleic acids expressing the PRO polypeptide or fragnicst. Drugs ame screened against such transfortned cells in competitive binding assay&. Such cells. either in viable or fixed form, can be used for standard binding assays. Om may masure. for exanple, the formation of complexes betweens PRO polypeptide or a fragment and the agent being tested. Alternavely, one an exaine the dininution in comnplex'formation between the PlO polypeptide and its target cell or target receptors caused by the agent being tested.

TIs, die present invention provides methods of screening for drugs or any other agents which can affect a PRO potptde-associated disease or disorder. Thes methods comprise contacting such in agent with an PRO polypeptide or fr-agment thereof and assaying for the presence of a complex between the agent and the PRO polypptde or fragmnt, or (ui) for the presece of a complex between the PRO polypeptide or fragment and the cell, by methd well known in the art. In such competitive binding assays, the PRO polypeptide or fraguient is typically lx~ted. After suitable incubation, free PRO polypeptide or fragment is separated from din present in bound form, and the amount of free or wncomplexed label is a measure of the ability of the particular agent to bind to PRO polypetide or to interfere with the PRO poypeptide/cell complex.

Another technique for drug screening provides high throgput screening for compounds having suitable binding affinity to a polypeptide and is described in detail in WO 84i/03564. published on September 13, 1984.

Briefly sated, large mmihers of different small peptide test cornpcumds arm synthesized on a solid substrate, such as plastic pins or some other surface. As applied to a PRO polypeptide, the peptide test compounds arc reacted with PRO polypeptide and washed. Bound PRO polypeptide is detected by methods well known in the art. Purified PRO polypeptide can also be coated directly onto plia for use in the aforemnentioned drug screening techniques. In addition, non-netr-alizig antibodies can be used to capure the peptide and immnobilize it on the solid support.

:This invention also contemplates the use of competitive drug screening assays in which neuatralizing antibodies 'capable of binding PRO polypeptide: specifically compete with a test compouind for binding to PRO polypeptide or fragments thereof. In this manner, the antibodies can be used to detem the presence of any peptide which shares one or more antigenic determinants with PRO polypeptide.

EXAMPLE 61 Rational Drug Design The goal of rational drug design is to produce strwltural analogs of biologically active polypeptide of interest a PRO polypeptide) or of small molecules with which they interact, agonists, antagonists, or inhibitors.

Any of these examples can be used to fashion drugs which are more active or stable forms of the PRO polypeptdec or which enhance or interfere with the function of the PRO polypeptide in Wvo (cf. Hodgson. Dio/ehnology. 2: 19-21 (1991)).

In one approach, the three-dimensional structure of dhe PRO polypeptide. or of an PRO polypeptide-inbibitor comlex, is datermin by x-ray Crystallography. by computer modeling or, most typically, by a combination of the two approaches. Both the shape and charges of the PRO polypeptide must be ascertained to elucidate the structure and to dctermine active site(s) of the molecule. Less often, useful information regarding the structure of the PRO polypeptide may be gained by modeling based on the structure of homologous proteins. In both cases, relevant stuctural inforfnion is used to design analogous PRO polypepihe-like molecules or to identify efficient inhibitors.

UseiW examples of rational drug design may includ molecules which have improved activity or stability as shown by Braxton and Wells, Biohmistry. UI:7796-7801 (1992) or which act as inhibitors. agosusts. or antagonists of native peptides; as shown by A rhauda ef at.. LIm. 11,3= :742-746 (1993).

his also possible to isolate a target-specific antibody, selected by functional assay, as described above, and then to solve its crystal structure. This approach, in principle, yields a-pharmacore upon whiclisubsequemt drug design can be based. ht is possible to bypass protein crystallography altogether by generating anti-idiotypic antibodies (andnds) to a functional, pharmacologically active antibody. As a mirror image of a mirror image, the binding site of the anti4ds would be expected to be an analog of the original receptor. The ann-id could then be used to identify and isolate peptides from banks of chemically or biologically produced peptides. The isolated peptides would the act as the pharmacorc.

By virtue of the present invention, sufficient amounts of the PRO polypeptide may be made available to perform such aml)*a studies as X-ray cystaflography. In addition. knowledge ofdePRO polypeptide amino acid sequence provided herein will provide guidanc to those emnploying computer modeling techniques in place of or in addition to x-ray crystallography.

EXAMPLE Diagnostic Test Usin, PRO317 PolviietIde-Sp~eific Antibodies Particular anti-PR0317 polypeptide: antibodies arc useful for the diagnosis of prepathologic conditions, and chronic or acuite diseases such as gynecological diseases or ischeniic diseases which arc characterized by differences in t saw= or distijInuion of PRO3 17. PR0317 has been found to be expressed in human kidney and is thus likely to be associated with abnormalities or pathologies which affect this organ. Further, since it is so closely related to EBAF-1, it is likely to affect the endometrium and other genital tissues. Further, due to library sources of certain :ESTs. it appears that Pi0317 may be involved as well in forming blood vessels and hence to be a modulator of angiogcnesis.

Diagnostc tests for PR0317 include methods utilizing die antibody and a label to detect PR0317 in human body fluids, tissues, or extracts of such tissues. The polypeptide and antibodies of the present invention may be used with or without modification. Frequently the polypeptide and antibodies will be labeled by joining them, either covalently or noncovalently, with a substance which provides for a detectable signal. A wide variety of labels and conjugation techniques are known and have been reported extensively in both the scientific and patent literature.

Suitable labels includec radionuclides. enzymes, substraes. cofactors, Inhibitors, fluorescent agents, cbemiltuminescent agents, magnetic particles, and dhe like. Patents teaching the use of such labels include U.S. Pat. Nos. 3,817,837: 3.850752. 3,939,350; 3,996,345; 4,27,437; 4.275,149; and 4,366,241. Also, recombinant imunwioglobulins may be produced as shown in U.S. Pat. No. 4,816,567.

A variety of protocols for measuring soluble or membrane-bound PRo3 17, using either polyclonal or monoclonal antibodies specific for that PROW 1, are known in the an. Examples include enzyme-linked iiiuuuisorbcnt assay (EWIA). radiolimmunoassay radioreceptor assay (RFLA), and fluorescent activated cell sorting (FACS). A two-site monoclonal-basod unmunoassay utilizing monoclonal antibodies reactive to two non-interfering cpitopes on PRO3 17 is preferred, but a competitive binding assay may be employed. These assays art described, among other places. In Maddox ef al. I Ex Me., Ja:121 1 EXAMPL.E63: Identification of PRO317 Rceptors Purifid PR0317 is useflul for characterization and purification of specific cell surface receptors and other bindling momIcsc. Cells which respond to PR0317 by metabolic changes or other specific responses are likely to express a receptor for PR0317. Such receptors include, but are not limited to. receptors associated with and activated by tyrosine and serire/tiuonine kinases. See Kolodziejczyk and Hall, supra, for a review on known seceptors for tiE TGF- supmerfamily. Candidate receptors for this superfamily fall into two primary groups, termed type I and typ UI receptors. Both types ame serinchreoninc kinases. Upon activation by the appropriate ligand, type I and tye 1I receptors physically interact to form hetero-oligomers and subsequently activate intracellular signaling cascades, ultimately regulating gene trnscription and expression. la addition, TOF- binds to a thirdI receptor class, type Ml, a membrane-anchored protooglycan la-king the kinase activity typical of signal transducIng molecules.

PR0317 recepunr or other PR03 17-binding molecules may be identified by interaction with radiolabelcd 0* PRO3 17. Radioactive labels may be incorporated into PR0317 by various methods known in the art. A preferred embodiment is the labeling of primary amino groups in PR0317 with Bolton-l-unrc reagent (Bolton and Hunter, Bicen 11:529 (1973)), which has been used to label other polypeptides without concomitant loss of biological *activity (Hecbert et al.. LI n~l-Chem-. 2M:18989 (1991); McColl et al.. JInmwpl.. INA:550-4555 (1993)).

Recptor-beaing cells are incubated with labeled PRO3 17. The cells arc then washed to removed unbound PROW 1.

and teceptor-bound PR0317 is quantified. T'he data obtained using different concentrations of PR0317 are used to calculate values for the rmmber and affinity of receptors.

Labeled PR0317 is useful as a reagent for purification of its specific receptor. In one embodiment of affinity purification, PR0317 is covalenitly coupled to a chromatography column. Receptor-bearing cells arc extracted, and .:the extract is passed over the coluimn. T7he receptor binds to the column by virtue of its biological affinity for **PRO3 17. The receptor is recovered from the column and subjected to N-terminal protein sequencing. 'This amino acid sequence is then used to design degenerate oligoniuceotide probes for cloning the receptor gene.

In an alternative method, rnRNA is obtained firom receptor-bearing cells and made into a eDNA library. The library is transfectd into apopulation of cells. an ioxse cells expressing the receptor are selected using fluorescently .***labeled PR0317. The receptor is identified by recovering and sequencing recombinant DNA from highly labeled cells.

In another alternative method. antibodies are raised against the surface of receptor bearing cells, specificaly monoclmnal antibodies. Thie monoclonal antibodies awe screened to idesuify those which inhibit the binding of labeled PRO3 17. These monoclonal antibodies are then used in affintity purification or expression cloning of the receptor.

Soluble receptors or other soluble binding molecules am identified in a similar manner. L.abeled PR0317 is incubated with extracts or other appropriate materials derived from the uerus. After incubation, PR0317 complexes larger than thme size of purified PR0317 are identified by a sizing technique such as size-exclusion chromatography or density gradient centrifugation and are purified by methods known in the art. The soluble receptors or binding protein(s) are subjected to N-terminal sequencing to obtain information sufficient for database identification, if the soluble protein is known, or for cloning, if the soluble protein is unknWn.

EXMPLE 64: Determination of PRm 17-Induced C1hr RPSU=n The biological activity of PR0317 is measured. for example. by binding of an PRO3 17 of the imnvedon to an PRO317 meptor. A testcompoundis screened as ananagoniut for itsability to block binding of PR0317 to the receptor. A test comspound is screened as an agonist of the PR0317 for its ability to bind an PR0317 receptor and luflrnce dramn physiological events as PR0317 using. for example. dhe KIRA-EU1SA assay described by Sadick et Anahytical Biochemistry- 2M:207-214 (1996) in which activation of'& receptor tyrosui ckinas e is monitored by immunmo-capaire of the activated receptor and quantitation of the evel of ligand-induced phosphorylation. The assay may be adapted to monitor PRO3l7-izihjed receptor activation through die use of an PR0317 receptor-specific antibody to capture the activated receptor. These techniques ame also applicable to other PRO polypeptides described herein.

EXAMPMERE65: Use of PRO224 for Screeninr Comeounds PR0224 is expressed in a ell stripped of mzembrane proteins and capablc of expressing PR0224. Low density lipoproteins having a dectectable label are added to the cells and incubated for a sufficient time for endocytosis.

The cells ame washed. flie cells ame then analysed for label bound to the membrane and within the cell after cell lysis.

Detection of the low density lipoproreins within the cell determines that PR0224 is within the family of low density lipoprotein receptor proeinus. Memabers found within this family ame then used for screening compounds which affect these receptors, and particularly the uptake of cholesterol via these receptors.

EXAPLE66: Abilty of PRO Polvoetides to Inhibit Vascular Fviothetial Growth Factor (VEGF) Stimulated Proliferation of Endothelial Cell Grot Th ability of various PRO polypeptides to inhibit VEGF stimulated proliferation of endotheliul cells was tse.Specifically, bovine adrenal cortical capillary endothelial (ACE) cells (from primary culture, maximwin 12-14 passages) were plated on 96-well mnicrotiter plates (Aniersham Life Science) at a density of 500 ceilstwell per 100 IL in low glucose DMEM, 10% calf serum, 2 mM glutanine, lx pen/strept and fungizone. supplemented with 3 nghrnL VEOF. Controls were plated the same way but some did not include VEGF. A test sample of the PRO polypeptide of interes was added in a 100pld volume for a 200 jAl final volume. Cells were incubated for 6-7 days at 370C. The media was aspirated and the cells washed Ix with PBS. An acid phosphatase reaction mixture (100 AL, 0. IM sodium acetate. pH 5.5. 0.1 Triton-iGO, 10 mM p-nitrophenyl phosphate) was added. After incubation for 2 hous at370C. hreaction was stopped by additionof 10 pl IN NaOH. OD was measuredoanmicrotiter plate reader at 405 am. Controls were no cells. cells alone, cells FGF (5 aglImL), cells VEGF (3 nglniL), cells VEGF (3 ng/mi) TGF-3 (I ag/mI). and cells VEGF (3ng/mL) UPF (5 ng/mL). (TGF-P at a I ng/Ml concentration is known to block 70-90% of VEGF stimulated cell proliferation.) The results were assessed by calculating the percentage inhibition of VEOF (3 ng/mi) stimulated cells proliferation, determined by measuring acid phosphatase activity at 0D405 am, relative to cells without stinmlation, and relative to the reference TGF-P inhibition of VEGF stimulated activity. The results, as shown in Table 2 below, are indicative of the utility of the PRO polypeptides in cancer therapy and specifically in inhibiting tumor angiogenesis. Mwe numerical values (relative inhibition) shown in Table 2 are determined by calcuilating the pelrv inbi~on of VEGF stilmated prolilcntion by the PRO polyppide retdve to cels without sziniation and dtn dividing dim prcentage iato the prcait inhibition obtained by TGF-P at I ng/Al which is known to block of VEGF stimulated cel prolicraation.

TaL 0 10 0 .0.

PRQName PR02 I PR0211 PRO211 PRO211 PR0211 PRO2I1 PR0217 PR0217 PR0217 PR0217 PR0217 PR0217 PRO187 PRO17 PR0187 PR0219 PR0219 PR0219 PR0246 25 PR0246 PR0246 PR0228 PR0228 PR0228 PR0228 PR0228 PR0228 PR0228 PR0228 PR0228 PR0245 PR0245 PR0245 PR0245 PR0245 PR0245 PR0221 PR0221 PR0221 PR0221 PR0221 PR0221 PR0258 PR0258 PRO258 PR0258 PR0258 PRO258 PRO Concenratio 0.01% 0.01% 0.1% 1.0% 1.0% 0.01% 0.1% 1.0% 2.5 p~M 25nM 250 nM 0.01% 0.1% 1.0% 5.7 #M 57nM 570 nM 0.01% 0.1% 1.0% 0.01% 0.1%7 1.0% 0.01% 0.01% 0.1% 0.1% 1.0% 1.0% 0.01% 0.1% 1.0% 0.48 nM 4.8 nM 48 nM 0.01% 0.01% 0.1% 0.1% 1.0% 1.0% 0.01% 0.01% 0.1% 0.1% 1.0% 1.0% Reb~ive lhbido 99.0 1.09 0.95 67.0 0.27 20.0 1.06 0.84 0.39 0.2 0.88 0.58 0.91 0.82 0.44 0.61 1.09 0.97 1.04 0.49 0.99 0.93 0.57 0.95 0.98 0.77 0.88 0.16 0.48 0.76 0.35 0.11 1.03 0.95 0.49 1.03 1.06 0.82 0.93 0.31 0.43 0.98 1.06 0.95 1.02 0.6 0.69 Table 2 cnt' PRO3ON PRO301 PRO301 PRO301 PRO301 PRO301 PR0224 PR0224 PRO224 PR0272 PRO272 PRO272 PR0328 PRO328 PRO328 PR0331 PR0331 PR0331 PRO Conrenrnion 7.0 juM 70 ;LM 70D0pM 0.01% 0.1% 1.0% 0.01% 0.1% 1.0% 0.01% 0.1% 1.0% 0.01% 0.1% 1.0% 0.01% 0.1% 1.0% 1.02 0.88 0.44 0.92 0.85 0.68 101.0 65.0 23.0 0.95 0.57 0.18 0.98 0.96 0.6 0.88 0.82 0.56

S

EXAMPE 67: Retinal Neuron Survival This example demonstrates that PR0220 polyPeptides have efficacy in enhancing the survival of retinal neuron cells.

Sprague Dawley rat pups at postnatal day 7 (mixed population: glia and retinal neuronal types) are killed by decapitation following CO, anesthesia and the eyes are removed wider sterile conditions. The neural retina is dissected away from the pigment epithelium and other ocular tissue and then dissociated into a single cell suspension using 0.25 trypsin in Ca*, Mg'*-frec PBS. The retinas arc incubated at 37*C for 7-10 minutes after which the trypsmn is inactivated by adding 1 ml soybean trypsin inhibitor. The cells are plated at 100,000 cells per well in %6 30 well plates in DMEW/Fl2 supplemented with N2 and with or without the specific test PRO polypeptide. Cells for all experimerts are grown at 37*C in a water saturated atmosphere of 5% CO,. After 2-3 days in culture, cells are staineI with calcein AM then fixed using 4% pamformaldehyde and stained with DAPI for determnation of total cell count The total cells (fluorescent) are quantified at 20X objective magnification using CCD camera and NIH image software for Macintosh. Fields in the well amc chosen at random.

The effect of various concentration of PR0220 polypptdes are reported in Table 3 below where percent survival is calculated by dividing the total number of calcein AM positive cells at 2-3 days in culture by the total number of DAPI-labeled Cells at 2-3 days in culture. Anything above 30% survival is considered positive.

Tabil ERQ±Nam PR0220 PRO220 PR0220 PR0220 PR0220 PRO220 PRO Cnncentation 0.01% 0.01% 0.1% 0.1% 1.0% 1.0% 2.4% 4.1% 3.0%9 3.1% 72.4 42.1% EXAMPLF 68: Rod Photorecpwr Survival This example demonstrates that PRO220 polypeptides have cfficacy in enhancing the survival of rod photoreceptor cells.

Sprigue Dawley rat pups at 7 day postnaral (mixed populatiort glia and retinal neuronal CCU type) am killed by decapitation following CO, anesthesis and the eyes arc removed under sterile conditions. The neural retina is dissected away form dic pigm= epidihlum and. other ocular tissue arnd then dissociated into a single cell suspentsion using 0.25% trypsin in Mg'-frec PBS. The retinas are incubated at 37C for 7-10 minutes after which dhe trypsin is inactivated by adding I ml soybean uypin inhibitor. The cells are plated at 100.000 cells per well in 96 well plates in DMEMIFI2 supplemented with N2 a" with or without: the specific test PRO polypeptide. Cells for all exer 1i are grown at 37*C in a water saturated atmosphere of 5 C02. After 2-3 days in culture. cells are 10 fixed using 4% paraforrnaldehyde. and then stained using CellTracker Green CMFDA. Rho 4D2 (ascites or IgG 1:.100). amorIxxconal antibody dircted towards the visual pigmet rhodopsin is used to detct= rod photoreceptor cells :9 ~by indirect inmmunofluoreseeie The results are reported as survival: total number of calceinl~elrrracker rhodopsin positive cells at 2-3 days in culture, divided byit total number of rhodopsin positive cells at time 2-3 days in culture. 7be total cells (fluorescent) are quantified at 20x objective magnification using a CCD cam and NI 15 image software for Maclntosh. Fields in the well are chosen at random.

The eifct of variouas concenratin of PR022D polypeptides are reported in Table 4 below. Anything above survival is considered positive..

Table 4 PRO Nam PRO Concentration PretSrh PR0220 0.01% 0.0% PRO220l 2.0% 0.0% PRO220 10% 0.0% PRO=2 20% 66.9% PR0220 1.0% 56.9% EXAMPLE 69: Induction of Endotheil Cell Agosis The ability of PR0228 polypeptides to induce apoptosis in endothela cells was tested in hunman venous umbilical vein cridotheil cells (HUVEC. Cell Systems), using a 96-weli format. in 0 serum media supplemented with 100 ngtml VEOF. (As HUVEC cells are easily dislodged from the plating surface, all pipetting in the wells must be done as gently as practicable.) Thmemedia was aspired and the cells washetdonce withbPBS. 5 mlof Ix tysin was added to the cells in a T-175 flask, and dhe cells were allowed to stand until they were released from the plate (about 5-10 minutes).

Tsiitinwas stopped by adding 5 ml of growth media. Ile cells were spun at 1000 rpm tar 5 minutes at VC.

The media was aspirated and the cells were resuspended in 10 ml of 10% serum complemented medium (Cell Systems), 1 x pern/strep.

The cells were plated on 96-well icrotiter plates (Arnershan Life Science. cytostar-T scintillating microplate. RPNQ16O. sterile, tissue-culture treated, individually wrapped), in 10% serum (CSG-medium. Cell Systems). at adensiy of 2x 10 celsper well in atotal volume ofIOD Te PRO228 polypeptide was added in trilicateazdifutions of 0.33% andO0.11%. Wells homt cells were used as ablank and wels withells only as a negative control. As a positive contol! 1:3 serial dilutions of 50 A1 of a 3x stock of staurosporne were used.

The ability of the PR0228 polypeptide to in&=r apoptosis was deternimd using Annexin V, a member of dhe calcium and phospholipid binding proteins. to detect apoptosis.

0.2 ml Aiuaexin V Bioti stock solution (100 pg/nil) were dibrtd in 4.6 mld 2 x Ca' 4 binding buffer and BSA (1:25 dilution). 50 ph of the diluted Annexin V Biotia solution were added to each well (except controls) to a final concentration of 1.0 pgail. The samples were incubated for 10-15 minutes with Annexin-Biotin prior to direct addition of"S-Streptavidin. iSte pain was diluted in 2x Ca 2 binding buffer. 2.5% BSA and! was aimed woall wells at a finalcoreaitrananof3 x 10'cpm/wefl. The plates were then sealed, centrifuged at 1000 rpm for 1S mitattes and placed on orbital shakr for 2 hours. The analysis was performed on 1450 Microbeta Trilux (Waline). The results ame shown in Table 5 bdlow where percent above background represents dhe percentage amount of counts per minute above the negative controls. Percents greawe duan or equal to 30% above background arc considered positive.

Table PRO Nam ~PRO Concentration e ntAoeBcg d PR0228 0.11% 07 PR0228 0.11% 47.6% *:PRO228 0.33% 92.2% *PRO228 1.0% 51.4% PR0228 1.0% 95.3% EXAMPLE 70: PDB12 Cell Inhibition Tis example demoenstrates; that various PRO polypeptides have efficacy in inhibiting protein production by PDB12 pancreatic ductal cells.

PDBI2 pancreaticductalccoft are paed on fibronectin coated 96 well plates at 1.5x10' cells per well in 100 I1. of growth media. 100 pL of growth media with the PRO polypeptide test sample or negative control lacking the PRO polypcptide is then added ID well. for a final volumne of 200 pL Controls contain growth medium containing a protein shown to be iniactive in this assay. Cells arc incubated for 4 days at 37*C. 20 puL of Alamar Blue Dye (AB) is then added to each well and the flourescent reading is measured at 4 hours post addition of AB. on a microtiter plate reader at 530 am excitation and 590 nm emission. The standard employed is cells without Bovine Pituitary Extract (BPE) and with various concentrations of BPE. Buffer or CM controls from unknowns are run 2 times on each 96 well plate.

The results from these assays are shown in Table 6 below wherein percent decrease in protein production is calculated by comparing the Alamar Blue Dye calculated protein concentration produced by the PRO polypeptidetreated cells with the Alarnar Blue Dye calculated protein concentration produced by the negative control cells. A percent decrease in protein production of greater than or equal to 25 as compared to the negative control cells is considered positive.

Tal PRO211I PRO211I PRO2I11 PR0287 PRO2W PRO2W PRO29 PRO2W PROW9 PRO301 PRO301 PRO301 PRO293 PRO293 PRO293 PRO Conceflraton 0.1% 0.01% 1.0% 2.0% 10% 50% 2.0% 10% 30% 2.0% 10% 50% 2.0% 10% 50% Percent Decrease n Protein Production 0.0% 0.6% 59.7% 2i.3 18.2% 67.5% 45.53% 57.3% 52.24% 0.0% 59.8% 65.6% 0.0% 40.4% 56.7%

C

EXAMPLJE 71: Stimulation of Adult Heart Hvntronh ibis assay is desigrid to measure the ability of various PRO polypeptides to stimulate hyperirophy of adult heart.

Ventricular nyocytes; freshly isolated from adult (250g) Sprague: Dawley rats are plated at 2000 cell/wdl in 180 p1 volume. Cells arm isolated and plated on day 1, the PRO polypeptide-containing test samples or growth mufum only (negative control) (20 volume) is added on day 2 and the cells are then fixed and stained on day After staining, cell size is visualized wherein cells showing no growth enhancement as compared to control cells are given a value of 0.0. cells showing small to moderate growth enhancement as compared to control cells are given a value of 1.0 and cells showing large growth enhancemen as compared to control cells are given a value Any degree of growth enhancement as compared to the negative control cells is considered Positivc for thc assay.

The results are shown in Table 7 below.

Table 7 PRO Name PROW9 PROW8 PR0301 PROmO PRO293 PRO293 PROM0 PR0303 PRO Q ncntin 20% 20% 20% 20% 20% 20% 20% 20% 1.0h naceet cr EXA&MPLE 72: PDB12 Cell Proliferation This example denris~tes tha various PRO polypeptdecs have efficacy in inducing proliferation of PDB12 pancreatic ductal cells.

PDBI2 pancreatic dluctal cells amt plated on fibroniectin coated 96 well plates at l.5xlO' Cells per well in 100 pLJ180 jL of growth media. 100 pL of growth media with the PRO polypeptide test sample or negative control laeking the PRO polyeptide Is d=n added to well. for a final volunt of 200 j&L Controls contain growth medium containing a protein shown to be inactive in this assay. Cells are incubated for 4 days at 37*C. 20 giL of Alamar Ble Dye (AB) is then added to each well and the fiourescent reading is measured at 4 hours post addition at AB. on a microtiter plate reader at 530 amn excitation and 590 mm emission. The standard employed is cells without Bovine Pituitary Extract (EPE) and with various conoeatations of BPE. Buffer or growth medium only controls from unimownsare nin 2 nines on each 96 well plate.

The results from these assays are shown in Table 8 below wherein percent increase in protein production is calculated by comparing the Abu=a Blue Dye calculated protein concentration produced by the PRO polypeptidetreated cells with the A12-a Blue Dye calculated protein concentration produced by the negative control cells. A percent increase in protein production of greater than or equal to 25% as compared to the negative control cells is considered positive.

I!Wl PR NaQ~1me PRO Conceiiuration Percent Increase in Protein Production *PRO301 2.0% 44.0% PR0301 10% 67.4% PRO301 -50% 185.8% PR0303 2.0% 27.9% PR0303 10% 174.9% PR0303 50% 193.1% 20 EXAMPLE..71. Enhancement of Heart Neonatal H3=arrnph Induced by PRO224 This assay is designied to measure the ability of PR0224 polypeptides to stimulate hypertrophy of neonatal heart.

Cardiac myocytcs fhum 1-day old Harlan Sprague Dawley rats were obtained. Cells (190 gil at 73 x seemr freshly isolated) are added on day Ito 96-well plates previously coated with DMEMIF12 4% FCS.

Test samples containing the test PR0224 polypcptide or growth mediulm only (hegative control) (20 p1l/well) ame added ~directly to the wells on day 1. POF (20 p1/well) is then added on day 2 at final concentration of W0 M. The cells then stained on day 4 and visually scored on day 5. wherein cells showing tno increase in size as compared to negative controls are scored 0.0, cells showing a small go moderate increase in size as compared to negative controls are scored 1.0 and cells showing. argse i11CrCas in size as compared to negative controls are scored 2.0. The results are shown in Table 9 below.

Tabl PRO Name FOCnetainGrowth Pnhancement Score PR0224 0.01% 0.0 PRO224 0.1% 0.0 PR0224 1.0% EAMPLE 74: In siu Hybridization In situ hybridization is a powerful and versatile technique for the detection and localization of nucleic acid sequences within cell or tissue preparations. It may be useful, for examnple, to identify sites of gene expression.

Ezulyze the tissue ditrbalon of transcription. Identify and localiz viral infection, follow changes in specific niRNA syntesiCs ard aid in chromosome mapping.

h, mat hybridization was performed following an optimized version of the protocol by Lu and Gilett W1 Ykido 1:169-176 (1994). uising PCR-generated "P-labeled riboprobes. Briefly. forinalin-fixed, paraffin-embedded humtan tissues were sectioned, deparaffinized. deproteinated in protciaase K (20g/m!) for 15 miu'utes at 37*C. andfurther processed for in situ hybridization as described by Lu and GWei. saipra. A W-abeled antisense riboprobe was generated from a PCR product andi hybridized at 55*C overnight. The slides were dipped in Kodak NTB2 nuclear track emaulsion and exposed for 4 weeks.

32 P.Rtlboprobe synhesis pd (125 mCi) of "P-UTP (Amersbam BF 1002, SA<2000 Ci/nmnol) were speed vac dried. To each tube containing dried "P.UTF. the following ingredients were added: p1 Sx rancription buffer p1 DTT (100 mM) IdNTP mix (2.SmM 1 L;each of10 mM GTP. CTP &ATP 10 pl H,0) Al UT? (50 AM 1.0 A Rnasin p 1 DNA template (Ilpg)

ILHO

Id RNA polymerase (for PCR products 13 AS, T7 S. usually) The tubes were incubated at 37*C for one hour. 1.0 pul RQ1 DNase were added, followed by incubation at 370C for 15 minutes. 90 y1 TE (10 mM Tns pH 7.6t1mM EDTA pH 8.0) were added, and the mixture was pipened onto DEBI paper. The remaining solution was loaded in a Microcon-SO ultrafiltration unit, and spun using program 10 (6 minutles). The filtration unit was inverted over a second tube and spun using program 2 (3 minutes).

After the final recovery spin. 100 zI TE were added. 1 #1lof the final product was pipetted on DEBI paper and counted in 6 ml of Biofluor 11.

The probe was run on aTBE/uca gel. 1-3 ldofthe probe or5 idof RNA MrklIIIwere added to 3Alof loading buffer. After heating on a 95*C heat block for three minuties. the gel was irinediately placed on ice. The wells of gel were flushed, the sample loaded, and run at 180-250 volts for 45 minutes. The gel was wrapped in saran wrap and exposed to XAR film with an intensifying screen in -70'C freezer one hour to overnight.

A. Pretreaunent of frozen sections The slides were removed from the freezer, placed on aluminium trays and thawed at room temperature for minute. Tflu trays were placed in 55'C incubator for five mimm te o reduce condensation. The slides were fixed for 10 minutes in 4% paraformaldehyde on ice in the fume hood, and washed in 0.5 x SSC for 5 minutes, at room temperature (25 ml 20 x SSC 975 ml SQ After deproteination, in 0.5 p~g/mI proteinass K for 10 minutes at 37*C (12.5 #1 of 10 mg/ml stock in 250 ml prewanmed RNasc-free RNAse buffer), the sections were washed in x SSC for 10 minutes at room temperature. The sections were dehydrated in 70%. 95%, 100% ethanol, 2 minutes each.

178 r r r B. Pfetreatmnt of paraffin-mbedded sections The slides were deparaffinized, place in SQ H 2 0, and rinsed twice in 2 x SSC at roacn tenperature, for 5 minutes each time. The sections were deproteinated in 20 p/ml proteinase K (500 #l of 10 nrg/ml in 250 ml RNase-free RNabse buffer; 37 0 C, 15 minutes) human embryo, or 8 x proteinase K (100 pl in 250 ml RNase buffer 37C, 30 minutes) formalin tissues. Subsequent rinsing in 0.5 x SSC and dehydration were performed as described above.

C. Prebybridization The slides were laid out in a plastic box lined with Box buffer (4 x SSC, 50% formamide) saturated filter paper. The tissue was covered with 50 #1 of hybridization buffer (3.75g Dextran Sulfate 6 mi SQ H 2 0), vortexed and heated in the microwave for 2 minutes with the cap loosened.

After cooling on ice, 18.75 ml formamide, 3.75 ml 20 x SSC and 9 ml SQ H 2 0 were added, the tissue was vortexed well, and incubated at 42 0 C for 1-4 hours.

D.ti x 10' cpn probe and 1.0 pl tUN (50 ng/ml stock) per slide were heated at 95 0 C for 3 minutes. The slides were cooled on ice, and 48 p1 20 hybridization buffer were added per slide. After vortexing, 50pl 33 P mix were added to 50 p1 prehybridization on slide. The slides were incubated overnight at E. Washes Washing was done 2 x 10 minrrutes with 2 x SSC, EDTA at room terperature (400 ml 20 x SSC 16 ml 0.25M EDTA, followed by RNaseA treatment at 37 0 C for 30 minutes (500 p of 10 ng/ml in 250 mil RNase buffer 20 p/ml). The slides were washed 2 x 10 minutes with 2 x SSC, EDITA at roan taeperature. The stringency wash conditions were as follows: 2 hours at 55 0 C, 0.1 x SSC, EDPA (20 ml 20-x SSC 16 ml EDTA, Vf=4L).

F. Oliganeleotides In situ analysis was performed on a variety of ENA sequences disclosed herein. The oligonucleotides enployed for these analyses are as follows: EMA33094-1131 (PRD217) pl 5' 3GGAT1TrAATMAACICACI (SEQ2 ID NO:347) p2 5' CTA'AAATIANAA3lA 3 (SEe ID 10:348) IM33223-1136 (F10230) pl 5' -A -3 (SD ID ND:349) 179 p2 5#3 (SBQ ID NOR: 350) (3) p1 p 2 (4) p1 p2 MW~443S-1140 (PRO232) 5' -3(s 5' -3-(E ID NO: 351) ID NO352) rzWA5639-3172 (PRO246) 5' -3 5' -3

(SEQ

(SBQ

ID 1'O:353) ID NOK:354) MD9435-32.9 (PRO533) IG- GCIIG3 (SEYQ ID c1ttLTA3' (SE ID N:355) NO:3 56) (6)

P

1 p2 (7) p 1 p2 (8) P1 p 2 ENAI35638-1141 (PRO245) 5' -3 Eta33O89-M12 (PW221) 3' rw,35918-174 (M=058) 51 AGGnc~ cG~ 51 'cCNU~AA3' ENA32286-1191 (PRO14) -rTAr1-30 E M 3321-1133 (PR 4) 5' 3'TthATAA'1~rT 5' AA~C~A~~~cGo3XGO~CG3' IIM35557-1137 (PIO234) 5' pxxtxPxxrrcnAAc-3' 5' '1ft tXA'33' (SEQ ID NOR:357) (SEQ ID NOZ:358) (SEQ ID IN):359) (SEQ ID 1ND:360)

(SEQ

(SEQ

ID N:361) ID NO7:362) (SEQ ID ND7:363) (SEQ ID NO7:364) p 1 p2

(SEQ

(SEQ

ID NO3: 365) MD 17:366) (11) p 1 p2

(SEQ

(SEQ

ID NO7: 367) ID NO7: 368) (12) rZM3310O-M19 (IIUO229) 180 p1 5' -GGIATACCTAT 3' (SE ID ND:369) p2 5' -CAIGAAAT (SEQ ID D:370) (13) MIU34431-l1177 (P263) pl 5' -3OATITAATAACICACT AG 3' (SED ID O: 371) p2 5' -CTA3AAA TAA0WAAGOAGTAA1GAG CACGAA-3' (SED ID ND:372) (14) IUA38268-1188 (PR295) p1 -3ATIATACCAcCDACAGGAGGAGG-3' (SEQ ID ND3:373) p2 5' -CIWtAAATIAACGITACTAAAGAT CAG'GAIGAIC1CAG-3' (SE ID NO: 374) G. Results In situ analysis was perforned on a variety of EA sequences i ****disclosed herein. The results fran these analyses are as follows.

15 NA33094-1131 (PW217) ~Highly distinctive expression pattern, that does not indicate an obvious biological function. In the human aebryo it was expressed in outer smooth nuscle layer of the GI tract, respiratory cartilage, branching respiratory epithelium, osteoblasts, tendons, gonad, in the optic nerve 20 head and developing dermis. In the adult expression was observed in the epidernal pegs of the chinp tongue, the basal epithelial/mnyoepithelial cells of the prostate and urinary bladder. Also expressed in the alveolar lining cells of the adult lung, nesenchynal cells juxtaposed to erectile tissue in the penis and the cerebral cortex (probably glial cells). In the kidney, expression was only seen in disease, in cells surrounding thyroidized renal tubules.

Hman fetal tissues eaamined (E12-E16 eeks) include: Placenta, umbilical cord, liver, kidney, adrenals, thyroid, lungs, heart, great vessels, oesophagus, stcnach, snall intestine, spleen, thynus, pancreas, brain, eye, spinal cord, body wall, pelvis and lower linb.

Adult human tissues enained: Kidney (normal and end-stage), adrenal, myocardium, aorta, spleen, lyaph node, gall bladder, pancreas, lung, skin, eye (inc. retina), prostate, bladder, liver (normnal, cirrhotic, acute failure) Non-Iman primate tissues xemuirned: Chinrp Tissues: Salivary gland, staach, thyroid, parathyroid, skin, thynus, ovary, lynph node Rhesus Monkey Tissues: Cerebral cortex, hippocanpus, cerebellum, penis.

180a EN.R33223-1236 (PRO23O) Sections show an intense signal associated with arterial and venous: vessels in the fetus. In arteries the signal appeared to be confined to smoth-nuscle/pericytic cells. The signal is also seen in capillary vessels and in glcueruli. It is not clear vhether or not enkothelial cells are expressing this nR~. Express icni is also observed in epithelial cells in the fetal lens. Strong expression was also seen in cells within placental trophoblastic villi, these cells lie between the trophoblast and the fibroblast-like cells that exp~ress H3F uncertain histogenes is. In the adult, there was no evidence of expression and the wall of the aorta and imst vessels appear to be negative. However, expression was seen over vascular channels in the norml prostate and in the epitheliumt lining the gallbladder. Insurers expression was seen in the vessels of the soft-tissue sarcacla and a renal cell carcincira. In sunmaxy, this is a molecule that shows relatively specific vascular expression in the fetus as well as in scim adult organs. Exqpression was also observed in the fetal lens and the adult gallbladder.

In a secondary screen, vascular expression was observed. similar to that observed above. seen in fetal blocks. Expression is on vascular smooth muscle. srlther than enidothelium. Expression also seen in smooth msc of the developing oesophagus. so as reported previously. this molecule is not vascular specific. Expression was examined in 4 lung and 4 breast carcinomas. Substantial expression was seen in vascular smooth muscle of at least 3/4 lung cances aund 2/4 breast cances. In addition, in one breast carcinoma. expression was observed in penitumoral stirnmal cells of uncertain histogenesis (possibly myofibroblasts). No endothelial cell expression was observed in this study.

DlNA34435-1 140 (P110232) Strong expression in prostauic epitheliumn and bladder epidheliumn, lower level of expression in brounchial epitielium. Eih background low level expression seen in a nmMber of sites, including among others, bone, blood, :9 droarcoma. adult heart and fetal liver. It is felt that this level of signal represents background. partly because signal at this level was seen over die blood. All other tissue negative.

Human fetal tissues examined (E12-E16 weeks) include: Placjcnta, umbilical cord, liver, kidney. adrenals, -thyroid.

lunigs, heart, great vessels. oesophagus, stomach, small intestine.

spleen, thymus. pancreas, brain, eye. spinal cord, body wall, pelvis, testis and lower limb.

Adul hi= t~sm xamied:Kidney (normnal and enid-stage), adrenal, spleen, lymph node, pancreas, lung, eye (inc.

9: retina), bladder, liver (normal, cirrhtloic. acute failur).

Nan-human pinate tissues examined: Cimpissues: adrenal ~~Rhetis Monkey Tissues: Cerebral cortex, ofth posaeth spefcil ayrso In secondary screen, expression was observed in the epithelium o h rsae h uefca aeso the urethelium of the urinary bladder, the ureielium lining the real pelvis and the uresheliumi of the ureter (1 out of 2 experiments). The urethra of a rhesus monkey was negative; it is unclear whether this represents a true lack of exptression by the urethra, or if it is the result of a failure of die probe to cross react with rhesus tissue. The findings in the prostate and bladde art ihilar to those previously described usng an isotopic detection technique. Expression of the mRNA for this antigen is NOT prostate epithelia specific. The antigen may serve as a useful marker for uretlal derived tissues. Expression in die superficial, post-mitotic cells, of the urinary tract epithelium also suggest that it is unlikely tompen a specific stem cell marker, as this would be expected to be expressed specifically in basal epithelium.

DNA35639-172 (PR0246) Strongly expressed in fetal vascular endodieitmi, including tissues of the CNS. Lower level of expression in adult vasculature, includin the CNS. Not obviously expressed at higher levels in tmor vascular endothelium.

Signal also seen over bone matrix andi adult spleen, not obviously cell associated, probably related to non-specific background at these sites.

Human fetal tissues eained (112-F16 weeks) include: Placenta, umbilical cord, liver, kidney. adrenals, thyroid, lungs. huamt great vessels. oesophagus. stomach, small intestine, spleen, thymus. pancreas, brain, eye. spinal cord, body *wall, pelvis, testis and lower limb.

Adult h m tissus cx du: y (normal and ezil-stage). adrenal, spleen, lymph node, pancreas, lung, eyc (nc.

retina), bladder. liver (normal, cirrhotic, acute failure).

Non-hurnn primate tissues examined: Ckuimn..Iiumz~: adrenal Rhesus Monkey Tissues: Cerebral cortex, hippocampus DNA49435-1219 (PRO533t Moderate expression over cortical neutrones in the fetal brain. Expression over the inner aspect of the fatal retina, possible expression in the developing lens. Expression over fetal skin, cartilage, snall intestine, placental vilti *and umbilic.al cord. In adult tissues there is an extremnely high evel of expression over the gallbladder epithelium.

Modeate expression over the adult kidney, gastric and clonic epithelia. Low-level expression was observed ovcr many cell types in many tissues, this may be related to stickiness of thre probe, these data should therefore be interpreted with a degree of caution.

15 Human feta tissues examined (E12-EI6 weeks) Includ: Placenta, umbilical cord, liver, kidney, adrenals, thyroid, lungs heart, great vessels. oesophagus. stomach, small intestine, spleen. thyms, pancreas, brain, eye, spinal cord, body wall, pelvis, testis and lower limb.

Adul hua is~ m Kidney (normal and eml-stage). adrenal, spleen, lymnph node, pancreas, lung, eye (mnc.

retina), bladder, liver (normal, cirrhotic, acute failure).

Non-human Uriratr tissues examined: ChimRiu=: adreiwl Rhesus Monkey Tissues: Cerebral cortex. hippocampus, cerebellum.' DNA35639-1141 (PR0245) :25 Expression cbsaerdu in the endothelium lining a subset of fetal andi placental vessels. Endothelial expression *was confined to these tisue blocks. Expression also observed over intermediate trophoblast cells of placenta. All other tissues negative.

Fe ise nria 132F1 ek)icir Placenta, umbilical cord, liver, kidney, adrenals, thyroid, lungs, heart, great vessels, csophagus. stomwAh small intestine, spleen, thymus. pancreas, brain. eye. spinal cord, body wall, pelvis and lower limb.

Adult tissues examined: Liver, kidney, adrenal, mnyocardium, aorta, spleen, lymph node, pancreas, lung, s"n cerebral cortex hippocanipus(rm). cerebdlum(rm). peris. eye, bladder, stomach, gastric carcinoma, colon, colonic carcinoma, thyroid (chimp), parathyroid (chimp) ovary (chimp) and chondrosarcorna.

Actorninophen induced liver injury and hepatic cirrhosis DNA330W91132 (PR0221) Specifc expressimon over feral cerebral white and grey matter, as well as over neurones inl the spinal cord.

Probe appears 10 cross react with rat. Low level of expression over cerbellar neurones in adult rhesus brain. All other tisses negative.

Fetal tissues examined (E12-Et weekcs) include: Placenta. umbilical cord, liver. kidney. adrenals, thyroid, hmngs, heart, great vessels. oesophagus. stomach, small intestine, spleen, thymus, pancreas, brain, eye, spinal cord, body wall, pelvis and lower limb.

Adult tissues examined: Liver, kidney, adrenal, myocardium. aorta. spleen, lymph node, panceas. bang, skin. cerebral cortex hippocainpus(rm), erbelluin(rm). penis, eye, bladder, stomach, gastric carcina, colon, colonic carcinoma and diomirosarcomn. Acetominophen induced liver injury and htepatic cirrhosis 0 CO DNA'319l1 174 (PRO258) Strong expression in the. nervous system. In the rheus nkeiy brain expression is observed in cortical, hippocampal and cerebellar neurons. Expression over spinal neurones in the fetal spinal cord, the developing brain and the inner aspects of the fetal retina. Expression over developing dorsal root and autonomic ganglia as well as entcrlc nerves. Expression observed over ganglion cells in the adulc prostate. In the rat, there is strong expression over the developing hind brain and spinal cord. Strong expression over interstitial cells in the placental villi. All other tissues were negative.

Fetal tissues examid (El2-E16 weekcs) include: Placenta, umbilical cord, liver, kidney, adrenals, thyroid, lungs, beamt great vessels, oesophagus, stomach, small intestine,* spleen, thymus, pancreas, brain, eye, spinal cord, body wall, pelvis and lower limb.

Adult tissues examined: Liver, kidney, renal cell carcinoma, adrenal, aorta, spleen, lymph node, pancreas, lung, niyocardium. skin, cerebral cortex hippocampus(ffn), cerebelun(rm), bladder, prostate, stomach, gastric carcinoma, colon, colonic carcinoma, thyroid (chimp), paradhyroid (chimp) ovary (chimp) and chondrosarcoma. Acetominophen induced liver injury and hepatic cirrhosis.

DNA32296-1191 (PRfl2141 Fetal tissue: Lo~w level throughout snesenchyme. Moderate expression in placental stromal cells in membranous tissues and in thyroid. Low level expression in cortical neurones. Adult tissue: all negative.

Fetal tissues examined (E12-Et6 weeks) iniclude: Placenta, umbilical cord, fiver, kidney, adrecals, thyrod,. lungs, hertn, great vessels, aesophagus, stomach, small intestine, spleen, thynmus, pancreas, brain, eye, spinal cord, body wall, pelvis and lower limb.

Adult fissues examined inchic-~ Liver, kidney, adrenal, myocardium, aorta, spleen, lymph node, pancreas, lung and Skin.

(10) DNA33221-1133 (PR02241 Expression limited 6o vascular endothelium in fetal spleen, adult spleen, fietal liver, adult thyroid and adult lymph node (chimp). Additionl site of expression is the devetoping spinal ganglia. All other tissue negative.

thuan feral ritties exambied (012-1116 weeks) include: Placenta, umbilical cord. liver, idneiy, adrenals, thyroid.

lungs, heart, great vessels, csophagus, stomach. small intestine, spleen. thymus. pancreas, brain, eye, spinal cord, body wall, pelvis and lower limb.

Adult humnan rsue, exaned~t: Kidney (normal and end-stage), adrenal. myocardium, aorta, spleen lymph node.

pancreas. lung, skin, eye (inc. retina), bladder. live (normal, cirrhotic, acute failure).

Nor-human primate tissues examined: bd= isues: Salivary gland, stomach. thyroid, parathyroid. skin, thymus. ovary, lymph node.

Rhesus Monkey Tissues: Cerebral cortex, bippocampus, cerebellum, penis.

(11) DNA35557-1 137 (PR0234) Specific expression over developing motor eurones in ventral aspect of die feWa spinal cord (will develop into ventral horns of spinal cord). All other tissucs negative. Possible role in growth, differentiation and/or development of spinal motor neurons.

Feudl tissues examined (El 2-F1 6 weeks) include: Placenta, umbilical cord, liver, idney, adrenals,-thyroid, lungs, beart, great vessels, oesopbagus, stomach, small intestine, spleen, thymus. pancreas, brain, eye, spinal cord, body wall, pelvis and lower limb.

Adult tissues examined: Liver, idney, adrenal, utyocardiulm, aorta, spleen, lymph node, pancrcas. hing, sin, cerebral cortex OWm. bippocanius(rm), cerebellum(rm). penis, eye, bladder, stomach, gastric canmcolon, colonic carcinoma and cliondrosarcoma. Actomninophcn induced liver injury and hepatic cirrhosis DNA331OD-1 159 (PRO229) Strilcing expresion in mononuclear phagocytes (macrophages) of fetal and adult spleen liver, lymph node and adult thynas (in fingible body macrophages). The highest expression is in the spleen. All other tissues negative.

Localisation and homology are entirely consistent with a role as a scavenger receptor for cells of the 25 rtisculoeraothelial System. Expression also observed in placocatal mononuecear cells.

HiUMa fetal tigses examined CE12-El16 weeks) include: Placenta, umbilical cord, liver, kIdney. adrenals. thyroid, hings. heart, great vessels, oesophagus. stomach, small intestizie, spleen, thymus. pancreas, brain. eye, spinal cord, body wall, pelvis and lower limb.

Aduth lmnar tissues examined: Kidney (normal and end-stage), adrenal, mayocardiumn, aorta, spleen, lymph node, gall bladder, pancreas. lung, skin, eye (inc. retina), prostate, bladder, liver (normal, cirrhotic, acute failure).

Non-human riniate tissues examingd: Oft Tisues: Salivary gland, stomach, thyroid, parathyroid, skin, thyms. ovary, lymph node.

Rhesus Mon Tissues: Cerebral cortex, hippocampus, cerebellum, pei.

(13) DNA3443-117 (PR0263) Wklepread expression in human fetal tissues and placenta over mononuclear cells, probably macrophages +1nlMhocytes. The cellular distilarrion follows a perivascular pattern in many tissues. Strong expression also seen in cpithelial cells of the fectal adrenal cortex. All adult tissues were negative.

Fetal tisame examined (E12-EI6 weeks) include: Placenta. unmbilcai cord, liver. kidney, adrenals. thyroid, lungs.

beaut, great vessels, oesophagus. stomach. small intestine, spleen, thymues, pancreas, brain, eye. spinal cord. body wall, pelvis and lower limb.

Adult tissues ezamined: Liver. idney. adrenal, spleen, lymph node, pancreas, lung, skinm cerebral cortex (in), hippocampus~rm), cerebellum(rmn), bladder, stomach, colon and colonic carcinoma. Acetomninopben induced liver izijury and hepatic cirrhosis.

A secondary screen evidenced expression over stromal. mononucelear cells probably histiocytes.

(14) DNA39269-1I R8 (PR0295) 10 Higji expression over ganglion cells in human fetal spinal ganglia and over large zturones in the anteior horns of thie developing spinal cord, In the adult there is expression in the chimp adrenal undula (neural), neurones of the sbesw monkcy brain (hippocampus I and cerebral cortex) and neurones in ganglia in the n~ormnal adult human prostate (the only section that contains gantglion cells. ic expression in this cell type is presumed NOT to be confined to the prostate). All other tissues negative.

Hirmrn fetal tisnes exained JE12-E16 weeks) hit-hide: Placenta, umbilical cord, liver, kidney. adirenals. thyroid.

hasp, great vessels. stomach, small intestine, spleen, thymus. pancreas, brain, eye, spinal cord, body wall, pelvis, testis and lower limb.

Adui hinnan tissues examined: Kidney (normal andi end-stge), adrenal, spleen, lymph node, pancreas, lung, eye (inc.

retina), bladder, liver (normal, cirrhotic, acute failure).

Nont-human primte tissues examined: *~Ci T issues: adrenal Rhesus Monkey Tissues: Cerebral cortex. hippocampus, cerebellum.

Denotit of Material The tollowinig materials have been deposited with the American Type Culture Collection, 12301 Parklawn Drive, Rockville, MD, USA (ATCC): DNA32292-1131 ATCC 209258 September 16, 1997 DNA33094-1131 ATCC 209256 September 16, 1997 DNA33223-1 136 ATCC 209264 September 16. 1997 DNA34435-1 140 ATCC 209250 September 16, 19917 DNA27864-1155 ATCC 209375 October 16. 19917 DNA,36350-115g ATCC 209378 October 16. 1997 DNA32290-1164 ATCC 209384 October 16, 1997 DNA35639-1172 ATCC 209396 October 17, 1997 DNA33092-1202 ATCC 209420 October 28. 1997 DNA49435-.1219 ATCC 209480 November 21, 1997 DNA35638-1141 ATCC 209265 September 16, 19917 DNA32298-1132 ATCC 20957 September 16, 1997 DNA33089-1 132 ATCC 209262 September 16, 1997 DNA33786-1 132 ATCC 209253 September 16,.1997 DNA35918-1174 ATCC 209402 October 17. 1997 186 DNA3 7150-1178 DNA3 8260-1180 DNA3 9969-1185 DNA3 2286-1191 DNA3 3461-1199 DNA40628-1216 DNA3 3221-1133 DNA3 3107-1135 DNA3 5557-1137 DNA3 4434-1139 DNA3 3100-1159 DNA3 5600-1162 DNA3 4436-1238 DNA3 3206-1165 DNA3 5558-1167 DNA3 5599-1168 DNA36992-1168 DNA3 4407-1169 DNA35841-1173 DNA3 3470-1175 DNA3 4431-1177 DNA3 9510 1181 DNA39423-1182 DNA4 0620-1183 DNA4 0604-1187 DNA3 8268-1188 DNA37151- 1193 DNA3 5673-1201 DNA40370-1217 DNA42551- 12 17 DNA39520-1217 DNA41225-1217 DNA43318*-1217 DNA40587-1231 DNA41338-1234 ATCC2 09401 ATCC2 09397 ATCC2 09400 ATCC2093 85 ATCC2 09367 ATCC209432 ATCC2 09263 ATCC209251 ATCC209255 ATCC2 092 52 ATCC2 09377 ATCC2093 70 ATCC209523 ATCC2093 72 ATCC2 09374 ATCC209373 ATCC2 09382 ATCC209383 ATCC2 094 03 ATCC209398 ATCC2 09399 ATCC2 09392 ATCC2093 87 ATCC209388 ATCC2 093 94 ATCC2 094 21 ATCC2093 93 ATCC2 094 18 ATCC2 094 85 ATCC2O94 83.

ATCC2 094 82 ATCC2 094 91 ATCC209481 ATCC209438 ATCC209927 October 17, 1997 October 17, 1997 October 17, 1997 October 16, 1997 October 15, 1997 November 7, 1997 September 16, 1997 September 16, 1997 September 16, 1997 September 16, 1997 October 16, 1997 October 16. 1997 December 10, 1997 October 16, 1997 October 16, 1997 October 16, 1997 October 16, 1997 October 16, 1997 October 17, 1997 October 17, 1997 October 17, 1997 October 17, 1997 October 17, 1997 October 17, 1997 October 17, 1997 October 28, 1997 October 17, 1997 October 28, 1997 November 21, 1997 November 21, 1997 November 21, 1997 November 21, 1997 Novem~ber 21, 1997 November 7, 1997 June 2, 1998 186a DNA40981-1234 DNA37140-1234 DNA40982-1235 DNA41379-1236 DNA44167-1243 DNA39427-1179 DNA40603-1232 DNA43466-1225 DNA43046-1225 DNA35668-1171 ATCC209439 ATCC209489 ATCC209433 ATCC209488 ATCC209434 ATCC209395 ATCC209486 ATCC209490 ATCC209484 ATCC209371 November 7, 1997 November 21, 1997 November 7, 1997 November 21, 1997 November 7, 1997 October 17, 1997 November 21, 1997 November 21, 1997 November 21, 1997 October 16, 1997 These deposits were made under the provisions of the Budapest Treaty on the International Recognition of the Deposit of Microorganisms for the Purpose of Patent Procedure and the Regulations thereunder (Budapest Treaty).

This assures maintenance of a viable culture of the deposit for 30 years from the date of deposit. The deposits will be made available by ATCC under the terms of the Budapest Treaty, and subject to an agreement between Genentech, Inc.

and ATCC, which assures permanent and unrestricted availability of the progeny of the culture of the deposit to the public upon issuance of the pertinent U.S. patent or upon laying open to the public of any U.S. or foreign patent application, whichever comes first, and assures availability of the progeny to one determined by the U.S. Conumissioner of Patents and Trademarks to be entitled thereto according to 35 usc 122 and te Commnissioner's rules pursuant thereto (including 37 CFR 1.14 with particular reference to 85600G638).

The assignee of the present application has agreed that if a culture of the mnaterials on deposit should die or be lost or desroyed when cultivated under sulohle conditions. the materials will be promptly replaced on notification with aziothc of the same. Xvailability of the deposited mnateuial is not to, be construed as a Ii c.ense to practice the invention in contravention of the rights granted under the authority of any goverment in accordance with its patent taws.

The foregoing written specification is considered wo be sufficient to enable one skilled in the anrt o practice the invention. The present invention is not to be limnited in scope by the constrc deposited, since the deposited embodiment is intended as a single illustration of certain aspects of the invention and any construs that arc functionally equivalent are within the scope of this invention. The deposit of material herein does not constitute an admnission that the written description herein contained is inadequate to enable tie practice of any aspect of dhe imunion nluding the besn mode theref, nor is it to be construed as limniting the scope of the claims to die specific illustrations that it represcrts. Indeed. various rnodifications of the invention in addition to those shown nd described herein will becomie apparent to dse silled in the art from the foregoing description and fall within the scope of the appended claims.

Claims (19)

1. An isolated nucleic acid molecule having at least sequence identity to a nucleotide sequence which encodes a polypeptide comprising an amino acid sequence which has the amino acid sequence shown in Figure 46 (SEQ ID NO:127).

2. A nucleic acid molecule according to Claim 1, wherein said nucleotide sequence comprises a nucleotide sequence as shown in Figure 45 (SEQ ID NO:126), or the complement thereof.

3. A nucleic acid molecule according to Claim 1, wherein said nucleotide sequence comprises the full-length coding sequence of the sequence shown in Figure 45 (SEQ ID NO:126), or 15 the complement thereof.

4. An isolated nucleic acid molecule which comprises the full-length coding sequence of the DNA deposited under accession number ATCC 209263. *o A vector comprising a nucleic acid according to any one of Claims 1 to 4.

6. A vector according to Claim 5, operably linked to 25 control sequences recognized by a host cell transformed with the vector.

7. A host cell comprising a vector according to Claim or Claim 6.

8. A host cell according to Claim 7, wherein said cell is a CHO cell.

9. A host cell according to Claim 7, wherein said cell is an E. coli. A host cell according to Claim 7, wherein said cell is a yeast cell. 189

11. A process for producing a PRO polypeptide, comprising the step of culturing a host cell according to any one of Claims 7 to 10 under conditions suitable for expression of said PRO polypeptide, and recovering said PRO polypeptide from the cell culture.

12. Isolated native sequence PRO polypeptide having at least 80% sequence identity to the amino acid sequence shown in Figure 46 (SEQ ID NO:127).

13. Isolated PRO polypeptide having at least 80% sequence identity to the amino acid sequence encoded by the nucleotide deposited under accession number ATCC 209263.

14. A chimeric molecule comprising a polypeptide according to Claim 12 or Claim 13, fused to a heterologous amino acid sequence.

15. A chimeric molecule according to Claim 14, wherein 20 said heterologous amino acid sequence is an epitope tag sequence.

16. A chimeric molecule according to Claim 14, wherein said heterologous amino acid sequence is a Fc region of an 25 immunoglobulin.

17. An antibody which specifically binds to a PRO polypeptide according to Claim 12 or Claim 13.

18. An antibody according to Claim 17, wherein said antibody is a monoclonal antibody.

19. A composition comprising a) a nucleic acid molecule according to any one of Claims 1 to 4; b) a polypeptide according to Claim 12 or Claim 13; c) a chimeric molecule according to any one of Claims 14 to 16; or 190 d) an antibody according to Claim 17 or Claim 18, together with a pharmaceutically-acceptable carrier.

20. A nucleic acid according to Claim 1 substantially as hereinbefore described with reference to any one of the examples.

21. A polypeptide according to Claim 12 or Claim 13, substantially as hereinbefore described with reference to any one of the examples. Dated this 1 s t day of February 2002 GENENTECH, INC. By their Patent Attorneys GRIFFITH HACK Fellows Institute of Patent and Trade Mark Attorneys of Australia *o