CA2367378A1 - Molecules of the immune system - Google Patents

Abstract

The invention provides human immune system molecules (IMOL) and polynucleotides which identify and encode IMOL. The invention also provides expression vectors, host cells, antibodies, agonists, and antagonists. The invention also provides methods for diagnosing, treating, or preventing disorders associated with expression of IMOL.

Description

MOLECULES OF THE IMMUNE SYSTEM
TECHNICAL FIELD
This invention relates to nucleic acid and amino acid sequences of immune system molecules and to the use of these sequences in the diagnosis, treatment, and prevention of immunological disorders, infections, and cell proliferative disorders including cancer.
BACKGROUND OF THE INVENTION
All vertebrates have developed sophisticated and complex immune systems that provide protection from viral, bacterial, fungal and parasitic infections and cancers.
Included in these systems are the processes of humoral immunity, the complement cascade, and the inflammatory response (See Paul, W.E. (1993) Fundamental ImmunoloQV, Raven Press, Ltd., New York NY pp.l-20).
The immune system responds to invading microorganisms in two major ways:
antibody production and cell mediated responses. The cells that recoginze and destroy pathogens include different types of leukocytes: monocytes, lymphocytes, neutrophils, eosinophils, and basophils.
Neutrophils and monocytes attack invading bacteria, viruses, and other pathogens and destroy them by phagocytosis. Monocytes enter tissues and differentiate into macrophages which are extremely phagocytic. Lymphocytes and plasma cells are a part of the immune system which recognizes specific foreign molecules and organisms and inactivates them, as well as signaling other cells to attack the invaders. Leukocytes are formed from two stem cell Iineages in bone marrow. The myeloid stem cell line produces granulocytes and monocytes and, the lymphoid stem cell produces lymphocytes. Lymphoid cells travel to the thymus, spleen and lymph nodes, where they mature and differentiate into lymphocytes. Two classes of lymphocytes are T- and B-lymphocytes, also called T
cells and B cells. The maturation of lymphocytes is subject to control by a variety of factors including the interleukins.
Cell-mediated immune responses involve T cells that react with foreign antigen on the surface of infected or transformed (cancerous) cells. There are two major types of T cells: cytotoxic T cells destroy antigen-bearing cells, whereas helper T cells activate other white blood cells via chemical signals such as the interleukins. One class of helper cell, TH1, activates macrophages to destroy ingested microorganisms, while another, TH2, stimulates the production of antibodies by B
cells.
Antibodies are immunoglobuIin proteins produced by B-lymphocytes which bind to specific antigens and cause inactivation or promote destruction of the antigen by other cells. The prototypical antibody is a tetramer consisting of two identical heavy polypeptide chains (H-chains) and two identical light polypeptide chains (L-chains) interlinked by disulfide bonds.
This arrangement confers the characteristic Y-shape to antibody molecules. Antibodies are classified based on their H-chain composition. The five antibody classes, IgA, IgD, IgE, IgG and IgM, are defined by the a, b, e, y, and p H-chain types. There are two types of L-chains, x and ~., either of which may associate as a pair with any H-chain pair. IgG, the most common class of antibody found in the circulation, is tetrameric, while the other classes of antibodies are generally variants or multimers of this basic structure.
H-chains and L-chains each contain an N-terminal variable region and a C-terminal constant region. The constant region consists of about 110 amino acids in L-chains and about 330 or 440 amino acids in H-chains. The amino acid sequence of the constant region is nearly identical among H- or L-chains of a particular class. The variable region consists of about 110 amino acids in both H-and L-chains. However; the amino acid sequence of the variable region differs among H- or L-chains of a particular class. Within each H- or L-chain variable region are three hypervariable regions of extensive sequence diversity, each consisting of about 5 to 10 amino acids. In the antibody molecule, the H- and L-chain hypervariable regions come together to form the antigen recognition site.
(Reviewed in Alberts, B. et al. (1994) Molecular Biology of the Cell, Garland Publishing, New York, NY, pp.1206-1213 and 1216-1217.) Both H-chains and L-chains contain repeated immunoglobulin domains. For example, a typical H-chain contains four Ig domains, three of which occur within the constant region and one of which occurs within the variable region and contributes to the formation of the antigen recognition site. Likewise, a typical L-chain contains two Ig domains, one of which occurs within the constant region and one of which occurs within the variable region. In addition, H
chains such as p have been shown to associate with other polypeptides during differentiation of the B-cell.
Host defense is further augmented by the complement system. The complement system serves as an effector system and is involved in infectious agent recognition.
It can function as an independent immune network or in conjunction with other humoral immune responses. The complement system is comprised of numerous plasma and membrane proteins that act in a cascade of reaction sequences whereby one component activates the next to generate a rapid and amplified response to infection through either an inflammatory response or increased phagocytosis. The first element in this cascade, C 1, is composed of subunits C 1 q, C 1 r, and C 1 s.
C 1 q-induced cellular responses are thought to be involved in host defense and in protection against autoimmunity since Clq-deficient humans are suceptible to infectious disorders and a very high incidence of autoimmune disease (Nicholson-Welter, A. and Klickstein, L.B. (1999) Curr. Opin. Immunol.
11:42-46).
The classical pathway requires antibody binding to infectious agent antigens.
The antibodies serve to define the target and initiate the complement system cascade, culminating in the destruction of the infectious agent. In this pathway, since the antibody guides initiation of the process, the complement can be seen as an effector arm of the humoral immune system.
The alternative pathway of the complement system does not require the presence of pre-existing antibodies for targeting infectious agent destruction. Rather, this pathway, through low levels of an activated component, remains constantly primed and provides surveillance in the non-immune host to enable targeting and destruction of infectious agents. In this case foreign material triggers the cascade, thereby facilitating phagocytosis or lysis (Paul, W.
(1993) Fundamental Immunolo~y Raven Press Ltd., New York NY pp.918-919).
Antigen Reco ign lion A key feature of the immune system is its ability to distinguish foreign molecules, or antigens, from "self' molecules. This ability is mediated primarily by secreted and transmembrane proteins expressed by leukocytes (white blood cells).such as lymphocytes, granulocytes, and moaocytes. Most of these proteins belong to the immunoglobulin (Ig) superfamily, members of which contain one or more repeats of a conserved structural domain. This Ig domain is comprised of IS antiparallel ~i sheets joined by a disulfide bond in an arrangement called the Ig fold. Members of the Ig superfamily include T-cell receptors, major histocompatibility (MHC) proteins, antibodies, and immune cell-specific surface markers such as CD4, CDB, and CD28.
MHC proteins are cell surface markers that bind to and present foreign antigens to T cells.
MHC molecules are classified as either class I or class II. Class I MHC
molecules (MHC I) are expressed on the surface of almost all cells and are involved in the presentation of antigen to cytotoxic T cells. For example, a cell infected with virus will degrade intracellular viral proteins and express the protein fragments bound to MHC I molecules on the cell surface.
The MHC I/antigen complex is recognized by cytotoxic T-cells which destroy the infected cell and the virus within.
Class II MHC molecules are expressed primarily on specialized antigen-presenting cells of the immune system, such as B-cells and macrophages. These cells ingest foreign proteins from the extracellular fluid and express MHC II/antigen complex on the cell surface.
This complex activates helper T-cells, which then secrete cytokines and other factors that stimulate the immune response.
MHC molecules also play an important role in organ rejection following transplantation. Rejection occurs when the recipient's T-cells respond to foreign MHC molecules on the transplanted organ in the same way as to self MHC molecules bound to foreign antigen. (Reviewed in Alberts, B. et al.
(1994) Molecular Biology of the Cell, Garland Publishing, New York, NY, pp.
1229-1246.) Antibodies, or immunoglobulins, bind and neutralize antigens in the circulation and other extracellular fluids. Antibody classes include IgG, IgA, IgM, IgD, and IgE.
IgA are found primarily in secretions and play an important role in mucosal immunity. IgA are transcytosed across epithelial cell sheets and secreted along with mucous into the lumenal space.
Transcytosis and secretion are mediated by the polymeric Ig receptor, which binds to and transports IgA. The polymeric Ig receptor is a transmembrane protein with about two to five Ig domains in its extracellular domain. (Reviewed in Alberts, supra, pp. 1210-1211; Kulseth, M. A. et al. (1995) DNA Cell Biol.
(1995) 14:251-256.) T-cell receptors are both structurally and functionally related to antibodies.
T-cell receptors are cell surface proteins that bind foreign antigens and mediate diverse aspects of the immune response. A typical T-cell receptor is a heterodimer comprised of two disulfide-linked polypeptide chains called a and (3. Each chain is about 280 amino acids in length and contains one variable region and one constant region. Each variable or constant region folds into an Ig domain. The variable regions from the a and ~i chains come together in the heterodimer to form the antigen recognition site.
T-cell receptor diversity is generated by somatic rearrangement of gene segments encoding the a and ~i chains. T-cell receptors recognize small peptide antigens that are expressed on the surface of antigen-presenting cells and pathogen-infected cells. These peptide antigens are presented on the cell surface in associatiomwith major histocompatibility proteins which provide the proper context for antigen recognition. (Reviewed in Alberts, supra, pp. 1228-1229.) Immune Cell Signnaling Cytokines comprise a family of signaling molecules that modulate the immune system and the inflammatory response. Cytokines are usually secreted by leukocytes, or white blood cells, in response to injury or infection. However, other tissues are capable of secreting cytokines in response to disease or trauma. Cytokines function as growth and differentiation factors that act primarily on cells of the immune system including B- and T-lymphocytes, monocytes, macrophages, granulocytes, and their progenitors, such as the myeloid stem cells and lymphoid stem cells.
Like other signaling molecules, cytokines bind to specific plasma membrane receptors and trigger intracellular signal transduction pathways which alter gene expression patterns. There is considerable potential for the use of cytokines in the treatment of inflammation and immune system disorders.
Cytokine structure and function have been extensively characterized in vitro.
Most cytokines are small polypeptides of about 30 kilodaltons or less. Over 50 cytokines have been identified from human and rodent sources. Examples of cytokine subfamilies include the interferons (IFN-a, -(3, and -y), the interleukins (IL-1 through IL-18), the tumor necrosis factors (TNF-a and -(3), and the chemokines. Many cytokines have been produced using recombinant DNA
techniques, and the activities of individual cytokines have been determined in vitro. These activities include regulation of leukocyte proliferation, differentiation, and motility. (Reviewed in Callard, R. E. and Gearing, A. J.
H. (1994) The Cvtokine Facts Book, Academic Press, San Diego, CA, pp. 2-6, 12-17; see, for example, Fossiez, F. et al. (1998) Int. Rev. Immunol. 16:541-551.) The activity of an individual cytokine in vitro may not reflect the full scope of that cytokine's activity in vivo. Cytokines are not expressed individually in vivo but are instead expressed in combination with a multitude of other cytokines when the organism is challenged with a stimulus.
Together, these cytokines collectively modulate the immune response in a manner appropriate for that particular stimulus. Therefore, the physiological activity of a cytokine is determined by the stimulus itself and by complex interactive networks among co-expressed cytokines which may demonstrate both synergistic and antagonistic relationships.
Chemokines comprise a cytokine subfamily with over 30 members. (Reviewed in Wells, T.
N. C. and Peitsch, M. C. (1997) J. Leukoc. Biol. 61:545-550.) Chemokines were initially identified as chemotactic proteins that recruit monocytes and macrophages to sites of inflammation. Recent evidence indicates that chemokines may also play key roles in hematopoiesis and HIV-1 infection.
Chemokines are small proteins which range from about 6-15 kilodaltons in molecular weight.
Chemokines are further classified as C, CC, CXC, or CX3C based on the number and position of critical cysteine residues. The CXC chemokines, for example, each contain a conserved motif consisting of two cysteines separated by a single residue followed by two additional cysteines which occur downstream at about 23- and 12-residue intervals (ExPASy PROSITE
database, documents PS00471 and PDOC00434). The presence and spacing of these four cysteine residues are highly conserved, whereas the intervening residues diverge significantly.
Organs of the Immune Svstem The major organs of the immune system are classified as either primary or secondary lymphoid organs. Primary lymphoid organs include the bone marrow, which produces B-cells, and the thymus, which produces T-cells (thymocytes). Bone marrow also contains blood vessels, nerves, fatty tissue, and stromal cells. Stromal cells produce a supporting meshwork of collagen fibers and other extracellular matrix components which are important for promoting the growth and differentiation of B-cells and other hematopoietic cells. Upon maturation, B-and T-cells travel through the lymphatic system and populate secondary lymphoid organs throughout the body such as the lymph nodes, adenoids, tonsils, spleen, and intestinal Peyer's patches.
Disorders of the Immune S sy tem Disorders of the immune system include various autoimmune and inflammatory diseases caused by failure of the immune system to discriminate self from non-self molecules. Other immune system disorders are caused by uncontrolled cell proliferation, including leukemias such as multiple myeloma and lymphomas such as Hodgkin's disease. Immunodeficiency, brought on by a variety of diseases and agents including HIV, renders afflicted individuals susceptible to severe and sometimes fatal bacterial and viral infections. (See, for example, Golub, E. S. et al.
(1987) ImmunoloQV: A

Synthesis, Sinauer Associates, Sunderland, MA,-pages 481 and 509-530.) Diseases which cause over- or under-abundance of any one type of leukocyte usually result in the entire immune defense system becoming involved. The most well known autoimmune disease is AIDS (Acquired Immunodeficiency Syndrome). This disease depletes the number of helper T cells and leaves the patient susceptible to infection by microorganisms and parasites.
Immunocompromised patients are also at increased risk for cancers.
Leukopenia or agranulocytosis occurs when the bone marrow stops producing white blood cells. This leaves the body unprotected against foreign microorganisms, including those which normally inhabit the skin, mucous membranes, and gastrointestinal tract.
Impaired phagocytosis occurs in several diseases, including monocytic leukemia, systemic lupus, and granulomatous disease.
Leukemias are an excess production of white blood cells, to the point where a major portion of the body's metabolic resources are directed solely at proliferation of white blood cells, leaving other tissues to starve. In myelogenous leukemias, cancerous young myelogenous cells spread from the bone marrow to other organs, especially the spleen, liver, lymph nodes and other highly vascularized regions. Usually, the extra leukemic cells released are immature, incapable of function, and undifferentiated. Leukemias may be caused by exposure to environmental factors such as radiation or toxic chemicals or by genetic aberration.
Transplant rejection and allergies are examples of situations in which it may be desirable to curtail the immune response. Interleukin activity may play a pivotal role in specific immune responses. An antagonist to IL-17 has been shown to promote survival in heart tissue grafts (Antonysamy, M.A. et al. ( 1999) Transplant. Proc. 31:93).
The discovery of new immune system molecules and the polynucleotides encoding them satisfies a need in the art by providing new compositions which are useful in the diagnosis, prevention, and treatment of immunological disorders, infections, and cell proliferative disorders including cancer.
SUMMARY OF THE INVENTION
The invention features purified polypeptides, immune system molecules, referred to collectively as "IMOL" and individually as "IMOL-I," "IMOL-2," "1MOL-3," "IMOL-4," "IMOL-5," "IMOL-6" "I1VIOL-7," "IMOL-8," "I1VIOL-9," "IMOL-10," "IMOL-1 I," "I1VIOL-12," "IMOL-13,"
"IMOL-14," and "1MOL-15." In one aspect, the invention provides an isolated polypeptide comprising a) an amino acid sequence selected from the group consisting of SEQ
)D NO:1-15, b) a naturally occurring amino acid sequence having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO:1-15, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ )D NO:1-15, or d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ 1D
NO:1-15. In one alternative, the invention provides an isolated polypeptide comprising the amino acid sequence of SEQ m NO:1-15.
The invention further provides an isolated polynucleotide encoding a polypeptide comprising a) an amino acid sequence selected from the group consisting of SEQ m NO:1-15, b) a naturally occurring amino acid sequence having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ )D NO:1-15, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ )D NO:1-15, or d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ
1D NO:1-15. In one alternative, the polynucleotide is selected from the group consisting of SEQ m N0:16-30.
Additionally, the invention provides a recombinant polynucleotide comprising a promoter sequence operably linked to a polynucleotide encoding a polypeptide comprising a) an amino acid sequence selected from the group consisting of SEQ 1D NO:1-15, b) a naturally occurring.amino acid sequence having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ )D NO:1-15, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ m NO:1-15, or d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-15. In one alternative, the invention provides a cell transformed with the recombinant polynucleotide. In another alternative, the invention provides a transgenic organism comprising the recombinant polynucleotide.
The invention also provides a method for producing a polypeptide comprising a) an amino acid sequence selected from the group consisting of SEQ )D NO:1-15, b) a naturally occurnng amino acid sequence having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ 1D NO:1-15, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ m NO:1-15, or d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ m NO:1-15. The method comprises a) culturing a cell under conditions suitable for expression of the polypeptide, wherein said cell is transformed with a recombinant polynucleotide comprising a promoter sequence operably linked to a polynucleotide encoding the polypeptide, and b) recovering the polypeptide so expressed.
Additionally, the invention provides an isolated antibody which specifically binds to a polypeptide comprising a) an amino acid sequence selected from the group consisting of SEQ )D
NO:1-15, b) a naturally occurnng amino acid sequence having at least 90%
sequence identity to an amino acid sequence selected from the group consisting of SEQ m NO:1-15, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ
)D NO:1-15, or d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ m NO:1-15.

The invention further provides an isolated polynucleotide comprising a) a polynucleotide sequence selected from the group consisting of SEQ m N0:16-30, b) a naturally occurring polynucleotide sequence having at least 90% sequence identity to a polynucleotide sequence selected from the group consisting of SEQ )D N0:16-30, c) a polynucleotide sequence complementary to a), or d) a polynucleotide sequence complementary to b). In one alternative, the polynucleotide comprises at least 60 contiguous nucleotides.
Additionally, the invention provides a method for detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide comprising a) a polynucleotide sequence selected from the group consisting of SEQ m N0:16-30, b) a naturally occurring polynucleotide sequence having at least 90% sequence identity to a polynucleotide sequence selected from the group consisting of SEQ )D N0:16-30, c) a polynucleotide sequence complementary to a), or d) a polynucleotide sequence complementary to b). The method comprises a) hybridizing the sample with a probe comprising at least l6 contiguous nucleotides comprising a sequence complementary to said target polynucleotide in the sample, and which probe specifically hybridizes to said target polynucleotide, under conditions whereby a hybridization complex is formed between said probe and said target polynucleotide, and b) detecting the presence or absence of said hybridization complex, and optionally, if present, the amount thereof. In one alternative, the probe comprises at least 30 contiguous nucleotides. In another alternative, the probe comprises at least 60 contiguous nucleotides.
The invention further provides a pharmaceutical composition comprising an effective amount of a polypeptide comprising a) an amino acid sequence selected from the group consisting of SEQ )D
NO:1-15, b) a naturally occurring amino acid sequence having at least 90%
sequence identity to an amino acid sequence selected from the group consisting of SEQ B7 NO:1-15, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ m NO:1-15, or d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ m NO:1-15, and a pharmaceutically acceptable excipient. The invention additionally provides a method of treating a disease or condition associated with decreased expression of functional IMOL, comprising administering to a patient in need of such treatment the pharmaceutical composition.
The invention also provides a method for screening a compound for effectiveness as an agonist of a polypeptide comprising a) an amino acid sequence selected from the group consisting of SEQ >D NO:1-15, b) a naturally occurring amino acid sequence having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ m NO:1-15, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ m NO:1-15, or d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ
>D NO:1-15. The method comprises a) exposing a sample comprising the polypeptide to a compound, and b) detecting agonist activity in the sample. In one alternative, the invention provides a pharmaceutical composition comprising an agonist compound identified by the method and a pharmaceutically acceptable excipient. In another alternative, the invention provides a method of treating a disease or condition associated with decreased expression of functional IMOL, comprising administering to a patient in need of such treatment the pharmaceutical composition.
Additionally, the invention provides a method for screening a compound for effectiveness as an antagonist of a polypeptide comprising a) an amino acid sequence selected from the group consisting of SEQ )D NO:1-15, b) a naturally occurring amino acid sequence having at least 90%
sequence identity to an amino acid sequence selected from the group consisting of SEQ 117 NO:1-15, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-15, or d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-15. The method comprises a) exposing a sample comprising the polypeptide to a compound, and b) detecting antagonist activity in the sample.
In one alternative, the invention provides a pharmaceutical composition comprising an antagonist compound identified by the method and a pharmaceutically acceptable excipient. In another alternative, the invention provides a method of treating a disease or condition associated with overexpression of functional IMOL, comprising administering to a patient in need of such treatment the pharmaceutical composition.
The invention further provides a method for screening a compound for effectiveness in altering expression of a target polynucleotide, wherein said target polynucleotide comprises a sequence selected from the group consisting of SEQ ID N0:16-30, the method comprising a) exposing a sample comprising the target polynucleotide to a compound, and b) detecting altered expression of the target polynucleotide.
BRIEF DESCRIPTION OF THE TABLES
Table 1 shows polypeptide and nucleotide sequence identification numbers (SEQ
ID NOs), clone identification numbers (clone IDs), cDNA libraries, and cDNA fragments used to assemble full-length sequences encoding IMOL.
Table 2 shows features of each polypeptide sequence, including potential motifs, homologous sequences, and methods, algorithms, and searchable databases used for analysis of IMOL.
Table 3 shows selected fragments of each nucleic acid sequence; the tissue-specific expression patterns of each nucleic acid sequence as determined by northern analysis; diseases, disorders, or conditions associated with these tissues; and the vector into which each cDNA was cloned.
Table 4 describes the tissues used to construct the cDNA libraries from which cDNA clones encoding IMOL were isolated.
Table 5 shows the tools, programs, and algorithms used to analyze IMOL, along with applicable descriptions, references, and threshold parameters.
DESCRIPTION OF THE INVENTION
Before the present proteins, nucleotide sequences, and methods are described, it is understood that this invention is not limited to the particular machines, materials and methods described, as these may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present invention which will be limited only by the appended claims.
It must be noted that as used herein and in the appended claims, the singular forms "a," "an,"
and "the" include plural reference unless the context clearly dictates otherwise. Thus, for example, a reference to "a host cell" includes a plurality of such host cells, and a reference to "an antibody" is a reference to one or more antibodies and equivalents thereof known to those skilled in the art, and so forth.
Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of ordinary skill in the art to which this invention belongs.
Although any machines, materials, and methods similar or equivalent to those described herein can be used to practice or test the present invention, the preferred machines, materials and methods are now described. All publications mentioned herein are cited for the purpose of describing and disclosing the cell lines, protocols, reagents and vectors which are reported in the publications and which might be used in connection with the invention. Nothing herein is to be construed as an admission that the invention is not entitled to antedate such disclosure by virtue of prior invention.
DEFINITIONS
"IMOL" refers'to the amino acid sequences of substantially purified IMOL
obtained from any species, particularly a mammalian species, including bovine, ovine, porcine, murine, equine, and human, and from any source, whether natural, synthetic, semi-synthetic, or recombinant.
The term "agonist" refers to a molecule which intensifies or mimics the biological activity of IMOL. Agonists may include proteins, nucleic acids, carbohydrates, small molecules, or any other compound or composition which modulates the activity of IMOL either by directly interacting with IMOL or by acting on components of the biological pathway in which IMOL
participates.
An "allelic variant" is an alternative form of the gene encoding IMOL. Allelic variants may result from at least one mutation in the nucleic acid sequence and may result in altered mRNAs or in polypeptides whose structure or function may or may not be altered. A gene may have none, one, or many allelic variants of its naturally occurring form. Common mutational changes which give rise to allelic variants are generally ascribed to natural deletions, additions, or substitutions of nucleotides.
Each of these types of changes may occur alone, or in combination with the others, one or more times in a given sequence.
"Altered" nucleic acid sequences encoding IMOL include those sequences with deletions, insertions, or substitutions of different nucleotides, resulting in a polypeptide the same as IMOL or a polypeptide with at least one functional characteristic of IMOL. Included within this definition are polymorphisms which may or may not be readily detectable using a particular oligonucleotide probe of the polynucleotide encoding IMOL, and improper or unexpected hybridization to allelic variants, with a locus other than the normal chromosomal locus for the polynucleotide sequence encoding IMOL. The encoded protein may also be "altered," and may contain deletions, insertions, or substitutions of amino acid residues which produce a silent change and result in a functionally equivalent IMOL. Deliberate amino acid substitutions may be made on the basis of similarity in polarity, charge, solubility, hydrophobicity, hydrophilicity, and/or the amphipathic nature of the residues, as long as the biological or immunological activity of IMOL is retained. For example, negatively charged amino acids may include aspartic acid and glutamic acid, and positively charged amino acids may include lysine and arginine. Amino acids with uncharged polar side chains having similar hydrophilicity values may include: asparagine and glutamine; and serine and threonine.
Amino acids with uncharged side chains having similar hydrophilicity values may include: leucine, isoleucine, and valine; glycine and alanine; and phenylalanine and tyrosine.
The terms "amino acid" and "amino acid sequence" refer to an oligopeptide, peptide, polypeptide, or protein sequence, or a fragment of any of these, and to naturally occurring or synthetic molecules. Where "amino acid sequence" is recited to refer to an amino acid sequence of a naturally occurring protein molecule, "amino acid sequence" and like terms are not meant to limit the amino acid sequence to the complete native amino acid sequence associated with the recited protein molecule.
"Amplification" relates to the production of additional copies of a nucleic acid sequence.
Amplification is generally carried out using polymerase chain reaction (PCR) technologies well known in the art.
The term "antagonist" refers to a molecule which inhibits or attenuates the biological activity of IMOL. Antagonists may include proteins such as antibodies, nucleic acids, carbohydrates, small molecules, or any other compound or composition which modulates the activity of IIvIOL either by directly interacting with IMOL or by acting on components of the biological pathway in which IMOL
participates.
The term "antibody" refers to intact immunoglobulin molecules as well as to fragments thereof, such as Fab, Flab' )z, and Fv fragments, which are capable of binding an epitopic determinant.

Antibodies that bind IMOL polypeptides can be prepared using intact polypeptides or using fragments containing small peptides of interest as the immunizing antigen. The polypeptide or oligopeptide used to immunize an animal (e.g., a mouse, a rat, or a rabbit) can be derived from the translation of RNA, or synthesized chemically, and can be conjugated to a carrier protein if desired. Commonly used carriers that are chemically coupled to peptides include bovine serum albumin, thyroglobulin, and keyhole limpet hemocyanin (KLH). The coupled peptide is then used to immunize the animal.
The term "antigenic determinant" refers to that region of a molecule (i.e., an epitope) that makes contact with a particular antibody. When a protein or a fragment of a protein is used to immunize a host animal, numerous regions of the protein may induce the production of antibodies which bind specifically to antigenic determinants (particular regions or three-dimensional structures on the protein). An antigenic determinant may compete with the intact antigen (i.e., the immunogen used to elicit the immune response) for binding to an antibody.
The term "antisense" refers to any composition capable of base-pairing with the "sense"
strand of a specific nucleic acid sequence. Antisense compositions may include DNA; RNA; peptide nucleic acid (PNA); oligonucleotides having modified backbone linkages such as phosphorothioates, methylphosphonates, or benzylphosphonates; oligonucleotides having modified sugar groups such as 2'-methoxyethyl sugars or 2'-methoxyethoxy sugars; or oligonucleotides having modified bases such as 5-methyl cytosine, 2'-deoxyuracil, or 7-deaza-2'-deoxyguanosine. Antisense molecules may be produced by any method including chemical synthesis or transcription. Once introduced into a cell, the complementary antisense molecule base-pairs with a naturally occurring nucleic acid sequence produced by the cell to form duplexes which block either transcription or translation. The designation "negative" or "minus" can refer to the antisense strand, and the designation "positive" or "plus" can refer to the sense strand of a reference DNA molecule.
The term "biologically active" refers to a protein having structural, regulatory, or biochemical functions of a naturally occurring molecule. Likewise, "immunologically active" refers to the capability of the natural, recombinant, or synthetic IMOL, or of any oligopeptide thereof, to induce a specific immune response in appropriate animals or cells and to bind with specific antibodies.
The terms "complementary" and "complementarity" refer to the natural binding of polynucleotides by base pairing. For example, the sequence "5' A-G-T 3"' bonds to the complementary sequence "3' T-C-A 5'." Complementarity between two single-stranded molecules may be "partial," such that only some of the nucleic acids bind, or it may be "complete," such that total complementarity exists between the single stranded molecules. The degree of complementarity between nucleic acid strands has significant effects on the efficiency and strength of the hybridization between the nucleic acid strands. This is of particular importance in amplification reactions, which depend upon binding between nucleic acid strands, and in the design and use of peptide nucleic acid (PNA) molecules.
A "composition comprising a given polynucleotide sequence" and a "composition comprising a given amino acid sequence" refer broadly to any composition containing the given polynucleotide or amino acid sequence. The composition may comprise a dry formulation or an aqueous solution.
Compositions comprising polynucleotide sequences encoding IMOL or fragments of IMOL may be employed as hybridization probes. The probes may be stored in freeze-dried form and may be associated with a stabilizing agent such as a carbohydrate. In hybridizations, the probe may be deployed in an aqueous solution containing salts (e.g., NaCI), detergents (e.g., sodium dodecyl sulfate; SDS), and other components (e.g., Denhardt's solution, dry milk, salmon sperm DNA, etc.).
"Consensus sequence" refers to a nucleic acid sequence which has been resequenced to resolve uncalled bases, extended using the XL-PCR kit (Perkin-Elmer, Norwalk CT) in the 5' and/or the 3' direction, and resequenced, or which has been assembled from the overlapping sequences of one or more Incyte Clones and, in some cases, one or more public domain ESTs, using a computer program for fragment assembly, such as the GELVIEW fragment assembly system (GCG, Madison Wn. Some sequences have been both extended and assembled to produce the consensus sequence.
"Conservative amino acid substitutions" are those substitutions that, when made, least interfere with the properties of the original protein, i.e., the structure and especially the function of the protein is conserved and not significantly changed by such substitutions.
The table below shows amino acids which may be substituted for an original amino acid in a protein and which are regarded as conservative amino acid substitutions.
Original Residue Conservative Substitution Ala Gly, Ser Arg His, Lys Asn Asp, Gln, His Asp Asn, Glu Cys Ala, Ser Gln Asn, Glu, His Glu Asp, Gln, His Gly Ala His Asn, Arg, Gln, Glu Ile Leu, Val Leu Ile, Val Lys Arg, Gln, Glu Met Leu, Ile Phe His, Met, Leu, Trp, Tyr Ser Cys, Thr Thr Ser, Val Trp Phe, Tyr Tyr His, Phe, Trp Val Ile. Leu, Thr Conservative amino acid substitutions generally maintain (a) the structure of the polypeptide backbone in the area of the substitution, for example, as a beta sheet or alpha helical conformation, (b) the charge or hydrophobicity of the molecule at the site of the substitution, and/or (c) the bulk of the side chain.
A "deletion" refers to a change in the amino acid or nucleotide sequence that results in the absence of one or more amino acid residues or nucleotides.
The term "derivative" refers to the chemical modification of a polypeptide sequence, or a polynucleotide sequence. Chemical modifications of a polynucleotide sequence can include, for example, replacement of hydrogen by an alkyl, acyl, hydroxyl, or amino group.
A derivative polynucleotide encodes a polypeptide which retains at least one biological or immunological function of the natural molecule. A derivative polypeptide is one modified by glycosylation, pegylation, or any similar process that retains at least one biological or immunological function of the polypeptide from which it was derived.
A "fragment" is a unique portion of IMOL or the polynucleotide encoding IMOL
which is identical in sequence to but shorter in length than the parent sequence. A
fragment may comprise up to the entire length of the defined sequence, minus one nucleotide/amino acid residue. For example, a fragment may comprise from 5 to 1000 contiguous nucleotides or amino acid residues. A fragment used as a probe, primer, antigen, therapeutic molecule, or for other purposes, may be at least 5, 10, 15, 16, 20, 25, 30, 40, 50, 60, 75, 100, 150, 250 or at least 500 contiguous nucleotides or amino acid residues in length. Fragments may be preferentially selected from certain regions of a molecule. For example, a polypeptide fragment may comprise a certain length of contiguous amino acids selected from the first 250 or 500 amino acids (or first 25% or 50% of a polypeptide) as shown in a certain defined sequence. Clearly these lengths are exemplary, and any length that is supported by the specification, including the Sequence Listing, tables, and figures, may be encompassed by the present embodiments.
A fragment of SEQ ID N0:16-30 comprises a region of unique polynucleotide sequence that specifically identifies SEQ ID N0:16-30, for example, as distinct from any other sequence in the same genome. A fragment of SEQ ID N0:16-30 is useful, for example, in hybridization and amplification technologies and in analogous methods that distinguish SEQ ID
N0:16-30 from related polynucleotide sequences. The precise length of a fragment of SEQ ID N0:16-30 and the region of SEQ ID N0:16-30 to which the fragment corresponds are routinely determinable by one of ordinary skill in the art based on the intended purpose for the fragment.
A fragment of SEQ ID NO:1-15 is encoded by a fragment of SEQ ID N0:16-30. A
fragment of SEQ ID NO:1-15 comprises a region of unique amino acid sequence that specifically identifies SEQ ID NO:1-15. For example, a fragment of SEQ ID NO:1-15 is useful as an immunogenic peptide for the development of antibodies that specifically recognize SEQ ID NO:1-15.
The precise length of a fragment of SEQ ID NO:1-15 and the region of SEQ ID NO:1-15 to which the fragment corresponds are routinely determinable by one of ordinary skill in the art based on the intended purpose for the fragment.
The term "similarity" refers to a degree of complementarity. There may be partial similarity or complete similarity. The word "identity" may substitute for the word "similarity." A partially complementary sequence that at least partially inhibits an identical sequence from hybridizing to a target nucleic acid is referred to as "substantially similar." The inhibition of hybridization of the completely complementary sequence to the target sequence may be examined using a hybridization assay (Southern or northern blot, solution hybridization, and the like) under conditions of reduced stringency. A substantially similar sequence or hybridization probe will compete for and inhibit the binding of a completely similar (identical) sequence to the target sequence under conditions of reduced stringency. This is not to say that conditions of reduced stringency are such that non-specific binding is permitted, as reduced stringency conditions require that the binding of two sequences to one another be a specific (i.e., a selective) interaction. The absence of non-specific binding may be tested by the use of a second target sequence which lacks even a partial degree of complementarity (e.g., less than about 30% similarity or identity). In the absence of non-specific binding, the substantially similar sequence or probe will not hybridize to the second non-complementary target sequence.
The phrases "percent identity" and "% identity," as applied to polynucleotide sequences, refer to the percentage of residue matches between at least two polynucleotide sequences aligned using a standardized algorithm. Such an algorithm may insert, in a standardized and reproducible way, gaps in the sequences being compared in order to optimize alignment between two sequences, and therefore achieve a more meaningful comparison of the two sequences.
Percent identity between polynucleotide sequences may be determined using the default parameters of the CLUSTAL V algorithm as incorporated into the MEGALIGN
version 3.12e sequence alignment program. This program is part of the LASERGENE software package, a suite of molecular biological analysis programs (DNASTAR, Madison Wn. CLUSTAL V is described in Higgins, D.G. and P.M. Sharp ( 1989) CABIOS 5:151-153 and in Higgins, D.G. et al. ( 1992) CABIOS
8:189-191. For pairwise alignments of polynucleotide sequences, the default parameters are set as follows: Ktuple=2, gap penalty=5, window=4, and "diagonals saved"=4. The "weighted" residue weight table is selected as the default. Percent identity is reported by CLUSTAL V as the "percent similarity" between aligned polynucleotide sequence pairs.
Alternatively, a suite of commonly used and freely available sequence comparison algorithms is provided by the National Center for Biotechnology Information (NCBI) Basic Local Alignment Search Tool (BLAST) (Altschul, S.F. et al. (1990) J. Mol. Biol. 215:403-410), which is available from several sources, including the NCBI, Bethesda, MD, and on the Internet at http://www.ncbi.nlm.nih.gov/BLAST/. The BLAST software suite includes various sequence analysis programs including "blastn," that is used to align a known polynucleotide sequence with other polynucleotide sequences from a variety of databases. Also available is a tool called "BLAST 2 Sequences" that is used for direct pairwise comparison of two nucleotide sequences. "BLAST 2 Sequences" can be accessed and used interactively at http://www.ncbi.nlm.nih.gov/gorf/bl2.html.
The "BLAST 2 Sequences" tool can be used for both blastn and blastp (discussed below). BLAST
programs are commonly used with gap and other parameters set to default settings. For example, to compare two nucleotide sequences, one may use blastn with the "BLAST 2 Sequences" tool Version 2Ø9 (May-07-1999) set at default parameters. Such default parameters may be, for example:
Matrix: BLOSUM62 Reward for match: I
Penalty for mismatch: -2 Open Gap: 5 and Extension Gap: 2 penalties Gap x drop-off. 50 Expect: 10 Word Size: 11 Filter: on Percent identity may be measured over the length of an entire defined sequence, for example, as defined by a particular SEQ ID number, or may be measured over a shorter length, for example, over the length of a fragment taken from a larger, defined sequence, for instance, a fragment of at least 20, at least 30, at least 40, at least 50, at least 70, at least 100, or at least 200 contiguous nucleotides. Such lengths are exemplary only, and it is understood that any fragment length supported by the sequences shown herein, in the tables, figures, or Sequence Listing, may be used to describe a length over which percentage identity may be measured.
Nucleic acid sequences that do not show a high degree of identity may nevertheless encode similar amino acid sequences due to the degeneracy of the genetic code. It is understood that changes in a nucleic acid sequence can be made using this degeneracy to produce multiple nucleic acid sequences that all encode substantially the same protein.
The phrases "percent identity" and "% identity," as applied to polypeptide sequences, refer to the percentage of residue matches between at least two polypeptide sequences aligned using a standardized algorithm. Methods of polypeptide sequence alignment are well-known. Some alignment methods take into account conservative amino acid substitutions.
Such conservative substitutions, explained in more detail above, generally preserve the hydrophobicity and acidity at the site of substitution, thus preserving the structure (and therefore function) of the polypeptide.

Percent identity between polypeptide sequences may be determined using the default parameters of the CLUSTAL V algorithm as incorporated into the MEGALIGN
version 3.12e sequence alignment program (described and referenced above). For pairwise alignments of polypeptide sequences using CLUSTAL V, the default parameters are set as follows: Ktuple=1, gap penalty=3, window=5, and "diagonals saved"=5. The PAM250 matrix is selected as the default residue weight table. As with polynucleotide alignments, the percent identity is reported by CLUSTAL V as the "percent similarity" between aligned polypeptide sequence pairs.
Alternatively the NCBI BLAST software suite may be used. For example, for a pairwise comparison of two polypeptide sequences, one may use the "BLAST 2 Sequences"
tool Version 2Ø9 (May-07-1999) with blastp set at default parameters. Such default parameters may be, for example:
Matrix: BLOSUM62 Open Gap: ll and Extension Gap: I penalties Gap x drop-off. SO
Expect: 10 Word Size: 3 Filter: on Percent identity may be measured over the length of an entire defined polypeptide sequence, for example, as defined by a particular SEQ ID number, or may be measured over a shorter length, for example, over the length of a fragment taken from a larger, defined polypeptide sequence, for instance, a fragment of at least 15, at least 20, at least 30, at least 40, at least 50, at least 70 or at least 150 contiguous residues. Such lengths are exemplary only, and it is understood that any fragment length supported by the sequences shown herein, in the tables, figures or Sequence Listing, may be used to describe a length over which percentage identity may be measured.
"Human artificial chromosomes" (HACs) are linear microchromosomes which may contain DNA sequences of about 6 kb to 10 Mb in size, and which contain all of the elements required for stable mitotic chromosome segregation and maintenance.
The term "humanized antibody" refers to antibody molecules in which the amino acid sequence in the non-antigen binding regions has been altered so that the antibody more closely resembles a human antibody, and still retains its original binding ability.
"Hybridization" refers to the process by which a polynucleotide strand anneals with a complementary strand through base pairing under defined hybridization conditions. Specific hybridization is an indication that two nucleic acid sequences share a high degree of identity.
Specific hybridization complexes form under permissive annealing conditions and remain hybridized after the "washing" step(s). The washing steps) is particularly important in determining the stringency of the hybridization process, with more stringent conditions allowing less non-specific binding, i.e., binding between pairs of nucleic acid strands that are not perfectly matched. Permissive conditions for annealing of nucleic acid sequences are routinely determinable by one of ordinary skill in the art and may be consistent among hybridization experiments, whereas wash conditions may be varied among experiments to achieve the desired stringency, and therefore hybridization specificity.
Permissive annealing conditions occur, for example, at 68°C in the presence of about 6 x SSC, about 1 % (w/v) SDS, and about 100 pg/ml denatured salmon sperm DNA.
Generally, stringency of hybridization is expressed, in part, with reference to the temperature under which the wash step is carried out. Generally, such wash temperatures are selected to be about 5°C to 20°C lower than the thermal melting point (T"~ for the specific sequence at a defined ionic strength and pH. The Tm is the temperature (under defined ionic strength and pH) at which 50% of the target sequence hybridizes to a perfectly matched probe. An equation for calculating Tm and conditions for nucleic acid hybridization are well known and can be found in Sambrook et al., 1989, Molecular Cloning: A Laboratory Manual, 2"° ed.; vol. 1-3, Cold Spring Harbor Press, Plainview NY;
specifically see volume 2, chapter 9.
High stringency conditions for hybridization between polynucleotides of the present invention include wash conditions of 68°C in the presence of about 0.2 x SSC and about 0.1% SDS, for 1 hour. Alternatively, temperatures of about 65°C, 60°C, 55°C, or 42°C may be used. SSC
concentration may be varied from about 0.1 to 2 x SSC, with SDS being present at about 0.1 %.
Typically, blocking reagents are used to block non-specific hybridization.
Such blocking reagents include, for instance, denatured salmon sperm DNA at about 100-200 ~g/ml.
Organic solvent, such as formamide at a concentration of about 35-50% v/v, may also be used under particular circumstances, such as for RNA:DNA hybridizations. Useful variations on these wash conditions will be readily apparent to those of ordinary skill in the art. Hybridization, particularly under high stringency conditions, may be suggestive of evolutionary similarity between the nucleotides. Such similarity is strongly indicative of a similar role for the nucleotides and their encoded polypeptides.
The term "hybridization complex" refers to a complex formed between two nucleic acid sequences by virtue of the formation of hydrogen bonds between complementary bases. A
hybridization complex may be formed in solution (e.g., Cot or Rot analysis) or formed between one nucleic acid sequence present in solution and another nucleic acid sequence immobilized on a solid support (e.g., paper, membranes, filters, chips, pins or glass slides, or any other appropriate substrate to which cells or their nucleic acids have been fixed).
The words "insertion" and "addition" refer to changes in an amino acid or nucleotide sequence resulting in the addition of one or more amino acid residues or nucleotides, respectively.
"Immune response" can refer to conditions associated with inflammation, trauma, immune disorders, or infectious or genetic disease, etc. These conditions can be characterized by expression of various factors, e.g., cytokines, chemokines, and other signaling molecules, which may affect cellular and systemic defense systems.
An "immunogenic fragment" is a polypeptide or oligopeptide fragment of IMOL
which is capable of eliciting an immune response when introduced into a living organism, for example, a mammal. The term "immunogenic fragment" also includes any polypeptide or oligopeptide fragment of IMOL which is useful in any of the antibody production methods disclosed herein or known in the art.
The term "microarray" refers to an arrangement of distinct polynucleotides on a substrate.
The terms "element" and "array element" in a microarray context, refer to hybridizable polynucleotides arranged on the surface of a substrate.
The term "modulate" refers to a change in the activity of IMOL. For example, modulation may cause an increase or a decrease in protein activity, binding characteristics, or any other biological, functional, or immunological properties of IMOL.
The phrases "nucleic acid" and "nucleic acid sequence" refer to a nucleotide, oligonucleotide, polynucleotide, or any fragment thereof. These phrases also refer to DNA or RNA of genomic or synthetic origin which may be single-stranded or double-stranded and may represent the sense or the antisense strand, to peptide nucleic acid (PNA), or to any DNA-like or RNA-like material.
"Operably linked" refers to the situation in which a first nucleic acid sequence is placed in a functional relationship with the second nucleic acid sequence. For instance, a promoter is operably linked to a coding sequence if the promoter affects the transcription or expression of the coding sequence. Generally, operably linked DNA sequences may be in close proximity or contiguous and, where necessary to join two protein coding regions, in the same reading frame.
"Peptide nucleic acid" (PNA) refers to an antisense molecule or anti-gene agent which comprises an oligonucleotide of at least about 5 nucleotides in length linked to a peptide backbone of amino acid residues ending in lysine. The terminal lysine confers solubility to the composition.
PNAs preferentially bind complementary single stranded DNA or RNA and stop transcript elongation, and may be pegylated to extend their lifespan in the cell.
"Probe" refers to nucleic acid sequences encoding IMOL, their complements, or fragments thereof, which are used to detect identical, allelic or related nucleic acid sequences. Probes are isolated oligonucleotides or polynucleotides attached to a detectable label or reporter molecule.
Typical labels include radioactive isotopes, ligands, chemiluminescent agents, and enzymes.
"Primers" are short nucleic acids, usually DNA oligonucleotides, which may be annealed to a target polynucleotide by complementary base-pairing. The primer may then be extended along the target DNA strand by a DNA polymerise enzyme. Primer pairs can be used for amplification (and identification) of a nucleic acid sequence, e.g., by the polymerise chain reaction (PCR).

Probes and primers as used in the present invention typically comprise at least 15 contiguous nucleotides of a known sequence. In order to enhance specificity, longer probes and primers may also be employed, such as probes and primers that comprise at least 20, 25, 30, 40, 50, 60, 70, 80, 90, 100, or at least 150 consecutive nucleotides of the disclosed nucleic acid sequences. Probes and primers may be considerably longer than these examples, and it is understood that any length supported by the specification, including the tables, figures, and Sequence Listing, may be used.
Methods for preparing and using probes and primers are described in the references, for example Sambrook et al., 1989, Molecular Clonins: A Laboratory Manual, 2"d ed., vol. 1-3, Cold Spring Harbor Press, Plainview NY; Ausubel et a1.,1987, Current Protocols in Molecular BioloQV, Greene Publ. Assoc. & Wiley-Intersciences, New York NY; Innis et al., 1990, PCR Protocols. A
Guide to Methods and Applications, Academic Press, San Diego CA. PCR primer pairs can be derived from a known sequence, for example, by using computer programs intended for that purpose such as Primer (Version 0.5, 1991, Whitehead Institute for Biomedical Research, Cambridge MA).
Oligonucleotides for use as primers are selected using software known in the art for such purpose. For example, OLIGO 4.06 software is useful for the selection of PCR
primer pairs of up to 100 nucleotides each, and for the analysis of oligonucleotides and larger polynucleotides of up to 5,000 nucleotides from an input polynucleotide sequence of up to 32 kilobases.
Similar primer selection programs have incorporated additional features for expanded capabilities. For example, the PrimOU primer selection program (available to the public from the Genome Center at University of Texas South West Medical Center, Dallas TX) is capable of choosing specific primers from megabase sequences and is thus useful for designing primers on a genome-wide scope. The Primer3 primer selection program (available to the public from the Whitehead Institute/MTT Center for Genome Research, Cambridge MA) allows the user to input a "mispriming library," in which sequences to avoid as primer binding sites are user-specified. Primer3 is useful, in particular, for the selection of oligonucleotides for microarrays. (The source code for the latter two primer selection programs may also be obtained from their respective sources and modified to meet the user's specific needs.) The PrimeGen program (available to the public from the UK Human Genome Mapping Project Resource Centre, Cambridge UK) designs primers based on multiple sequence alignments, thereby allowing selection of primers that hybridize to either the most conserved or least conserved regions of aligned nucleic acid sequences. Hence, this program is useful for identification of both unique and conserved oligonucleotides and polynucleotide fragments. The oligonucleotides and polynucleotide fragments identified by any of the above selection methods are useful in hybridization technologies, for example, as PCR or sequencing primers, microarray elements, or specific probes to identify fully or partially complementary polynucleotides in a sample of nucleic acids. Methods of oligonucleotide selection are not limited to those described above.

A "recombinant nucleic acid" is a sequence that is not naturally occurring or has a sequence that is made by an artificial combination of two or more otherwise separated segments of sequence.
This artificial combination is often accomplished by chemical synthesis or, more commonly, by the artificial manipulation of isolated segments of nucleic acids, e.g., by genetic engineering techniques such as those described in Sambrook, supra. The term recombinant includes nucleic acids that have been altered solely by addition, substitution, or deletion of a portion of the nucleic acid. Frequently, a recombinant nucleic acid may include a nucleic acid sequence operably linked to a promoter sequence. Such a recombinant nucleic acid may be part of a vector that is used, for example, to transform a cell.
Alternatively, such recombinant nucleic acids may be part of a viral vector, e.g., based on a vaccinia virus, that could be use to vaccinate a mammal wherein the recombinant nucleic acid is expressed, inducing a protective immunological response in the mammal.
An "RNA equivalent," in reference to a DNA sequence, is composed of the same linear sequence of nucleotides as the reference DNA sequence with the exception that all occurrences of the nitrogenous base thymine are replaced with uracil, and the sugar backbone is composed of ribose instead of deoxyribose.
The term "sample" is used in its broadest sense. A sample suspected of containing nucleic acids encoding IMOL, or fragments thereof, or IMOL itself, may comprise a bodily fluid; an extract from a cell, chromosome, organelle, or membrane isolated from a cell; a cell;
genomic DNA, RNA, or cDNA, in solution or bound to a substrate; a tissue; a tissue print; etc.
The terms "specific binding" and "specifically binding" refer to that interaction between a protein or peptide and an agonist, an antibody, an antagonist, a small molecule, or any natural or synthetic binding composition. The interaction is dependent upon the presence of a particular structure of the protein, e.g., the antigenic determinant or epitope, recognized by the binding molecule. For example, if an antibody is specific for epitope "A," the presence of a polypeptide containing the epitope A, or the presence of free unlabeled A, in a reaction containing free labeled A
and the antibody will reduce the amount of labeled A that binds to the antibody.
The term "substantially purified" refers to nucleic acid or amino acid sequences that are removed from their natural environment and are isolated or separated, and are at least 60% free, preferably at least 75% free, and most preferably at least 90% free from other components with which they are naturally associated.
A "substitution" refers to the replacement of one or more amino acids or nucleotides by different amino acids or nucleotides, respectively.
"Substrate" refers to any suitable rigid or semi-rigid support including membranes, filters, chips, slides, wafers, fibers, magnetic or nonmagnetic beads, gels, tubing, plates, polymers, microparticles and capillaries. The substrate can have a variety of surface forms, such as wells, trenches, pins, channels and pores, to which polynucleotides or polypeptides are bound.
"Transformation" describes a process by which exogenous DNA enters and changes a recipient cell. Transformation may occur under natural or artificial conditions according to various methods well known in the art, and may rely on any known method for the insertion of foreign nucleic acid sequences into a prokaryotic or eukaryotic host cell. The method for transformation is selected based on the type of host cell being transformed and may include, but is not limited to, viral infection, electroporation, heat shock, lipofection, and particle bombardment.
The term "transformed" cells includes stably transformed cells in which the inserted DNA is capable of replication either as an autonomously replicating plasmid or as part of the host chromosome, as well as transiently transformed cells which express the inserted DNA or RNA for limited periods of time.
A "transgenic organism;" as used herein, is any organism, including but not limited to animals and plants, in which one or more of the cells of the organism contains heterologous nucleic acid introduced by way of human intervention, such as by transgenic techniques well known in the art. The nucleic acid is introduced into the cell, directly or indirectly by introduction into a precursor of the cell, by way of deliberate genetic manipulation, such as by microinjection or by infection with a recombinant virus. The term genetic manipulation does not include classical cross-breeding, or in vitro fertilization, but rather is directed to the introduction of a recombinant DNA molecule. The transgenic organisms contemplated in accordance with the present invention include bacteria, cyanobacteria, fungi, and plants and animals. The isolated DNA of the present invention can be introduced into the host by methods known in the art, for example infection, transfection, transformation or transconjugation. Techniques for transferring the DNA of the present invention into such organisms are widely known and provided in references such as Sambrook et al. (1989), s_~ra.
A "variant" of a particular nucleic acid sequence is defined as a nucleic acid sequence having at least 40% sequence identity to the particular nucleic acid sequence over a certain length of one of the nucleic acid sequences using blastn with the "BLAST 2 Sequences" tool Version 2Ø9 (May-07-1999) set at default parameters. Such a pair of nucleic acids may show, for example, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95% or at least 98% or greater sequence identity over a certain defined length. A variant may be described as, for example, an "allelic" (as defined above), "splice," "species," or "polymorphic"
variant. A splice variant may have significant identity to a reference molecule, but will generally have a greater or lesser number of polynucleotides due to alternate splicing of exons during mRNA processing. The corresponding polypeptide may possess additional functional domains or lack domains that are present in the reference molecule. Species variants are polynucleotide sequences that vary from one species to another. The resulting polypeptides generally will have significant amino acid identity relative to each other. A polymorphic variant is a variation in the polynucleotide sequence of a particular gene between individuals of a given species. Polymorphic variants also may encompass "single nucleotide polymorphisms" (SNPs) in which the polynucleotide sequence varies by one nucleotide base. The presence of SNPs may be indicative of, for example, a certain population, a disease state, or a propensity for a disease state.
A "variant" of a particular polypeptide sequence is defined as a polypeptide sequence having at least 40% sequence identity to the particular polypeptide sequence over a certain length of one of the polypeptide sequences using blastp with the "BLAST 2 Sequences" tool Version 2Ø9 (May-07-1999) set at default parameters. Such a pair of polypeptides may show, for example, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 98% or greater sequence identity over a certain defined length of one of the polypeptides.
THE INVENTION
The invention is based on the discovery of new human immune system molecules (IMOL), the polynucleotides encoding IMOL, and the use of these compositions for the diagnosis, treatment, or prevention of immunological disorders, infections, and cell proliferative disorders including cancer.
Table 1 lists the Incyte clones used to assemble full length nucleotide sequences encoding IMOL. Columns 1 and 2 show the sequence identification numbers (SEQ m NOs) of the polypeptide and nucleotide sequences, respectively. Column 3 shows the clone IDs of the Incyte clones in which nucleic acids encoding each IMOL were identified, and column 4 shows the cDNA
libraries from which these clones were isolated. Column 5 shows Incyte clones and their corresponding cDNA
libraries. Clones for which cDNA libraries are not indicated were derived from pooled cDNA
libraries. The Incyte clones in column 5 were used to assemble the consensus nucleotide sequence of each IMOL and are useful as fragments in hybridization technologies.
The columns of Table 2 show various properties of each of the polypeptides of the invention:
column 1 references the SEQ ID NO; column 2 shows the number of amino acid residues in each polypeptide; column 3 shows potential phosphorylation sites; column 4 shows potential glycosylation sites; column 5 shows the amino acid residues comprising signature sequences and motifs; column 6 shows homologous sequences as identified by BLAST analysis; and column 7 shows analytical methods and in some cases, searchable databases to which the analytical methods were applied. The methods of column 7 were used to characterize each polypeptide through sequence homology and protein motifs.
The columns of Table 3 show the tissue-specificity and diseases, disorders, or conditions associated with nucleotide sequences encoding IMOL. The first column of Table 3 lists the nucleotide SEQ ID NOs. Column 2 lists fragments of the nucleotide sequences of column 1. These fragments are useful, for example, in hybridization or amplification technologies to identify SEQ 1D
N0:16-30 and to distinguish between SEQ )D N0:16-30 and related polynucleotide sequences. The polypeptides encoded by these fragments are useful, for example, as immunogenic peptides. Column 3 lists tissue categories which express IMOL as a fraction of total tissues expressing IMOL. Column 4 lists diseases, disorders, or conditions associated with those tissues expressing IMOL as a fraction of total tissues expressing IMOL. Column 5 lists the vectors used to subclone each cDNA library.
The columns of Table 4 show descriptions of the tissues used to construct the cDNA libraries from which cDNA clones encoding IMOL were isolated. Column I references the nucleotide SEQ B7 NOs, column 2 shows the cDNA libraries from which these clones were isolated, and column 3 shows the tissue origins and other descriptive information relevant to the cDNA
libraries in column 2.
SEQ )D N0:26 maps to chromosome 16 within the interval from 19.70 to 33.30 centiMorgans. This interval also contains genes and ESTs associated with B
cell maturation and MHC Class II transactivation. SEQ )D N0:29 maps to chromosome 1 I within the interval from 104.80 to 123.50 centiMorgans. This interval also contains genes and ESTs associated with human lymphoma.
The invention also encompasses IMOL variants. A preferred IMOL variant is one which has at least about 80%, or alternatively at least about 90%, or even at least about 95% amino acid sequence identity to the IMOL amino acid sequence, and which contains at least one functional or structural characteristic of IMOL.
The invention also encompasses polynucleotides which encode IMOL. In a particular embodiment, the invention encompasses a polynucleotide sequence comprising a sequence selected from the group consisting of SEQ )D N0:16-30, which encodes IMOL. The polynucleotide sequences of SEQ B? N0:16-30, as presented in the Sequence Listing, embrace the equivalent RNA
sequences, wherein occurrences of the nitrogenous base thymine are replaced with uracil, and the sugar backbone is composed of ribose instead of deoxyribose.
The invention also encompasses a variant of a polynucleotide sequence encoding IMOL. In particular, such a variant polynucleotide sequence will have at least about 80%, or alternatively at least about 90%, or even at least about 95% polynucleotide sequence identity to the polynucleotide sequence encoding IMOL. A particular aspect of the invention encompasses a variant of a polynucleotide sequence comprising a sequence selected from the group consisting of SEQ )D
N0:16-30 which has at least about 80%, or alternatively at least about 90%, or even at least about 95% polynucleotide sequence identity to a nucleic acid sequence selected from the group consisting of SEQ )D N0:16-30. Any one of the polynucleotide variants described above can encode an amino acid sequence which contains at least one functional or structural characteristic of IMOL.

It will be appreciated by those skilled in the art that as a result of the degeneracy of the genetic code, a multitude of polynucleotide sequences encoding IMOL, some bearing minimal similarity to the polynucleotide sequences of any known and naturally occurnng gene, may be produced. Thus, the invention contemplates each and every possible variation of polynucleotide sequence that could be made by selecting combinations based on possible codon choices. These combinations are made in accordance with the standard triplet genetic code as applied to the polynucleotide sequence of naturally occurring IMOL, and all such variations are to be considered as being specifically disclosed.
Although nucleotide sequences which encode IMOL and its variants are generally capable of hybridizing to the nucleotide sequence of the naturally occurring IMOL under appropriately selected conditions of stringency, it may be advantageous to produce nucleotide sequences encoding IMOL or its derivatives possessing a substantially different codon usage, e.g., inclusion of non-naturally occurring codons. Codons may be selected to increase the rate at which expression of the peptide occurs in a particular prokaryotic or eukaryotic host in accordance with the frequency with which particular codons are utilized by the host. Other reasons for substantially altering the nucleotide sequence encoding IMOL and its derivatives without altering the encoded amino acid sequences include the production of RNA transcripts having more desirable properties, such as a greater half life, than transcripts produced from the naturally occurring sequence.
The invention also encompasses production of DNA sequences which encode IMOL
and IMOL derivatives, or fragments thereof, entirely by synthetic chemistry. After production, the synthetic sequence may be inserted into any of the many available expression vectors and cell systems using reagents well known in the art. Moreover, synthetic chemistry may be used to introduce mutations into a sequence encoding IMOL or any fragment thereof.
Also encompassed by the invention are polynucleotide sequences that are capable of hybridizing to the claimed polynucleotide sequences, and, in particular, to those shown in SEQ 1D
N0:16-30 and fragments thereof under various conditions of stringency. (See, e.g., Wahl, G.M. and S.L. Berger ( 1987) Methods Enzymol. 152:399-407; Kimmel, A.R. ( 1987) Methods Enzymol.
152:507-51 l.) Hybridization conditions, including annealing and wash conditions, are described in "Definitions."
Methods for DNA sequencing are well known in the art and may be used to practice any of the embodiments of the invention. The methods may employ such enzymes as the Klenow fragment of DNA polymerase I, SEQUENASE (US Biochemical, Cleveland OH), Taq polymerase (Perkin-Elmer), thermostable T7 polymerase (Amersham Pharmacia Biotech, Piscataway NJ), or combinations of polymerases and proofreading exonucleases such as those found in the ELONGASE
amplification system (Life Technologies, Gaithersburg MD). Preferably, sequence preparation is automated with machines such as the MICROLAB 2200 liquid transfer system (Hamilton, Reno NV), PTC200 thermal cycler (MJ Research, Watertown MA) and ABI CATALYST 800 thermal cycler (Perkin-Elmer). Sequencing is then carned out using either the ABI 373 or 377 DNA sequencing system (Perkin-Elmer), the MEGABACE 1000 DNA sequencing system (Molecular Dynamics, Sunnyvale CA), or other systems known in the art. The resulting sequences are analyzed using a variety of algorithms which are well known in the art. (See, e.g., Ausubel, F.M. (1997) Short Protocols in Molecular Biolo~y> John Wiley & Sons, New York NY, unit 7.7;
Meyers, R.A. (1995) Molecular Biolo~v and Biotechnology, Wiley VCH, New York NY, pp. 856-853.) The nucleic acid sequences encoding IMOL may be extended utilizing a partial nucleotide sequence and employing various PCR-based methods known in the art to detect upstream sequences, such as promoters and regulatory elements. For example, one method which may be employed, restriction-site PCR, uses universal and nested primers to amplify unknown sequence from genomic DNA within a cloning vector. (See; e.g., Sarkar, G. (1993) PCR Methods Applic.
2:318-322.) Another method, inverse PCR, uses primers that extend in divergent directions to amplify unknown sequence from a circularized template. The template is derived from restriction fragments comprising a known genomic locus and surrounding sequences. (See, e.g., Triglia, T. et al. (1988) Nucleic Acids Res. 16:8186.) A third method, capture PCR, involves PCR amplification of DNA
fragments adjacent to known sequences in human and yeast artificial chromosome DNA.
(See, e.g., Lagerstrom, M. et al. ( 1991 ) PCR Methods Applic. 1:111-119.) In this method, multiple restriction enzyme digestions and ligations may be used to insert an engineered double-stranded sequence into a region of unknown sequence before performing PCR. Other methods which may be used to retrieve unknown sequences are known in the art. (See, e.g., Parker, J.D. et al. (1991) Nucleic Acids Res.
19:3055-3060). Additionally, one may use PCR, nested primers, and PROMOTERFINDER libraries (Clontech, Palo Alto CA) to walk genomic DNA. This procedure avoids the need to screen libraries and is useful in finding intron/exon junctions. For all PCR-based methods, primers may be designed using commercially available software, such as OLIGO 4.06 Primer Analysis software (National Biosciences, Plymouth MN) or another appropriate program, to be about 22 to 30 nucleotides in length, to have a GC content of about 50% or more, and to anneal to the template at temperatures of about 68°C to 72°C.
When screening for full-length cDNAs, it is preferable to use libraries that have been size-selected to include larger cDNAs. In addition, random-primed libraries, which often include sequences containing the 5' regions of genes, are preferable for situations in which an oligo d(T) library does not yield a full-length cDNA. Genomic libraries may be useful for extension of sequence into 5' non-transcribed regulatory regions.
Capillary electrophoresis systems which are commercially available may be used to analyze the size or confirm the nucleotide sequence of sequencing or PCR products. In particular, capillary sequencing may employ flowable polymers for electrophoretic separation, four different nucleotide-specific, laser-stimulated fluorescent dyes, and a charge coupled device camera for detection of the emitted wavelengths. Output/light intensity may be converted to electrical signal using appropriate software (e.g., GENOTYPER and SEQUENCE NAVIGATOR, Perkin-Elmer), and the entire process from loading of samples to computer analysis and electronic data display may be computer controlled. Capillary electrophoresis is especially preferable for sequencing small DNA fragments which may be present in limited amounts in a particular sample.
In another embodiment of the invention, polynucleotide sequences or fragments thereof which encode IMOL may be cloned in recombinant DNA molecules that direct expression of IMOL, or fragments or functional equivalents thereof, in appropriate host cells. Due to the inherent degeneracy of the genetic code, other DNA sequences which encode substantially the same or a functionally equivalent amino acid sequence may be produced and used to express IMOL.
The nucleotide sequences of the present invention can be engineered using methods generally known in the art in order to alter IMOL-encoding sequences for a variety of purposes including, but not limited to, modification of the cloning, processing, and/or expression of the gene product. DNA
shuffling by random fragmentation and PCR reassembly of gene fragments and synthetic oligonucleotides may be used to engineer the nucleotide sequences. For example, oligonucleotide-mediated site-directed mutagenesis may be used to introduce mutations that create new restriction sites, alter glycosylation patterns, change codon preference, produce splice variants, and so forth.
The nucleotides of the present invention may be subjected to DNA shuffling techniques such as MOLECULARBREEDING (Maxygen Inc., Santa Clam CA; described in U.S. Patent Number 5,837,458; Chang, C.-C. et al. (1999) Nat. Biotechnol. 17:793-797; Christians, F.C. et al. (1999) Nat.
Biotechnol. 17:259-264; and Crameri, A. et al. (1996) Nat. Biotechnol. 14:315-319) to alter or improve the biological properties of IMOL, such as its biological or enzymatic activity or its ability to bind to other molecules or compounds. DNA shuffling is a process by which a library of gene variants is produced using PCR-mediated recombination of gene fragments. The library is then subjected to selection or screening procedures that identify those gene variants with the desired properties. These preferred variants may then be pooled and further subjected to recursive rounds of DNA shuffling and selection/screening. Thus, genetic diversity is created through "artificial"
breeding and rapid molecular evolution. For example, fragments of a single gene containing random point mutations may be recombined, screened, and then reshuffled until the desired properties are optimized. Alternatively, fragments of a given gene may be recombined with fragments of homologous genes in the same gene family, either from the same or different species, thereby maximizing the genetic diversity of multiple naturally occurring genes in a directed and controllable manner.
In another embodiment, sequences encoding IMOL may be synthesized, in whole or in part, using chemical methods well known in the art. (See, e.g., Caruthers, M.H. et al. ( 1980) Nucleic Acids Symp. Ser. 7:215-223; and Horn, T. et al. ( 1980) Nucleic Acids Symp. Ser.
7:225-232.) Alternatively, IMOL itself or a fragment thereof may be synthesized using chemical methods. For example, peptide synthesis can be performed using various solid-phase techniques. (See, e.g., Roberge, J.Y. et al. (1995) Science 269:202-204.) Automated synthesis may be achieved using the ABI 431A peptide synthesizer (Perkin-Elmer). Additionally, the amino acid sequence of IMOL, or any part thereof, may be altered during direct synthesis and/or combined with sequences from other proteins, or any part thereof, to produce a variant polypeptide.
The peptide may be substantially purified by preparative high performance liquid chromatography. (See, e.g., Chiez, R.M. and F.Z: Regnier (1990) Methods Enzymol. 182:392-421.) The composition of the synthetic peptides may be confirmed by amino acid analysis or by sequencing. (See, e.g., Creighton, T. ( 1984) Proteins. Structures and Molecular Properties, WH
Freeman, New York NY.) In order to express a biologically active IMOL, the nucleotide sequences encoding IMOL or derivatives thereof may be inserted into an appropriate expression vector, i.e., a vector which contains the necessary elements for transcriptional and translational control of the inserted coding sequence in a suitable host. These elements include regulatory sequences, such as enhancers, constitutive and inducible promoters, and 5' and 3' untranslated regions in the vector and in polynucleotide sequences encoding IMOL. Such elements may vary in their strength and specificity.
Specific initiation signals may also be used to achieve more efficient translation of sequences encoding IMOL. Such signals include the ATG initiation codon and adjacent sequences, e.g. the Kozak sequence. In cases where sequences encoding IMOL and its initiation codon and upstream regulatory sequences are inserted into the appropriate expression vector, no additional transcriptional or translational control signals may be needed. However, in cases where only coding sequence, or a fragment thereof, is inserted, exogenous translational control signals including an in-frame ATG initiation codon should be provided by the vector. Exogenous translational elements and initiation codons may be of various origins, both natural and synthetic. The efficiency of expression may be enhanced by the inclusion of enhancers appropriate for the particular host cell system used. (See, e.g., Scharf, D. et al. ( 1994) Results Probl. Cell Differ. 20:125-162.) Methods which are well known to those skilled in the art may be used to construct expression vectors containing sequences encoding IMOL and appropriate transcriptional and translational control elements. These methods include in vitro recombinant DNA techniques, synthetic techniques, and in vivo genetic recombination. (See, e.g., Sambrook, J. et al. (1989) Molecular Cloning, A

Laboratory Manual, Cold Spring Harbor Press, Plainview NY, ch. 4, 8, and 16-17; Ausubel, F.M. et al. ( 1995) Current Protocols in Molecular Biolo~y, John Wiley & Sons, New York NY, ch. 9, 13, and 16.) A variety of expression vector/host systems may be utilized to contain and express sequences encoding IMOL. These include, but are not limited to, microorganisms such as bacteria transformed with recombinant bacteriophage, plasmid, or cosmid DNA expression vectors;
yeast transformed with yeast expression vectors; insect cell systems infected with viral expression vectors (e.g., baculovirus);
plant cell systems transformed with viral expression vectors (e.g., cauliflower mosaic virus, CaMV, or tobacco mosaic virus, TMV) or with bacterial expression vectors (e.g., Ti or pBR322 plasmids); or animal cell systems. The invention is not limited by the host cell employed.
In bacterial systems, a number of cloning and expression vectors may be selected depending upon the use intended for polynucleotide sequences encoding IMOL. For example, routine cloning, subcloning; and propagation of polynucleotide sequences encoding IMOL can be achieved using a multifunctional E. coli vector such as PBLUESCRIPT (Stratagene, La Jolla CA) or PSPORT1 plasmid (Life Technologies). Ligation of sequences encoding IMOL into the vector's multiple cloning site disrupts the lacZ gene, allowing a colorimetric screening procedure for identification of transformed bacteria containing recombinant molecules. In addition, these vectors may be useful for in vitro transcription, dideoxy sequencing, single strand rescue with helper phage, and creation of nested deletions in the cloned sequence. (See, e.g., Van Heeke, G. and S.M.
Schuster ( 1989) J. Biol.
Chem. 264:5503-5509.) When large quantities of IMOL are needed, e.g. for the production of antibodies, vectors which direct high level expression of IMOL may be used.
For example, vectors containing the strong, inducible TS or T7 bacteriophage promoter may be used.
Yeast expression systems may be used for production of IMOL. A number of vectors containing constitutive or inducible promoters, such as alpha factor, alcohol oxidase, and PGH
promoters, may be used in the yeast Saccharomyces cerevisiae or Pichia nastoris. In addition, such vectors direct either the secretion or intracellular retention of expressed proteins and enable integration of foreign sequences into the host genome for stable propagation.
(See, e.g., Ausubel, 1995, supra; Bitter, G.A. et al. ( 1987) Methods Enzymol. 153:516-544; and Scorer, C.A. et al. ( 1994) Bio/Technology 12:181-184.) Plant systems may also be used for expression of IMOL. Transcription of sequences encoding IMOL may be driven viral promoters, e.g., the 35S and 19S promoters of CaMV used alone or in combination with the omega leader sequence from TMV (Takamatsu, N.
(1987) EMBO J.

6:307-311). Alternatively, plant promoters such as the small subunit of RUBISCO or heat shock promoters may be used. (See, e.g., Coruzzi, G. et al. (1984) EMBO J. 3:1671-1680; Brogue, R. et al.
(1984) Science 224:838-843; and Winter, J. et al. (1991) Results Probl. Cell Differ. 17:85-105.) These constructs can be introduced into plant cells by direct DNA
transformation or pathogen-mediated transfection. (See, e.g., The McGraw Hill Yearbook of Science and TechnoloQv ( 1992) McGraw Hill, New York NY, pp. 191-196.) In mammalian cells, a number of viral-based expression systems may be utilized. In cases where an adenovirus is used as an expression vector, sequences encoding IMOL
may be ligated into an adenovirus transcription/translation complex consisting of the late promoter and tripartite leader sequence. Insertion in a non-essential E 1 or E3 region of the viral genome may be used to obtain infective virus which expresses IMOL in host cells. (See, e.g., Logan, J. and T. Shenk ( 1984) Proc.
Natl. Acad. Sci. USA 81:3655-3659.) In addition, transcription enhancers, such as the Rous sarcoma virus (RSV) enhancer, may be used to increase expression in mammalian host cells. SV40 or EBV-based vectors may also be used for high-level protein expression.
Human artificial chromosomes (HACs) may also be employed to deliver larger fragments of DNA than can be contained in and expressed from a plasmid. HACs of about 6 kb to 10 Mb are constructed and delivered via conventional delivery methods (liposomes, polycationic amino polymers, or vesicles) for therapeutic purposes. (See, e.g., Harrington, J.J.
et al. ( 1997) Nat. Genet.
15:345-355.) For long term production of recombinant proteins in mammalian systems, stable expression of IMOL in cell lines is preferred. For example, sequences encoding IMOL can be transformed into cell lines using expression vectors which may contain viral origins of replication and/or endogenous expression elements and a selectable marker gene on the same or on a separate vector. Following the introduction of the vector, cells may be allowed to grow for about 1 to 2 days in enriched media before being switched to,selective media. The purpose of the selectable marker is to confer resistance to a selective agent, and its presence allows growth and recovery of cells which successfully express the introduced sequences: Resistant clones of stably transformed cells may be propagated using tissue culture techniques appropriate to the cell type.
Any number of selection systems may be used to recover transformed cell lines.
These include, but are not limited to, the herpes simplex virus thymidine kinase and adenine phosphoribosyltransferase genes, for use in tk~ and apr cells, respectively.
(See, e.g., Wigler, M. et al. (1977) Cell 11:223-232; Lowy, I. et al. (1980) Cell 22:817-823.) Also, antimetabolite, antibiotic, or herbicide resistance can be used as the basis for selection. For example, dhfr confers resistance to methotrexate; neo confers resistance to the aminoglycosides neomycin and G-418; and als and pat confer resistance to chlorsulfuron and phosphinotricin acetyltransferase, respectively. (See, e.g., Wigler, M. et al. (1980) Proc. Natl. Acad. Sci. USA 77:3567-3570; Colbere-Garapin, F. et al. (1981) J. Mol. Biol. 150:1-14.) Additional selectable genes have been described, e.g., trpB and hisD, which alter cellular requirements for metabolites. (See, e.g., Hartman, S.C. and R.C. Mulligan ( 1988) Proc.

Natl. Acad. Sci. USA 85:8047-8051.) Visible markers, e.g., anthocyanins, green fluorescent proteins (GFP; Clontech), B glucuronidase and its substrate B-glucuronide, or luciferase and its substrate luciferin may be used. These markers can be used not only to identify transformants, but also to quantify the amount of transient or stable protein expression attributable to a specific vector system.
(See, e.g., Rhodes, C.A. ( 1995) Methods Mol. Biol. 55:121-131.) Although the presence/absence of marker gene expression suggests that the gene of interest is also present, the presence and expression of the gene may need to be confirmed. For example, if the sequence encoding IMOL is inserted within a marker gene sequence, transformed cells containing sequences encoding IMOL can be identified by the absence of marker gene function. Alternatively, a marker gene can be placed in tandem with a sequence encoding IMOL under the control of a single promoter. Expression of the marker gene in response to induction or selection usually indicates expression of the tandem gene as well.
In general, host cells that contain the nucleic acid sequence encoding IMOL
and that express IMOL may be identified by a variety of procedures known to those of skill in the art. These procedures include, but are not limited to, DNA-DNA or DNA-RNA hybridizations, PCR
amplification, and protein bioassay or immunoassay techniques which include membrane, solution, or chip based technologies for the detection and/or quantification of nucleic acid or protein sequences.
Immunological methods for detecting and measuring the expression of IMOL using either specific polyclonal or monoclonal antibodies are known in the art. Examples of such techniques include enzyme-linked immunosorbent assays (ELISAs), radioimmunoassays (RIAs), and fluorescence activated cell sorting (FACS). A two-site, monoclonal-based immunoassay utilizing monoclonal antibodies reactive to two non-interfering epitopes on IMOL is preferred, but a competitive binding assay may be employed. These and other assays are well known in the art. (See, e.g., Hampton, R. et al. (1990) Serological Methods, a Laboratory Manual, APS
Press, St. Paul MN, Sect. IV; Coligan, J.E. et al. (1997) Current Protocols in Immunoloay, Greene Pub. Associates and Wiley-Interscience, New York NY; and Pound, J.D. (1998) Immunochemical Protocols, Humana Press, Totowa NJ.) A wide variety of labels and conjugation techniques are known by those skilled in the art and may be used in various nucleic acid and amino acid assays. Means for producing labeled hybridization or PCR probes for detecting sequences related to polynucleotides encoding IMOL
include oligolabeling, nick translation, end-labeling, or PCR amplification using a labeled nucleotide.
Alternatively, the sequences encoding IMOL, or any fragments thereof, may be cloned into a vector for the production of an mRNA probe. Such vectors are known in the art, are commercially available, and may be used to synthesize RNA probes in vitro by addition of an appropriate RNA polymerase such as T7, T3, or SP6 and labeled nucleotides. These procedures may be conducted using a variety of commercially available kits, such as those provided by Amersham Pharmacia Biotech, Promega (Madison Wn, and US Biochemical. Suitable reporter molecules or labels which may be used for ease of detection include radionuclides, enzymes, fluorescent, chemiluminescent, or chromogenic agents, as well as substrates, cofactors, inhibitors, magnetic particles, and the like.
Host cells transformed with nucleotide sequences encoding IMOL may be cultured under conditions suitable for the expression and recovery of the protein from cell culture. The protein produced by a transformed cell may be secreted or retained intracellularly depending on the sequence and/or the vector used. As will be understood by those of skill in the art, expression vectors containing polynucleotides which encode IMOL may be designed to contain signal sequences which direct secretion of IMOL through a prokaryotic or eukaryotic cell membrane.
In addition, a host cell strain may be chosen for its ability to modulate expression of the inserted sequences or to process the expressed protein in the desired fashion.
Such modifications of the polypeptide include, but are not limited to, acetylation, carboxylation;
glycosylation, phosphorylation, lipidation, and acylation. Post-translational processing which cleaves a "prepro" or "pro" form of the protein may also be used to specify protein targeting, folding, and/or activity.
Different host cells which have specific cellular machinery and characteristic mechanisms for post-translational activities (e.g., CHO, HeLa, MDCK, HEK293, and WI38) are available from the American Type Culture Collection (ATCC, Manassas VA) and may be chosen to ensure the correct modification and processing of the foreign protein.
In another embodiment of the invention, natural, modified, or recombinant nucleic acid sequences encoding IMOL may be ligated to a heterologous sequence resulting in translation of a fusion protein in any of the aforementioned host systems. For example, a chimeric IMOL protein containing a heterologous moiety that can be recognized by a commercially available antibody may facilitate the screening of peptide libraries for inhibitors of IMOL activity.
Heterologous protein and peptide moieties may also facilitate purification of fusion proteins using commercially available affinity matrices. Such moieties include, but are not limited to, glutathione S-transferase (GST), maltose binding protein (MBP), thioredoxin (Trx), calmodulin binding peptide (CBP), 6-His, FLAG, c-myc, and hemagglutinin (HA). GST, MBP, Trx, CBP, and 6-His enable purification of their cognate fusion proteins on immobilized glutathione, maltose, phenylarsine oxide, calmodulin, and metal-chelate resins, respectively. FLAG, c-myc, and hemagglutinin (HA) enable immunoaffinity purification of fusion proteins using commercially available monoclonal and polyclonal antibodies that specifically recognize these epitope tags. A fusion protein may also be engineered to contain a proteolytic cleavage site located between the IMOL encoding sequence and the heterologous protein sequence, so that IMOL may be cleaved away from the heterologous moiety following purification.
Methods for fusion protein expression and purification are discussed in Ausubel (1995, supra, ch. 10).

A variety of commercially available kits may also be used to facilitate expression and purification of fusion proteins.
In a further embodiment of the invention, synthesis of radiolabeled IMOL may be achieved in vitro using the TNT rabbit reticulocyte lysate or wheat germ extract system (Promega). These systems couple transcription and translation of protein-coding sequences operably associated with the T7, T3, or SP6 promoters. Translation takes place in the presence of a radiolabeled amino acid precursor, for example, 35S-methionine.
Fragments of IMOL may be produced not only by recombinant means, but also by direct peptide synthesis using solid-phase techniques. (See, e.g., Creighton, supra, pp. 55-60.) Protein synthesis may be performed by manual techniques or by automation. Automated synthesis may be achieved, for example, using the ABI 431A peptide synthesizer (Perkin-Elmer).
Various fragments of IMOL may be synthesized separately and then combined to produce the full length molecule.
THERAPEUTICS
Chemical and structural similarity, e.g., in the context of sequences and motifs, exists between regions of IMOL and immune system molecules. In addition, the expression of IMOL is closely associated with immunological disorders, infections, and cell proliferative disorders including cancer. SEQ ID N0:26 maps to a chromosomal interval which also contains genes and ESTs associated with B cell maturation and MHC Class II transactivation. SEQ ID
N0:29 maps to a chromosomal interval which also contains genes and ESTs associated with human lymphoma.
Therefore, IMOL appears to play a role in immunological disorders, infections, and cell proliferative disorders including cancer. In the treatment of disorders associated with increased IMOL expression or activity, it is desirable to decrease the expression or activity of IMOL.
In the treatment of disorders associated with decreased IMOL expression or activity, it is desirable to increase the expression or activity of IMOL.
Therefore, in one embodiment, IMOL or a fragment or derivative thereof may be administered to a subject to treat or prevent a disorder associated with decreased expression or activity of IMOL. Examples of such disorders include, but are not limited to, an immunological disorder such as inflammation, actinic keratosis, acquired immunodeficiency syndrome (AIDS), Addison's disease, adult respiratory distress syndrome, allergies, ankylosing spondylitis, amyloidosis, anemia, arteriosclerosis, asthma, atherosclerosis, autoimmune hemolytic anemia, autoimmune thyroiditis, autoimune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED), bronchitis, bursitis, cholecystitis, cirrhosis, contact dermatitis, Crohn's disease, atopic denmatitis, dermatomyositis, diabetes mellitus, emphysema, erythroblastosis fetalis, erythema nodosum, atrophic gastritis, glomerulonephritis, Goodpasture's syndrome, gout, Graves' disease, Hashimoto's thyroiditis, paroxysmal nocturnal hemoglobinuria, hepatitis, hypereosinophilia, irritable bowel syndrome, episodic lymphopenia with lymphocytotoxins, mixed connective tissue disease (MCTD), multiple sclerosis, myasthenia gravis, myocardial or pericardial inflammation, myelofibrosis, osteoarthritis, osteoporosis, pancreatitis, polycythemia vera, polymyositis, psoriasis, Reiter's syndrome, rheumatoid arthritis, scleroderma, Sjogren's syndrome, systemic anaphylaxis, systemic lupus erythematosus, systemic sclerosis, primary thrombocythemia, thrombocytopenic purpura, ulcerative colitis, uveitis, Werner syndrome, complications of cancer, hemodialysis, and extracorporeal circulation, trauma, and hematopoietic cancer including lymphoma, leukemia, and myeloma; and an infection caused by a viral agent classified as adenovirus, arenavirus, bunyavirus, calicivirus, coronavirus, filovirus, hepadnavirus, herpesvirus, flavivirus, orthomyxovirus, parvovirus, papovavirus, paramyxovirus, picomavirus, poxvirus, reovirus, retrovirus, rhabdovirus, or togavirus;
an infection caused by a bacterial agent classified as pneumococcus, staphylococcus, streptococcus, bacillus, corynebacterium, clostridium, meningococcus, gonococcus, listeria, moraxella,.kingella, haemophilus, legionella, bordetella, gram-negative enterobacterium including shigella, salmonella, or campylobacter, pseudomonas, vibrio, brucella, francisella, yersinia, bartonella, norcardium, actinomyces, mycobacterium, spirochaetale, rickettsia, chlamydia, or mycoplasma; an infection caused by a fungal agent classified as aspergillus, blastomyces, dermatophytes, cryptococcus, coccidioides, malasezzia, histoplasma, or other mycosis-causing fungal agent;
and an infection caused by a parasite classified as plasmodium or malaria-causing, parasitic entamoeba, leishmania, trypanosoma, toxoplasma, pneumocystis carinii, intestinal protozoa such as giardia, trichomonas, tissue nematode such as trichinella, intestinal nematode such as ascaris, lymphatic filarial nematode, trematode such as schistosoma, and cestode such as tapeworm; and a cell proliferative disorder such as actinic keratosis, arteriosclerosis, atherosclerosis, bursitis, cirrhosis, hepatitis, mixed connective tissue disease (MCTD), myelofibrosis, paroxysmal nocturnal hemoglobinuria, polycythemia vera, psoriasis, primary thrombocythemia, and cancers including adenocarcinoma, leukemia, lymphoma, melanoma, myeloma, sarcoma, teratocarcinoma, and, in particular, cancers of the adrenal gland, bladder, bone, bone marrow, brain, breast, cervix, gall bladder, ganglia, gastrointestinal tract, heart, kidney, liver, lung, muscle, ovary, pancreas, parathyroid, penis, prostate, salivary glands, skin, spleen, testis, thymus, thyroid, and uterus.
In another embodiment, a vector capable of expressing IMOL or a fragment or derivative thereof may be administered to a subject to treat or prevent a disorder associated with decreased expression or activity of IMOL including, but not limited to, those described above.
In a further embodiment, a pharmaceutical composition comprising a substantially purified IMOL in conjunction with a suitable pharmaceutical carrier may be administered to a subject to treat or prevent a disorder associated with decreased expression or activity of IMOL
including, but not limited to, those provided above.

In still another embodiment, an agonist which modulates the activity of IMOL
may be administered to a subject to treat or prevent a disorder associated with decreased expression or activity of IMOL including, but not limited to, those listed above.
In a further embodiment, an antagonist of IMOL may be administered to a subject to treat or prevent a disorder associated with increased expression or activity of IMOL.
Examples of such disorders include, but are not limited to, those immunological disorders, infections, and cell proliferative disorders including cancer, listed above. In one aspect, an antibody which specifically binds IMOL may be used directly as an antagonist or indirectly as a targeting or delivery mechanism for bringing a pharmaceutical agent to cells or tissues which express IMOL.
In an additional embodiment, a vector expressing the complement of the polynucleotide encoding IMOL may be administered to a subject to treat or prevent a disorder associated with increased expression or activity of IMOL including, but not limited to, those described above.
In other embodiments, any of the proteins, antagonists, antibodies, agonists, complementary sequences, or vectors of the invention may be administered in combination with other appropriate therapeutic agents. Selection of the appropriate agents for use in combination therapy may be made by one of ordinary skill in the art, according to conventional pharmaceutical principles. The combination of therapeutic agents may act synergistically to effect the treatment or prevention of the various disorders described above. Using this approach, one may be able to achieve therapeutic efficacy with lower dosages of each agent, thus reducing the potential for adverse side effects.
An antagonist of IMOL may be produced using methods which are generally known in the art. In particular, purified IMOL may be used to produce antibodies or to screen libraries of pharmaceutical agents to identify those which specifically bind IMOL.
Antibodies to IMOL may also be generated using methods that are well known in the art. Such antibodies may include, but are not limited to, polyclonal, monoclonal, chimeric, and single chain antibodies, Fab fragments, and fragments produced by a Fab expression library. Neutralizing antibodies (i.e., those which inhibit dimer formation) are generally preferred for therapeutic use.
For the production of antibodies, various hosts including goats, rabbits, rats, mice, humans, and others may be immunized by injection with 1MOL or with any fragment or oligopeptide thereof which has immunogenic properties. Depending on the host species, various adjuvants may be used to increase immunological response. Such adjuvants include, but are not limited to, Freund's, mineral gels such as aluminum hydroxide, and surface active substances such as lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, KL,H, and dinitrophenol. Among adjuvants used in humans, BCG (bacilli Calmette-Guerin) and Corvnebacterium parvum are especially preferable.
It is preferred that the oligopeptides, peptides, or fragments used to induce antibodies to IMOL have an amino acid sequence consisting of at least about 5 amino acids, and generally will consist of at least about 10 amino acids. It is also preferable that these oligopeptides, peptides, or fragments are identical to a portion of the amino acid sequence of the natural protein and contain the entire amino acid sequence of a small, naturally occurring molecule. Short stretches of IMOL amino acids may be fused with those of another protein, such as KLH, and antibodies to the chimeric molecule may be produced.
Monoclonal antibodies to IMOL may be prepared using any technique which provides for the production of antibody molecules by continuous cell lines in culture. These include, but are not limited to, the hybridoma technique, the human B-cell hybridoma technique, and the EBV-hybridoma technique. (See, e.g., Kohler, G. et al. (1975) Nature 256:495-497; Kozbor, D.
et al. (1985) J.
Immunol. Methods 81:31-42; Cote, R.J. et al. ( 1983) Proc. Natl. Acad. Sci.
USA 80:2026-2030; and Cole, S.P. et al. (1984) Mol. Cell Biol. 62:109-120.) In addition, techniques developed for the production of "chimeric antibodies,"
such as the splicing of mouse antibody genes to human antibody genes to obtain a molecule with appropriate antigen specificity and biological activity, can be used. (See, e.g., Morrison, S.L. et al. (1984) Proc.
Natl. Acad. Sci. USA 81:6851-6855; Neuberger, M.S. et al. (1984) Nature 312:604-608; and Takeda, S. et al. (1985) Nature 314:452-454.) Alternatively, techniques described for the production of single chain antibodies may be adapted, using methods known in the art, to produce IMOL-specific single chain antibodies. Antibodies with related specificity, but of distinct idiotypic composition, may be generated by chain shuffling from random combinatorial immunoglobulin libraries. (See, e.g., Burton, D.R. (1991) Proc. Natl. Acad. Sci. USA 88:10134-10137.) Antibodies may also be produced by inducing in vivo production in the lymphocyte population or by screening immunoglobulin libraries or panels of highly specific binding reagents as disclosed in the literature. (See, e.g., Orlandi, R. et al. (1989) Proc. Natl.
Acad. Sci. USA
86:3833-3837; Winter, G. et al. (1991) Nature 349:293-299.) Antibody fragments which contain specific binding sites for IMOL may also be generated.
For example, such fragments include, but are not limited to, F(ab')2 fragments produced by pepsin digestion of the antibody molecule and Fab fragments generated by reducing the disulfide bridges of the F(ab')2 fragments. Alternatively, Fab expression libraries may be constructed to allow rapid and easy identification of monoclonal Fab fragments with the desired specificity.
(See, e.g., Huse, W.D.
et al. (1989) Science 246:1275-1281.) Various immunoassays may be used for screening to identify antibodies having the desired specificity. Numerous protocols for competitive binding or immunoradiometric assays using either polyclonal or monoclonal antibodies with established specificities are well known in the art. Such immunoassays typically involve the measurement of complex formation between IMOL and its specific antibody. A two-site, monoclonal-based immunoassay utilizing monoclonal antibodies reactive to two non-interfering IMOL epitopes is generally used, but a competitive binding assay may also be employed (Pound; su ra .
Various methods such as Scatchard analysis in conjunction with radioimmunoassay techniques may be used to assess the affinity of antibodies for IMOL. Affinity is expressed as an association constant, K~, which is defined as the molar concentration of IMOL-antibody complex divided by the molar concentrations of free antigen and free antibody under equilibrium conditions.
The Ka determined for a preparation of polyclonal antibodies, which are heterogeneous in their affinities for multiple IMOL epitopes, represents the average affinity, or avidity, of the antibodies for IMOL. The Ka determined for a preparation of monoclonal antibodies, which are monospecific for a particular IMOL epitope, represents a true measure of affinity. High-affinity antibody preparations with K~ ranging from about 109 to 10'z L/mole are preferred for use in immunoassays in which the IMOL-antibody complex must withstand rigorous manipulations. Low-affinity antibody preparations with K~ ranging from about 106 to 10' L/mole are preferred for use in immunopurification and similar procedures which ultimately require dissociation of IMOL, preferably in active form, from the antibody (Catty, D. (1988) Antibodies. Volume I: A Practical Approach, IRL
Press, Washington, DC;
Liddell, J.E. and Cryer, A. (1991) A Practical Guide to Monoclonal Antibodies, John Wiley & Sons, New York NY).
The titer and avidity of polyclonal antibody preparations may be further evaluated to determine the quality and suitability of such preparations for certain downstream applications. For example, a polyclonal antibody preparation containing at least 1-2 mg specific antibody/ml, preferably 5-10 mg specific antibody/ml, is generally employed in procedures requiring precipitation of IMOL-antibody complexes. Procedures for evaluating antibody specificity, titer, and avidity, and guidelines for antibody quality and usage in various applications, are generally available. (See, e.g., Catty, supra, and Coligan et al. supra.) In another embodiment of the invention, the polynucleotides encoding IMOL, or any fragment or complement thereof, may be used for therapeutic purposes. In one aspect, the complement of the polynucleotide encoding IMOL may be used in situations in which it would be desirable to block the transcription of the mRNA. In particular, cells may be transformed with sequences complementary to polynucleotides encoding IMOL. Thus, complementary molecules or fragments may be used to modulate IMOL activity, or to achieve regulation of gene function. Such technology is now well known in the art, and sense or antisense oligonucleotides or larger fragments can be designed from various locations along the coding or control regions of sequences encoding IMOL.
Expression vectors derived from retroviruses, adenoviruses, or herpes or vaccinia viruses, or from various bacterial plasmids, may be used for delivery of nucleotide sequences to the targeted organ, tissue, or cell population. Methods which are well known to those skilled in the art can be used to construct vectors to express nucleic acid sequences complementary to the polynucleotides encoding IMOL. (See, e.g., Sambrook, s-unra; Ausubel, 1995, supra.) Genes encoding IMOL can be turned off by transforming a cell or tissue with expression vectors which express high levels of a polynucleotide, or fragment thereof, encoding IMOL. Such constructs may be used to introduce untranslatable sense or antisense sequences into a cell. Even in the absence of integration into the DNA, such vectors may continue to transcribe RNA molecules until they are disabled by endogenous nucleases. Transient expression may last for a month or more with a non-replicating vector, and may last even longer if appropriate replication elements are part of the vector system.
As mentioned above, modifications of gene expression can be obtained by designing complementary sequences or antisense molecules (DNA, RNA, or PNA) to the control, 5', or regulatory regions of the gene encoding IMOL. Oligonucleotides derived from the transcription initiation site, e.g., between about positions -10 and +10 from the start site, may be employed.
Similarly, inhibition can be achieved using triple helix base-pairing methodology. Triple helix pairing is useful because it causes inhibition of the ability of the double helix to open sufficiently for the binding of polymerases, transcription factors, or regulatory molecules.
Recent therapeutic advances using triplex DNA have been described in the literature. (See, e.g., Gee, J.E. et al. (1994) in Huber, B.E. and B.I. Carr, Molecular and Immunolo ig c Approaches, Futura Publishing, Mt. Kisco NY, pp. 163-177.) A complementary sequence or antisense molecule may also be designed to block translation of mRNA by preventing the transcript from binding to ribosomes.
Ribozymes, enzymatic RNA molecules, may also be used to catalyze the specific cleavage of RNA. The mechanism of ribozyme action involves sequence-specific hybridization of the ribozyme molecule to complementary target RNA, followed by endonucleolytic cleavage.
For example, engineered hammerhead motif ribozyme molecules may specifically and efficiently catalyze endonucleolytic cleavage of sequences encoding IMOL.
Specific ribozyme cleavage sites within any potential RNA target are initially identified by scanning the target molecule for ribozyme cleavage sites, including the following sequences: GUA, GUU, and GUC. Once identified, short RNA sequences of between 15 and 20 ribonucleotides, corresponding to the region of the target gene containing the cleavage site, may be evaluated for secondary structural features which may render the oligonucleotide inoperable.
The suitability of candidate targets may also be evaluated by testing accessibility to hybridization with complementary oligonucleotides using ribonuclease protection assays.
Complementary ribonucleic acid molecules and ribozymes of the invention may be prepared by any method known in the art for the synthesis of nucleic acid molecules.
These include techniques for chemically synthesizing oligonucleotides such as solid phase phosphoramidite chemical synthesis.
Alternatively, RNA molecules may be generated by in vitro and in vivo transcription of DNA
sequences encoding IMOL. Such DNA sequences may be incorporated into a wide variety of vectors with suitable RNA polymerase promoters such as T7 or SP6. Alternatively, these cDNA constructs that synthesize complementary RNA, constitutively or inducibly, can be introduced into cell lines, cells, or tissues.
RNA molecules may be modified to increase intracellular stability and half life. Possible modifications include, but are not limited to, the addition of flanking sequences at the 5' and/or 3' ends of the molecule, or the use of phosphorothioate or 2' O-methyl rather than phosphodiesterase linkages within the backbone of the molecule. This concept is inherent in the production of PNAs and can be extended in all of these molecules by the inclusion of nontraditional bases such as inosine, queosine, and wybutosine, as well as acetyl-, methyl-, thio-, and similarly modified forms of adenine, cytidine, guanine, thymine, and uridine which are not as easily recognized by endogenous endonucleases.
Many methods for introducing vectors into cells or tissues are available and equally suitable for use in vivo, in vitro, and ex vivo. For ex vivo therapy, vectors may be introduced into stem cells taken from the patient and clonally propagated for autologous transplant back into that same patient.
Delivery by transfection, by liposome injections, or by polycationic amino polymers may be achieved using methods which are well known in the art. (See, e.g., Goldman, C.K. et al. (1997) Nat.
Biotechno1.15:462-466.) Any of the therapeutic methods described above may be applied to any subject in need of such therapy, including, for example, mammals such as humans, dogs, cats, cows, horses, rabbits, and monkeys.
An additional embodiment of the invention relates to the administration of a pharmaceutical or sterile composition, in conjunction with a pharmaceutically acceptable Garner, for any of the therapeutic effects discussed above. Such pharmaceutical compositions may consist of IMOL, antibodies to IMOL, and mimetics, agonists, antagonists, or inhibitors of IMOL. The compositions may be administered alone or in combination with at least one other agent, such as a stabilizing compound, which may be administered in any sterile, biocompatible pharmaceutical carrier including, but not limited to, saline, buffered saline, dextrose, and water. The compositions may be administered to a patient alone, or in combination with other agents, drugs, or hormones.
The pharmaceutical compositions utilized in this invention may be administered by any number of routes including, but not limited to, oral, intravenous, intramuscular, intra-arterial, intramedullary, intrathecal, intraventricular, transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical, sublingual, or rectal means.

In addition to the active ingredients, these pharmaceutical compositions may contain suitable pharmaceutically-acceptable carnets comprising excipients and auxiliaries which facilitate processing of the active compounds into preparations which can be used pharmaceutically.
Further details on techniques for formulation and administration may be found in the latest edition of Remin on's Pharmaceutical Sciences (Maack Publishing, Easton PA).
Pharmaceutical compositions for oral administration can be formulated using pharmaceutically acceptable carnets well known in the art in dosages suitable for oral administration.
Such carriers enable the pharmaceutical compositions to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions, and the like, for ingestion by the patient.
Pharmaceutical preparations for oral use can be obtained through combining active compounds with solid excipient and processing the resultant mixture of granules (optionally, after grinding) to obtain tablets or dragee cores. Suitable auxiliaries can be added, if desired. Suitable excipients include carbohydrate or protein fillers, such as sugars, including lactose, sucrose, mannitol, and sorbitol; starch from corn, wheat, rice, potato, or other plants;
cellulose, such as methyl cellulose, hydroxypropylmethyl-cellulose, or sodium carboxymethylcellulose; gums, including arabic and tragacanth; and proteins, such as gelatin and collagen. If desired, disintegrating or solubilizing agents may be added, such as the cross-linked polyvinyl pyrrolidone, agar, and alginic acid or a salt thereof, such as sodium alginate.
Dragee cores may be used in conjunction with suitable coatings, such as concentrated sugar solutions, which may also contain gum arabic, talc, polyvinylpyrrolidone, carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures.
Dyestuffs or pigments may be added to the tablets or dragee coatings for product identification or to characterize the quantity of active compound, i.e., dosage.
Pharmaceutical preparations which can be used orally include push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a coating, such as glycerol or sorbitol.
Push-fit capsules can contain active ingredients mixed with fillers or binders, such as lactose or starches, lubricants, such as talc or magnesium stearate, and, optionally, stabilizers. In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid, or liquid polyethylene glycol with or without stabilizers.
Pharmaceutical formulations suitable for parenteral administration may be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hanks' solution, Ringer's solution, or physiologically buffered saline. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Additionally, suspensions of the active compounds may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils, such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate, triglycerides, or liposomes. Non-lipid polycationic amino polymers may also be used for delivery. Optionally, the suspension may also contain suitable stabilizers or agents to increase the solubility of the compounds and allow for the preparation of highly concentrated solutions.
For topical or nasal administration, penetrants appropriate to the particular barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.
The pharmaceutical compositions of the present invention may be manufactured in a manner that is known in the art, e.g., by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping, or lyophilizing processes.
The pharmaceutical composition may be provided as a salt and can be formed with many acids, including but not limited to, hydrochloric, sulfuric, acetic, lactic, tartaric, malic, and succinic acids. Salts tend to be more soluble in aqueous or other protonic solvents than are the corresponding free base forms. In other cases, the preparation may be a lyophilized powder which may contain any or all of the following: 1 mM to 50 mM histidine, 0.1 % to 2% sucrose, and 2%
to 7% mannitol, at a pH range of 4.5 to 5.5, that is combined with buffer prior to use.
After pharmaceutical compositions have been prepared, they can be placed in an appropriate container and labeled for treatment of an indicated condition. For administration of IMOL, such labeling would include amount, frequency, and method of administration.
Pharmaceutical compositions suitable for use in the invention include compositions wherein the active ingredients are contained in an effective amount to achieve the intended purpose. The determination of an effective dose is well within the capability of those skilled in the art.
For any compound, the therapeutically effective dose can be estimated initially either in cell culture assays, e.g., of neoplastic cells, or in animal models such as mice, rats, rabbits, dogs, or pigs.
An animal model may also be used to determine the appropriate concentration range and route of administration. Such information can then be used to determine useful doses and routes for administration in humans.
A therapeutically effective dose refers to that amount of active ingredient, for example IMOL
or fragments thereof, antibodies of IMOL, and agonists, antagonists or inhibitors of IMOL, which ameliorates the symptoms or condition. Therapeutic efficacy and toxicity may be determined by standard pharmaceutical procedures in cell cultures or with experimental animals, such as by calculating the EDSO (the dose therapeutically effective in 50% of the population) or LDSO (the dose lethal to 50% of the population) statistics. The dose ratio of toxic to therapeutic effects is the therapeutic index, which can be expressed as the LDSO/EDSO ratio.
Pharmaceutical compositions which exhibit large therapeutic indices are preferred. The data obtained from cell culture assays and animal studies are used to formulate a range of dosage for human use. The dosage contained in such compositions is preferably within a range of circulating concentrations that includes the EDSO with little or no toxicity. The dosage varies within this range depending upon the dosage form employed, the sensitivity of the patient, and the route of administration.
The exact dosage will be determined by the practitioner, in light of factors related to the subject requiring treatment. Dosage and administration are adjusted to provide sufficient levels of the active moiety or to maintain the desired effect. Factors which may be taken into account include the severity of the disease state, the general health of the subject, the age, weight, and gender of the subject, time and frequency of administration, drug combination(s), reaction sensitivities, and response to therapy. Long-acting pharmaceutical compositions may be administered every 3 to 4 days, every week, or biweekly depending on the half life and clearance rate of the particular formulation.
Normal dosage amounts may vary from about 0.1 ~g to 100,000 fig, up to a total dose of about 1 gram, depending upon the route of administration. Guidance as to particular dosages and methods of delivery is provided in the literature and generally available to practitioners in the art.
Those skilled in the art will employ different formulations for nucleotides than for proteins or their inhibitors. Similarly, delivery of polynucleotides or polypeptides will be specific to particular cells, conditions, locations, etc.
DIAGNOSTICS
In another embodiment, antibodies which specifically bind IMOL may be used for the diagnosis of disorders characterized by expression of IMOL, or in assays to monitor patients being treated with IMOL or agonists, antagonists, or inhibitors of IMOL. Antibodies useful for diagnostic purposes may be prepared in the same manner as described above for therapeutics. Diagnostic assays for IMOL include methods which utilize the antibody and a label to detect IMOL
in human body fluids or in extracts of cells or tissues. The antibodies may be used with or without modification, and may be labeled by covalent or non-covalent attachment of a reporter molecule.
A wide variety of reporter molecules, several of which are described above, are known in the art and may be used.
A variety of protocols for measuring IMOL, including ELISAs, RIAs, and FACS, are known in the art and provide a basis for diagnosing altered or abnormal levels of IMOL expression. Normal or standard values for IMOL expression are established by combining body fluids or cell extracts taken from normal mammalian subjects, for example, human subjects, with antibody to IMOL under conditions suitable for complex formation. The amount of standard complex formation may be quantitated by various methods, such as photometric means. Quantities of IMOL
expressed in subject, control, and disease samples from biopsied tissues are compared with the standard values.
Deviation between standard and subject values establishes the parameters for diagnosing disease.
In another embodiment of the invention, the polynucleotides encoding IMOL may be used for diagnostic purposes. The polynucleotides which may be used include oligonucleotide sequences, complementary RNA and DNA molecules, and PNAs. The polynucleotides may be used to detect and quantify gene expression in biopsied tissues in which expression of IMOL
may be correlated with disease. The diagnostic assay may be used to determine absence, presence, and excess expression of IMOL, and to monitor regulation of IMOL levels during therapeutic intervention.
In one aspect, hybridization with PCR probes which are capable of detecting polynucleotide sequences, including genomic sequences, encoding IMOL or closely related molecules may be used to identify nucleic acid sequences which encode IMOL. The specificity of the probe, whether it is made from a highly specific region, e.g., the 5' regulatory region, or from a less specific region, e.g., a conserved motif, and the stringency of the hybridization or amplification will determine whether the probe identifies only naturally occurring sequences encoding IMOL, allelic variants, or related sequences.
Probes may also be used for the detection of related sequences, and may have at least 50%
sequence identity to any of the IMOL encoding sequences. The hybridization probes of the subject invention may be DNA or RNA and may be derived from the sequence of SEQ ID
N0:16-30 or from genomic sequences including promoters, enhancers, and introns of the IMOL
gene.
Means for producing specific hybridization probes for DNAs encoding IMOL
include the cloning of polynucleotide sequences encoding IMOL or IMOL derivatives into vectors for the production of mRNA probes. Such vectors are known in the art, are commercially available, and may be used to synthesize RNA probes in vitro by means of the addition of the appropriate RNA
polymerases and the appropriate labeled nucleotides. Hybridization probes may be labeled by a variety of reporter groups, for example, by radionuclides such as 32P or 355, or by enzymatic labels, such as alkaline phosphatase coupled to the probe via avidin/biotin coupling systems, and the like.
Polynucleotide sequences encoding IMOL may be used for the diagnosis of disorders associated with expression of IMOL. Examples of such disorders include, but are not limited to, an immunological disorder such as inflammation, actinic keratosis, acquired immunodeficiency syndrome (AIDS), Addison's disease, adult respiratory distress syndrome, allergies, ankylosing spondylitis, amyloidosis, anemia, arteriosclerosis, asthma, atherosclerosis, autoimmune hemolytic anemia, autoimmune thyroiditis, autoimune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED), bronchitis, bursitis, cholecystitis, cirrhosis, contact dermatitis, Crohn's disease, atopic dermatitis, dermatomyositis, diabetes mellitus, emphysema, erythroblastosis fetalis, erythema nodosum, atrophic gastritis, glomerulonephritis, Goodpasture's syndrome, gout, Graves' disease, Hashimoto's thyroiditis, paroxysmal nocturnal hemoglobinuria, hepatitis, hypereosinophilia, irritable bowel syndrome, episodic lymphopenia with lymphocytotoxins, mixed connective tissue disease (MCTD), multiple sclerosis, myasthenia gravis, myocardial or pericardial inflammation, myelofibrosis, osteoarthritis, osteoporosis, pancreatitis, polycythemia vera, polymyositis, psoriasis, Reiter's syndrome, rheumatoid arthritis, scleroderma, Sjogren's syndrome, systemic anaphylaxis, systemic lupus erythematosus, systemic sclerosis, primary thrombocythemia, thrombocytopenic purpura, ulcerative colitis, uveitis, Werner syndrome, complications of cancer, hemodialysis, and extracorporeal circulation, trauma, and hematopoietic cancer including lymphoma, leukemia, and myeloma; and an infection caused by a viral agent classified as adenovirus, arenavirus, bunyavirus, calicivirus, coronavirus, filovirus, hepadnavirus, herpesvirus, flavivirus, orthomyxovirus, parvovirus, papovavirus, paramyxovirus, picornavirus, poxvirus, reovirus, retrovirus, rhabdovirus, or togavirus;
an infection caused by a bacterial agent classified as pneumococcus, staphylococcus, streptococcus, bacillus, corynebacterium, clostridium, meningococcus, gonococcus, listeria, moraxella, kingella, haemophilus, legionella, bordetella, gram-negative enterobacterium including shigella, salmonella, or campylobacter, pseudomonas, vibrio, brucella, francisella, yersinia, bartonella, norcardium, actinomyces, mycobacterium, spirochaetale, rickettsia, chlamydia, or mycoplasma; an infection caused by a fungal agent classified as aspergillus, blastomyces, dermatophytes, cryptococcus, coccidioides, malasezzia, histoplasma, or other mycosis-causing fungal agent;
and an infection caused by a parasite classified as plasmodium or malaria-causing, parasitic entamoeba, leishmania, trypanosoma, toxoplasma, pneumocystis carinii, intestinal protozoa such as giardia, trichomonas, tissue nematode such as trichinella, intestinal nematode such as ascaris, lymphatic filarial nematode, trematode such as schistosoma, and cestrode such as tapeworm; and a cell proliferative disorder such as actinic keratosis, arteriosclerosis, atherosclerosis, bursitis, cirrhosis, hepatitis, mixed connective tissue disease (MCTD), myelofibrosis, paroxysmal nocturnal hemoglobinuria, polycythemia vera, psoriasis, primary thrombocythemia, and cancers including adenocarcinoma, leukemia, lymphoma, melanoma, myeloma, sarcoma, teratocarcinoma, and, in particular, cancers of the adrenal gland, bladder, bone, bone marrow, brain, breast, cervix, gall bladder, ganglia, gastrointestinal tract, heart, kidney, liver, lung, muscle, ovary, pancreas, parathyroid, penis, prostate, salivary glands, skin, spleen, testis, thymus, thyroid, and uterus. The polynucleotide sequences encoding IMOL may be used in Southern or northern analysis, dot blot, or other membrane-based technologies;
in PCR technologies;
in dipstick, pin, and multiformat ELISA-like assays; and in microarrays utilizing fluids or tissues from patients to detect altered IMOL expression. Such qualitative or quantitative methods are well known in the art.
In a particular aspect, the nucleotide sequences encoding IMOL may be useful in assays that detect the presence of associated disorders, particularly those mentioned above. The nucleotide sequences encoding IMOL may be labeled by standard methods and added to a fluid or tissue sample from a patient under conditions suitable for the formation of hybridization complexes. After a suitable incubation period, the sample is washed and the signal is quantified and compared with a standard value. If the amount of signal in the patient sample is significantly altered in comparison to a control sample then the presence of altered levels of nucleotide sequences encoding IMOL in the sample indicates the presence of the associated disorder. Such assays may also be used to evaluate the efficacy of a particular therapeutic treatment regimen in animal studies, in clinical trials, or to monitor the treatment of an individual patient.
In order to provide a basis for the diagnosis of a disorder associated with expression of IMOL, a normal or standard profile for expression is established. This may be accomplished by combining body fluids or cell extracts taken from normal subjects, either animal or human, with a sequence, or a fragment thereof, encoding IMOL, under conditions suitable for hybridization or amplification. Standard hybridization may be quantified by comparing the values obtained from normal subjects with values from an experiment in which a known amount of a substantially purified polynucleotide is used. Standard values obtained in this manner may be compared with values obtained from samples from patients who are symptomatic for a disorder.
Deviation from standard values is used to establish the presence of a disorder.
Once the presence of a disorder is established and a treatment protocol is initiated, hybridization assays may be repeated on a regular basis to determine if the level of expression in the patient begins to approximate that which is observed in the normal subject.
The results obtained from successive assays may be used to show the efficacy of treatment over a period ranging from several days to months.
With respect to cancer, the presence of an abnormal amount of transcript (either under- or overexpressed) in biopsied tissue from an individual may indicate a predisposition for the development of the disease, or may provide a means for detecting the disease prior to the appearance of actual clinical symptoms. A more definitive diagnosis of this type may allow health professionals to employ preventative measures or aggressive treatment earlier thereby preventing the development or further progression of the cancer.
Additional diagnostic uses for oligonucleotides designed from the sequences encoding IMOL
may involve the use of PCR. These oligomers may be chemically synthesized, generated enzymatically, or produced in vitro. Oligomers will preferably contain a fragment of a polynucleotide encoding IMOL, or a fragment of a polynucleotide complementary to the polynucleotide encoding IMOL, and will be employed under optimized conditions for identification of a specific gene or condition. Oligomers may also be employed under less stringent conditions for detection or quantification of closely related DNA or RNA sequences.
Methods which may also be used to quantify the expression of IMOL include radiolabeling or biotinylating nucleotides, coamplification of a control nucleic acid, and interpolating results from standard curves. (See, e.g., Melby, P.C. et al. (1993) J. Immunol. Methods 159:235-244; Duplaa, C.

et al. (1993) Anal. Biochem. 212:229-236.) The speed of quantitation of multiple samples may be accelerated by running the assay in a high-throughput format where the oligomer of interest is presented in various dilutions and a spectrophotometric or colorimetric response gives rapid quantitation.
In further embodiments, oligonucleotides or longer fragments derived from any of the polynucleotide sequences described herein may be used as targets in a microarray. The microarray can be used to monitor the expression level of large numbers of genes simultaneously and to identify genetic variants, mutations, and polymorphisms. This information may be used to determine gene function, to understand the genetic basis of a disorder, to diagnose a disorder, and to develop and monitor the activities of therapeutic agents.
Microarrays may be prepared, used, and analyzed using methods known in the art. (See, e.g., Brennan, T.M. et al. (1995) U.S. Patent No. 5,474,796; Schena, M. et al.
(1996) Proc. Natl. Acad. Sci.
USA 93:10614-10619; Baldeschweiler et al. ( 1995) PCT application W095/251116;
Shalon, D. et al.
(1995) PCT application W095/35505; Heller, R.A. et al. (1997) Proc. Natl.
Acad. Sci. USA 94:2150-2155; and Heller, M.J. et al. (1997) U.S. Patent No. 5,605,662.) In another embodiment of the invention, nucleic acid sequences encoding IMOL
may be used to generate hybridization probes useful in mapping the naturally occurring genomic sequence. The sequences may be mapped to a particular chromosome, to a specific region of a chromosome, or to artificial chromosome constructions, e.g., human artificial chromosomes (HACs), yeast artificial chromosomes (YACs), bacterial artificial chromosomes (BACs), bacterial P1 constructions, or single chromosome cDNA libraries. (See, e.g., Harrington, J.J. et al. (1997) Nat.
Genet. 15:345-355; Price, C.M. (1993) Blood Rev. 7:127-134; and Trask, B.J. (1991) Trends Genet. 7:149-154.) Fluorescent in situ hybridization (FISH) may be correlated with other physical chromosome mapping techniques and genetic map data. (See, e.g., Heinz-Ulrich, et al.
(1995) in Meyers, su~a, pp. 965-968.) Examples of genetic map data can be found in various scientific journals or at the Online Mendelian Inheritance in Man (OMIM) World Wide Web site. Correlation between the location of the gene encoding IMOL on a physical chromosomal map and a specific disorder, or a predisposition to a specific disorder, may help define the region of DNA
associated with that disorder. The nucleotide sequences of the invention may be used to detect differences in gene sequences among normal, carrier, and affected individuals.
In situ hybridization of chromosomal preparations and physical mapping techniques, such as linkage analysis using established chromosomal markers, may be used for extending genetic maps.
Often the placement of a gene on the chromosome of another mammalian species, such as mouse, may reveal associated markers even if the number or arm of a particular human chromosome is not known. New sequences can be assigned to chromosomal arms by physical mapping.
This provides valuable information to investigators searching for disease genes using positional cloning or other gene discovery techniques. Once the disease or syndrome has been crudely localized by genetic linkage to a particular genomic region, e.g., ataxia-telangiectasia to l 1q22-23, any sequences mapping to that area may represent associated or regulatory genes for further investigation. (See, e.g., Gatti, R.A. et al. (1988) Nature 336:577-580.) The nucleotide sequence of the subject invention may also be used to detect differences in the chromosomal location due to translocation, inversion, etc., among normal, carrier, or affected individuals.
In another embodiment of the invention, IMOL, its catalytic or immunogenic fragments, or oligopeptides thereof can be used for screening libraries of compounds in any of a variety of drug screening techniques. The fragment employed in such screening may be free in solution, affixed to a solid support, borne on a cell surface, or located intracellularly. The formation of binding complexes between IMOL and the agent being tested may be measured.
Another technique for drug screening provides for high throughput screening of compounds having suitable binding affinity to the protein of interest. (See, e.g., Geysen, et al. ( 1984) PCT
application W084/03564.) In this method, large numbers of different small test compounds are synthesized on a solid substrate. The test compounds are reacted with IMOL, or fragments thereof, and washed. Bound IMOL is then detected by methods well known in the art.
Purified IMOL can also be coated directly onto plates for use in the aforementioned drug screening techniques.
Alternatively, non-neutralizing antibodies can be used to capture the peptide and immobilize it on a solid support.
In another embodiment, one may use competitive drug screening assays in which neutralizing antibodies capable of binding IMOL specifically compete with a test compound for binding IMOL.
In this manner, antibodies can be used to detect the presence of any peptide which shares one or more antigenic determinants with IMOL.
In additional embodiments, the nucleotide sequences which encode IMOL may be used in any molecular biology techniques that have yet to be developed, provided the new techniques rely on properties of nucleotide sequences that are currently known, including, but not limited to, such properties as the triplet genetic code and specific base pair interactions.
Without further elaboration, it is believed that one skilled in the art can, using the preceding description, utilize the present invention to its fullest extent. The following preferred specific embodiments are, therefore, to be construed as merely illustrative, and not limitative of the remainder of the disclosure in any way whatsoever.
The disclosures of all patents, applications, and publications mentioned above and below, in particular U.S. Ser. No. 60/127,852 and U.S. Ser. No. 60/132,647, are hereby expressly incorporated by reference.

EXAMPLES
I. Construction of cDNA Libraries RNA was purchased from Clontech or isolated from tissues described in Table 4.
Some tissues were homogenized and lysed in guanidinium isothiocyanate, while others were homogenized and lysed in phenol or in a suitable mixture of denaturants, such as TRIZOL
(Life Technologies), a monophasic solution of phenol and guanidine isothiocyanate. The resulting lysates were centrifuged over CsCI cushions or extracted with chloroform. RNA was precipitated from the lysates with either isopropanol or sodium acetate and ethanol, or by other routine methods.
Phenol extraction and precipitation of RNA were repeated as necessary to increase RNA
purity. In some cases, RNA was treated with DNase. For most libraries, poly(A+) RNA was isolated using oligo d(T)-coupled paramagnetic particles (Promega), OLIGOTEX latex particles (QIAGEN, Chatsworth CA), or an OLIGOTEX mRNA purification kit (QIAGEN). Alternatively, RNA was isolated directly from tissue lysates using other RNA isolation kits, e.g., the POLY(A)PURE mRNA
purification kit (Ambion, Austin TX).
In some cases, Stratagene was provided with RNA and constructed the corresponding cDNA
libraries. Otherwise, cDNA was synthesized and cDNA libraries were constructed with the UNIZAP
vector system (Stratagene) or SUPERSCRIPT plasmid system (Life Technologies), using the recommended procedures or similar methods known in the art. (See, e.g., Ausubel, 1997, supra, units 5.1-6.6.) Reverse transcription was initiated using oligo d(T) or random primers. Synthetic oligonucleotide adapters were ligated to double stranded cDNA, and the cDNA
was digested with the appropriate restriction enzyme or enzymes. For most libraries, the cDNA was size-selected (300-1000 bp) using SEPHACRYL S 1000, SEPHAROSE CL2B, or SEPHAROSE CL4B column chromatography (Amersham Pharmacia Biotech) or preparative agarose gel electrophoresis. cDNAs were ligated into compatible restriction enzyme sites of the polylinker of a suitable plasmid, e.g., PBLUESCR1PT plasmid (Stratagene), PSPORT1 plasmid (Life Technologies), pcDNA2.l plasmid (Invitrogen, Carlsbad CA), or pINCY plasmid (Incyte Pharmaceuticals, Palo Alto CA). Recombinant plasmids were transformed into competent E. coli cells including XL1-Blue, XL1-BIueMRF, or SOLR from Stratagene or DHSa, DH10B, or ElectroMAX DH10B from Life Technologies.
II. Isolation of cDNA Clones Plasmids were recovered from host cells by in vivo excision using the UNIZAP
vector system (Stratagene) or by cell lysis. Plasmids were purified using at least one of the following: a Magic or WIZARD Minipreps DNA purification system (Promega); an AGTC Miniprep purification kit (Edge Biosystems, Gaithersburg MD); and QIAWELL 8 Plasmid, QIAWELL 8 Plus Plasmid, Ultra Plasmid purification systems or the R.E.A.L. PREP 96 plasmid purification kit from QIAGEN.
Following precipitation, plasmids were resuspended in 0.1 ml of distilled water and stored, with or without lyophilization, at 4°C.
Alternatively, plasmid DNA was amplified from host cell lysates using direct link PCR in a high-throughput format (Rao, V.B. (1994) Anal. Biochem. 216:1-14). Host cell lysis and thermal cycling steps were carned out in a single reaction mixture. Samples were processed and stored in 384-well plates, and the concentration of amplified plasmid DNA was quantified fluorometrically using PICOGREEN dye (Molecular Probes, Eugene OR) and a FLUOROSKAN II
fluorescence scanner (Labsystems Oy, Helsinki, Finland).
III. Sequencing and Analysis cDNA sequencing reactions were processed using standard methods or high-throughput instrumentation such as the ABI CATALYST 800 (Perkin-Elmer) thermal cycler or the PTC-200 thermal cycler (MJ Research) in conjunction with the HYDRA microdispenser (Robbins Scientific) or the MICROLAB 2200 (Hamilton) liquid transfer system. cDNA sequencing reactions were prepared using reagents provided by Amersham Pharmacia Biotech or supplied in ABI sequencing kits such as the ABI PRISM BIGDYE Terminator cycle sequencing ready reaction kit (Perkin-Elmer).
Electrophoretic separation of cDNA sequencing reactions and detection of labeled polynucleotides were carned out using the MEGABACE 1000 DNA sequencing system (Molecular Dynamics); the ABI PRISM 373 or 377 sequencing system (Perkin-Elmer) in conjunction with standard ABI
protocols and base calling software; or other sequence analysis systems known in the art. Reading frames within the cDNA sequences were identified using standard methods (reviewed in Ausubel, 1997, supra, unit 7.7). Some of the cDNA sequences were selected for extension using the techniques disclosed in Example VI.
The polynucleotide sequences derived from cDNA sequencing were assembled and analyzed using a combination of software programs which utilize algorithms well known to those skilled in the art. Table 5 summarizes the tools, programs, and algorithms used and provides applicable descriptions, references, and threshold parameters. The first column of Table 5 shows the tools, programs, and algorithms used, the second column provides brief descriptions thereof, the third column presents appropriate references, all of which are incorporated by reference herein in their entirety, and the fourth column presents, where applicable, the scores, probability values, and other parameters used to evaluate the strength of a match between two sequences (the higher the score, the greater the homology between two sequences). Sequences were analyzed using MACDNASIS PRO
software (Hitachi Software Engineering, South San Francisco CA) and LASERGENE
software (DNASTAR). Polynucleotide and polypeptide sequence alignments were generated using the default parameters specified by the clustal algorithm as incorporated into the MEGALIGN multisequence alignment program (DNASTAR), which also calculates the percent identity between aligned sequences.

The polynucleotide sequences were validated by removing vector, linker, and polyA
sequences and by masking ambiguous bases, using algorithms and programs based on BLAST, dynamic programing, and dinucleotide nearest neighbor analysis. The sequences were then queried against a selection of public databases such as the GenBank primate, rodent, mammalian, vertebrate, and eukaryote databases, and BLOCKS, PRINTS, DOMO, PRODOM, and PFAM to acquire annotation using programs based on BLAST, FASTA, and BLIMPS. The sequences were assembled into full length polynucleotide sequences using programs based on Phred, Phrap, and Consed, and were screened for open reading frames using programs based on GeneMark, BLAST, and FASTA.
The full length polynucleotide sequences were translated to derive the corresponding full length amino acid sequences, and these full length sequences were subsequently analyzed by querying against databases such as the GenBank databases (described above), SwissProt, BLOCKS, PRINTS, DOMO, PRODOM, Prosite, and Hidden Markov Model (HMM)-based protein family databases such as PFAM. HMM is a probabilistic approach which analyzes consensus primary structures of gene families. (See, e.g., Eddy, S.R. (1996) Curr. Opin. Struct. Biol. 6:361-365.) The programs described above for the assembly and analysis of full length polynucleotide and amino acid sequences were also used to identify polynucleotide sequence fragments from SEQ ID
N0:16-30. Fragments from about 20 to about 4000 nucleotides which are useful in hybridization and amplification technologies were described in The Invention section above.
IV. Northern Analysis Northern analysis is a laboratory technique used to detect the presence of a transcript of a gene and involves the hybridization of a labeled nucleotide sequence to a membrane on which RNAs from a particular cell type or tissue have been bound. (See, e.g., Sambrook, supra, ch. 7; Ausubel, 1995, supra, ch. 4 and 16.) Analogous computer techniques applying BLAST were used to search for identical or related molecules in nucleotide databases such as GenBank or LIFESEQ (Incyte Pharmaceuticals). This analysis is much faster than multiple membrane-based hybridizations. In addition, the sensitivity of the computer search can be modified to determine whether any particular match is categorized as exact or similar. The basis of the search is the product score, which is defined as:
% seguence identity.x % maximum BLAST score The product score takes into account both the degree of similarity between two sequences and the length of the sequence match. For example, with a product score of 40, the match will be exact within a 1% to 2% error, and, with a product score of 70, the match will be exact. Similar molecules are usually identified by selecting those which show product scores between 15 and 40, although lower scores may identify related molecules.
The results of northern analyses are reported as a percentage distribution of libraries in which the transcript encoding IMOL occurred. Analysis involved the categorization of cDNA libraries by organ/tissue and disease. The organ/tissue categories included cardiovascular, dermatologic, developmental, endocrine, gastrointestinal, hematopoietic/immune, musculoskeletal, nervous, reproductive, and urologic. The disease/condition categories included cancer, inflammation, trauma, cell proliferation, neurological, and pooled. For each category, the number of libraries expressing the sequence of interest was counted and divided by the total number of libraries across all categories.
Percentage values of tissue-specific and disease- or condition-specific expression are reported in Table 3.
V. Chromosomal Mapping of IMOL Encoding Polynucleotides The cDNA sequences which were used to assemble SEQ ID N0:16-30 were compared with sequences from the Incyte LIFESEQ database and public domain databases using BLAST and other implementations of the Smith-Waterman algorithm. Sequences from these databases that matched SEQ ID N0:16-30 were assembled into clusters of contiguous and overlapping sequences using assembly algorithms such as Phrap (Table 5). Radiation hybrid and genetic mapping data available from public resources such as the Stanford Human Genome Center (SHGC), Whitehead Institute for Genome Research (WIGR), and Genethon were used to determine if any of the clustered sequences had been previously mapped. Inclusion of a mapped sequence in a cluster resulted in the assignment of all sequences of that cluster, including its particular SEQ ID NO:, to that map location. The genetic map locations of SEQ ID N0:26 and SEQ ID N0:29 are described in the Invention as a range, or interval, of a particular human chromosome. The map position of an interval, in centiMorgans, is measured relative to the terminus of the chromosome's p-arm.
(The centiMorgan (cM) is a unit of measurement based on recombination frequencies between chromosomal markers.
On average, 1 cM is roughly equivalent to 1 megabase (Mb) of DNA in humans, although this can vary widely due to hot and cold spots of recombination.) The cM distances are based on genetic markers mapped by Genethon which provide boundaries for radiation hybrid markers whose sequences were included in each of the clusters. Diseases associated with the public and Incyte sequences located within the indicated intervals are also reported in the Invention.
VI. Extension of IMOL Encoding Polynucleotides The full length nucleic acid sequences of SEQ ID N0:16-30 were produced by extension of an appropriate fragment of the full length molecule using oligonucleotide primers designed from this fragment. One primer was synthesized to initiate 5' extension of the known fragment, and the other primer, to initiate 3' extension of the known fragment. The initial primers were designed using OLIGO 4.06 software (National Biosciences), or another appropriate program, to be about 22 to 30 nucleotides in length, to have a GC content of about 50% or more, and to anneal to the target sequence at temperatures of about 68°C to about 72°C. Any stretch of nucleotides which would result in hairpin structures and primer-primer dimerizations was avoided.
Selected human cDNA libraries were used to extend the sequence. If more than one extension was necessary or desired, additional or nested sets of primers were designed.
High fidelity amplification was obtained by PCR using methods well known in the art. PCR
was performed in 96-well plates using the PTC-200 thermal cycler (MJ Research, Inc.). The reaction mix contained DNA template, 200 nmol of each primer, reaction buffer containing Mg'-+, (NH4)ZS04, and ~i-mercaptoethanol, Taq DNA polymerase (Amersham Pharmacia Biotech), ELONGASE enzyme (Life Technologies), and Pfu DNA polymerase (Stratagene), with the following parameters for primer pair PCI A and PCI B: Step 1: 94°C, 3 min; Step 2: 94°C, 15 sec;
Step 3: 60°C, 1 min; Step 4: 68°C, 2 min; Step 5: Steps 2, 3, and 4 repeated 20 times; Step 6: 68°C, 5 min; Step 7: storage at 4°C. In the alternative, the parameters for primer pair T7 and SK+ were as follows: Step 1: 94°C, 3 min; Step 2:
94°C, 15 sec; Step 3: 57°C, 1 min; Step 4: 68°C, 2 min;
Step 5: Steps 2, 3, and 4 repeated 20 times;
Step 6: 68°C, 5 min; Step 7: storage at 4°C.
The concentration of DNA in each well was determined by dispensing 100 pl PICOGREEN
quantitation reagent (0.25% (v/v) PICOGREEN; Molecular Probes, Eugene OR) dissolved in 1X TE
and 0.5 pl of undiluted PCR product into each well of an opaque fluorimeter plate (Corning Costar, Acton MA), allowing the DNA to bind to the reagent. The plate was scanned in a Fluoroskan II
(Labsystems Oy, Helsinki, Finland) to measure the fluorescence of the sample and to quantify the concentration of DNA. A 5 ~l to 10 ~1 aliquot of the reaction mixture was analyzed by electrophoresis on a 1 % agarose mini-gel to determine which reactions were successful in extending the sequence.
The extended nucleotides were desalted and concentrated, transferred to 384-well plates, digested with CviJI cholera virus endonuclease (Molecular Biology Research, Madison Wn, and sonicated or sheared prior to religation into pUC 18 vector (Amersham Pharmacia Biotech). For shotgun sequencing, the digested nucleotides were separated on low concentration (0.6 to 0.8%) agarose gels, fragments were excised, and agar digested with Agar ACE
(Promega). Extended clones were religated using T4 ligase (New England Biolabs, Beverly MA) into pUC 18 vector (Amersham Pharmacia Biotech), treated with Pfu DNA polymerase (Stratagene) to fill-in restriction site overhangs, and transfected into competent E. coli cells. Transformed cells were selected on antibiotic-containing media, individual colonies were picked and cultured overnight at 37°C in 384-well plates in LB/2x carb liquid media.
The cells were lysed, and DNA was amplified by PCR using Taq DNA polymerase (Amersham Pharmacia Biotech) and Pfu DNA polymerase (Stratagene) with the following parameters: Step 1: 94°C, 3 min; Step 2: 94°C, 15 sec; Step 3:
60°C, 1 min; Step 4: 72°C, 2 min;
Step 5: steps 2, 3, and 4 repeated 29 times; Step 6: 72°C, 5 min; Step 7: storage at 4°C. DNA was quantified by PICOGREEN reagent (Molecular Probes) as described above. Samples with low DNA
recoveries were reamplified using the same conditions as described above.
Samples were diluted with 20% dimethysulfoxide ( 1:2, v/v), and sequenced using DYENAMIC energy transfer sequencing primers and the DYENAMIC DIRECT kit (Amersham Pharmacia Biotech) or the ABI
PRISM
BIGDYE Terminator cycle sequencing ready reaction kit (Perkin-Elmer).
In like manner, the nucleotide sequences of SEQ m N0:16-30 are used to obtain 5' regulatory sequences using the procedure above, oligonucleotides designed for such extension, and an appropriate genomic library.
VII. Labeling and Use of Individual Hybridization Probes Hybridization probes derived from SEQ ID N0:16-30 are employed to screen cDNAs, genomic DNAs, or mRNAs. Although the labeling of oligonucleotides, consisting of about 20 base pairs, is specifically described, essentially the same procedure is used with larger nucleotide fragments. Oligonucleotides are designed using state-of the-art software such as OLIGO 4.06 software (National Biosciences) and labeled by combining 50 pmol of each oligomer, 250 ~cCi of [Y szP] adenosine triphosphate (Amersham Pharmacia Biotech), and T4 polynucleotide kinase (DuPont NEN, Boston MA). The labeled oligonucleotides are substantially purified using a SEPHADEX G-25 superf'me size exclusion dextran bead column (Amersham Pharmacia Biotech).
An aliquot containing 10' counts per minute of the labeled probe is used in a typical membrane-based hybridization analysis of human genomic DNA digested with one of the following endonucleases:
Ase I, Bgl II, Eco RI, Pst I, Xba I, or Pvu II (DuPont NEN).
The DNA from each digest is fractionated on a 0.7% agarose gel and transferred to nylon membranes (Nytran Plus, Schleicher & Schuell, Durham NH). Hybridization is carried out for 16 hours at 40°C. To remove nonspecific signals, blots are sequentially washed at room temperature under conditions of up to, for example, 0.1 x saline sodium citrate and 0.5%
sodium dodecyl sulfate.
Hybridization patterns are visualized using autoradiography or an alternative imaging means and compared.
VIII. Microarrays A chemical coupling procedure and an ink jet device can be used to synthesize array elements on the surface of a substrate. (See, e.g., Baldeschweiler, supra.) An array analogous to a dot or slot blot may also be used to arrange and link elements to the surface of a substrate using thermal, W, chemical, or mechanical bonding procedures. A typical array may be produced by hand or using available methods and machines and contain any appropriate number of elements. After hybridization, nonhybridized probes are removed and a scanner used to determine the levels and patterns of fluorescence. The degree of complementarity and the relative abundance of each probe which hybridizes to an element on the microarray may be assessed through analysis of the scanned images.
Full-length cDNAs, Expressed Sequence Tags (ESTs), or fragments thereof may comprise the elements of the microarray. Fragments suitable for hybridization can be selected using software well known in the art such as LASERGENE software (DNASTAR). Full-length cDNAs, ESTs, or fragments thereof corresponding to one of the nucleotide sequences of the present invention, or selected at random from a cDNA library relevant to the present invention, are arranged on an appropriate substrate, e.g., a glass slide. The cDNA is fixed to the slide using, e.g., UV cross-linking followed by thermal and chemical treatments and subsequent drying. (See, e.g., Schena, M. et al.
( 1995) Science 270:467-470; Shalon, D. et al. ( 1996) Genome Res. 6:639-645.) Fluorescent probes are prepared and used for hybridization to the elements on the substrate. The substrate is analyzed by procedures described above.
IX. Complementary Polynucleotides Sequences complementary to the IMOL-encoding sequences, or any parts thereof, are used to detect, decrease, or inhibit expression of naturally occurring IMOL.
Although use of oligonucleotides comprising from about 15 to 30 base pairs is described, essentially the same procedure is used with smaller or with larger sequence fragments. Appropriate oligonucleotides are designed using OLIGO 4.06 software (National Biosciences) and the coding sequence of IMOL. To inhibit transcription, a complementary oligonucleotide is designed from the most unique 5' sequence and used to prevent promoter binding to the coding sequence. To inhibit translation, a complementary oligonucleotide is designed to prevent ribosomal binding to the IMOL-encoding transcript.
X. Expression of IMOL
Expression and purification of IMOL is achieved using bacterial or virus-based expression systems. For expression of IMOL in bacteria, cDNA is subcloned into an appropriate vector containing an antibiotic resistance gene and an inducible promoter that directs high levels of cDNA
transcription. Examples of such promoters include, but are not limited to, the trp-lac (tac) hybrid promoter and the TS or T7 bacteriophage promoter in conjunction with the lac operator regulatory element. Recombinant vectors are transformed into suitable bacterial hosts, e.g., BL21(DE3).
Antibiotic resistant bacteria express IMOL upon induction with isopropyl beta-D-thiogalactopyranoside (IPTG). Expression of IMOL in eukaryotic cells is achieved by infecting insect or mammalian cell lines with recombinant Autog-phica californica nuclear polyhedrosis virus (AcMNPV), commonly known as baculovirus. The nonessential polyhedrin gene of baculovirus is replaced with cDNA encoding IMOL by either homologous recombination or bacterial-mediated transposition involving transfer plasmid intermediates. Viral infectivity is maintained and the strong polyhedrin promoter drives high levels of cDNA transcription. Recombinant baculovirus is used to infect Spodoptera frugiperda (Sf9) insect cells in most cases, or human hepatocytes, in some cases.
Infection of the latter requires additional genetic modifications to baculovirus. (See Engelhard, E.K.
et al. (1994) Proc. Natl. Acad. Sci. USA 91:3224-3227; Sandig, V. et al.
(1996) Hum. Gene Ther.
7:1937-1945.) In most expression systems, IMOL is synthesized as a fusion protein with, e.g., glutathione S-transferase (GST) or a peptide epitope tag, such as FLAG or 6-His, permitting rapid, single-step, affinity-based purification of recombinant fusion protein from crude cell lysates. GST, a 26-kilodalton enzyme from Schistosoma janonicum, enables the purification of fusion proteins on immobilized glutathione under conditions that maintain protein activity and antigenicity (Amersham Pharmacia Biotech). Following purification, the GST moiety can be proteolytically cleaved from IMOL at specifically engineered sites. FLAG, an 8-amino acid peptide, enables immunoaffinity purification using commercially available monoclonal and polyclonal anti-FLAG
antibodies (Eastman Kodak). 6-His, a stretch of six consecutive histidine residues, enables purification on metal-chelate resins (QIAGEN). Methods for protein expression and purification are discussed in Ausubel (1995, supra, ch. 10 and 16). Purified IMOL obtained by these methods can be used directly in the following activity assay.
XI. Demonstration of IMOL Activity An assay for IMOL activity measures the proliferation of leukocytes in response to IMOL.
In this assay, the amount of tritiated thymidine incorporated into newly synthesized DNA is used to estimate proliferative activity. Varying amounts of IMOL are added to cultured leukocytes, such as granulocytes, monocytes, or lymphocytes, in the presence of [3H]thymidine, a radioactive DNA
precursor. IMOL for this assay can be obtained by recombinant means or from biochemical preparations. Incorporation of [3H]thymidine into acid-precipitable DNA is measured over an appropriate time interval, and the amount incorporated is directly proportional to the amount of newly synthesized DNA. A linear dose-response curve over at least a hundred-fold IMOL concentration range is indicative of IMOL activity. One unit of activity per milliliter is conventionally defined as the concentration of IMOL producing a SO% response level, where 100%
represents maximal incorporation of [3H]thymidine into acid-precipitable DNA.
Alternatively, an assay for IMOL activity measures the expression of IMOL on the cell surface. cDNA encoding IMOL is transfected into a non-leukocytic cell line.
Cell surface proteins are labeled with biotin as described (de la Fuente, M. A. et al. (1997) Blood 90:2398-2405).
Immunoprecipitations are performed using IMOL-specific antibodies, and immunoprecipitated samples are analyzed using SDS-PAGE and immunoblotting techniques. The ratio of labeled immunoprecipitant to unlabeled immunoprecipitant is proportional to the amount of IMOL expressed on the cell surface.
Alternatively, IMOL activity is exemplified by that of immunoglobulins, which recognize and precipitate antigens from serum. The quantitative precipitin reaction measures this activity.
(Golub, E. S. et al. ( 1987) Immunology: A Synthesis, Sinauer Associates, Sunderland, MA, pages 113-I 15.) IMOL is isotopically labeled using methods known in the art.
Various serum concentrations are added to constant amounts of labeled IMOL. IMOL-antigen complexes precipitate out of solution and are collected by centrifugation. The amount of precipitable IMOL-antigen complex is proportional to the amount of radioisotope detected in the precipitate. The amount of precipitable 1MOL-antigen complex is plotted against the serum concentration.
For various serum concentrations, a characteristic precipitin curve is obtained, in which the amount of precipitable 1MOL-antigen complex initially increases proportionately with increasing serum concentration, peaks at the equivalence point, and then decreases proportionately with further increases in serum concentration. Thus, the amount of precipitable IMOL-antigen complex is a measure of IMOL
activity which is characterized by sensitivity to both limiting and excess quantities of antigen.
XII. Functional Assays 1MOL function is assessed by expressing the sequences encoding 1MOL at physiologically elevated levels in mammalian cell culture systems. cDNA is subcloned into a mammalian expression vector containing a strong promoter that drives high levels of cDNA
expression. Vectors of choice include pCMV SPORT plasmid (Life Technologies) and pCR3.1 plasmid (Invitrogen), both of which contain the cytomegalovirus promoter. 5-10 ~cg of recombinant vector are transiently transfected into a human cell line, for example, an endothelial or hematopoietic cell line, using either liposome formulations or electroporation. 1-2 ~g of an additional plasmid containing sequences encoding a marker protein are co-transfected. Expression of a marker protein provides a means to distinguish transfected cells from nontransfected cells and is a reliable predictor of cDNA expression from the recombinant vector. Marker proteins of choice include, e.g., Green Fluorescent Protein (GFP;
Clontech), CD64, or a CD64.-GFP fusion protein. Flow cytometry (FCM), an automated, laser optics-based technique, is used to identify transfected cells expressing GFP or CD64-GFP and to evaluate the apoptotic state of the cells and other cellular properties. FCM detects and quantifies the uptake of fluorescent molecules that diagnose events preceding or coincident with cell death. These events include changes in nuclear DNA content as measured by staining of DNA with propidium iodide;
changes in cell size and granularity as measured by forward light scatter and 90 degree side light scatter; down-regulation of DNA synthesis as measured by decrease in bromodeoxyuridine uptake;
alterations in expression of cell surface and intracellular proteins as measured by reactivity with specific antibodies; and alterations in plasma membrane composition as measured by the binding of fluorescein-conjugated Annexin V protein to the cell surface. Methods in flow cytometry are discussed in Ormerod, M:G. (1994) Flow Cytometry, Oxford, New York NY.
The influence of IMOL on gene expression can be assessed using highly purified populations of cells transfected with sequences encoding IMOL and either CD64 or CD64-GFP.
CD64 and CD64-GFP are expressed on the surface of transfected cells and bind to conserved regions of human immunoglobulin G (IgG). Transfected cells are efficiently separated from nontransfected cells using magnetic beads coated with either human IgG or antibody against CD64 (DYNAL, Lake Success NY). mRNA can be purified from the cells using methods well known by those of skill in the art. Expression of mRNA encoding 1MOL and other genes of interest can be analyzed by northern analysis or microarray techniques.
XIII. Production of IMOL Specific Antibodies IMOL substantially purified using polyacrylamide gel electrophoresis (PAGE;
see, e.g:, Harrington, M.G. ( 1990) Methods Enzymol. 182:488-495), or other purification techniques, is used to immunize rabbits and to produce antibodies using standard protocols.
Alternatively, the IMOL amino acid sequence is analyzed using LASERGENE
software (DNASTAR) to determine regions of high immunogenicity, and a corresponding oligopeptide is synthesized and used to raise antibodies by means known to those of skill in the art. Methods for selection of appropriate epitopes, such as those near the C-terminus or in hydrophilic regions are well described in the art. (See, e.g., Ausubel, 1995, supra, ch. 11.) Typically, oligopeptides of about 15 residues in length are synthesized using an ABI 431A
peptide synthesizer (Perkin-Elmer) using fmoc-chemistry and coupled to KLH
(Sigma-Aldrich, St.
Louis MO) by reaction with N-maleimidobenzoyl-N-hydroxysuccinimide ester (MBS) to increase immunogenicity. (See, e.g., Ausubel, 1995, supra.) Rabbits are immunized with the oligopeptide-KLH complex in complete Freund's adjuvant. Resulting antisera are tested for antipeptide and anti-IMOL activity by, for example, binding the peptide or IMOL to a substrate, blocking with 1 % BSA, reacting with rabbit antisera, washing, and reacting with radio-iodinated goat anti-rabbit IgG.
Xl V. Purification of Naturally Occurring IMOL Using Specific Antibodies Naturally occurring or recombinant IMOL is substantially purified by immunoaffmity chromatography using antibodies specific for 1MOL. An immunoaffinity column is constructed by covalently coupling anti-IMOL antibody to an activated chromatographic resin, such as CNBr-activated SEPHAROSE (Amersham Pharmacia Biotech). After the coupling, the resin is blocked and washed according to the manufacturer's instructions.
Media containing IMOL are passed over the immunoaffinity column, and the column is washed under conditions that allow the preferential absorbance of IMOL (e.g., high ionic strength buffers in the presence of detergent). The column is eluted under conditions that disrupt antibody/IMOL binding (e.g., a buffer of pH 2 to pH 3, or a high concentration of a chaotrope, such as urea or thiocyanate ion), and IMOL is collected.
XV. Identification of Molecules Which Interact with IMOL
IMOL, or biologically active fragments thereof, are labeled with'z5I Bolton-Hunter reagent.
(See, e.g., Bolton A.E. and W.M. Hunter ( 1973) Biochem. J. 133:529-539.) Candidate molecules previously arrayed in the wells of a mufti-well plate are incubated with the labeled IMOL, washed, and any wells with labeled IMOL complex are assayed. Data obtained using different concentrations of IMOL are used to calculate values for the number, affinity, and association of IMOL with the candidate molecules.
Alternatively, molecules interacting with IMOL are analyzed using the yeast two-hybrid system as described in Fields, S. and O. Song (1989, Nature 340:245-246), or using commercially available kits based on the two-hybrid system, such as the MATCHMAKER system (Clontech).
Various modifications and variations of the described methods and systems of the invention will be apparent to those skilled in the art without departing from the scope and spirit of the invention. Although the invention has been described in connection with certain embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the described modes for carrying out the invention which are obvious to those skilled in molecular biology or related fields are intended to be within the scope of the following claims.

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SEQUENCE LISTING
<110> INCYTE PHARMACEUTICALS, INC.
YUE, Henry LAL, Preeti TANG, Y. Tom BAUGHN, Mariah R.
AZIMZAI, Yalda LU, Dyung Aina M.
<120> MOLECULES OF THE IMMUNE SYSTEM
<130> PF-0680 PCT
<140> To Be Assigned <141> Herewith <150> 60/127,852; 60/132,647 <151> 1999-04-05; 1999-05-05 <160> 30 <170> PERL Program <210> 1 <211> 613 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte 2705028CD1 <400> 1 Met Gly Ala Leu Arg Pro Thr Leu Leu Pro Pro Ser Leu Pro Leu Leu Leu Leu Leu Met Leu Gly Met Gly Cys Trp Ala Arg Glu Val Leu Val Pro Glu Gly Pro Leu Tyr Arg Val Ala Gly Thr Ala Val Ser Ile Ser Cys Asn Val Thr Gly Tyr Glu Gly Pro Ala Gln Gln Asn Phe Glu Trp Phe Leu Tyr Arg Pro Glu Ala Pro Asp Thr Ala 65 , , . 70 . 75 Leu Gly Ile Val Ser Thr Lys Asp Thr Gln Phe Ser Tyr Ala Val Phe Lys Ser Arg Val Val Ala Gly Glu Val Gln Val Gln Arg Leu Gln Gly Asp Ala Val Val Leu Lys Ile Ala Arg Leu Gln Ala Gln Asp Ala Gly Ile Tyr Glu Cys His Thr Pro Ser Thr Asp Thr Arg Tyr Leu Gly Ser Tyr Ser Gly Lys Val Glu Leu Arg Val Leu Pro Asp Val Leu Gln Val Ser Ala Ala Pro Pro Gly Pro Arg Gly Arg Gln Ala Pro Thr Ser Pro Pro Arg Met Thr Val His Glu Gly Gln Glu Leu Ala Leu Gly Cys Leu Ala Arg Thr Ser Thr Gln Lys His Thr His Leu Ala Val Ser Phe Gly Arg Ser Val Pro Glu Ala Pro Val Gly Arg Ser Thr Leu Gln Glu Val Val Gly Ile Arg Ser Asp Leu Ala Val Glu Ala Gly Ala Pro Tyr Ala Glu Arg Leu Ala Ala Gly Glu Leu Arg Leu Gly Lys Glu Gly Thr Asp Arg Tyr Arg Met Val Val Gly Gly Ala Gln Ala Gly Asp Ala Gly Thr Tyr His Cys Thr Ala Ala Glu Trp Ile Gln Asp Pro Asp Gly Ser Trp Ala Gln Ile Ala Glu Lys Arg Ala Val Leu Ala His Val Asp Val Gln Thr Leu Ser Ser Gln Leu Ala Val Thr Val Gly Pro Gly Glu Arg Arg Ile Gly Pro Gly Glu Pro Leu Glu Leu Leu Cys Asn Val Ser Gly.

Ala Leu Pro Pro Ala Gly Arg His Ala Ala Tyr Ser Val Gly Trp Glu Met Ala Pro Ala Gly Ala Pro Gly Pro Gly Arg Leu Val Ala Gln Leu Asp Thr Glu Gly Val Gly Ser Leu Gly Pro Gly Tyr Glu Gly Arg His Ile Ala Met Glu Lys Val Ala Ser Arg Thr Tyr Arg Leu Arg Leu Glu Ala Ala Arg Pro Gly Asp Ala Gly Thr Tyr Arg Cys Leu Ala Lys Ala Tyr Val Arg Gly Ser Gly Thr Arg Leu Arg Glu Ala Ala Ser Ala Arg Ser Arg Pro Leu Pro Val His Val Arg Glu Glu Gly Val Val Leu Glu Ala Val Ala Trp Leu Ala Gly Gly Thr Val Tyr Arg Gly Glu Thr Ala Ser Leu Leu Cys Asn Ile Ser Val Arg Gly Gly Pro Pro Gly Leu Arg Leu Ala Ala Ser Trp Trp Val Glu Arg Pro Glu Asp Gly Glu Leu Ser Ser Val Pro Ala Gln Leu Val Gly Gly Val Gly Gln Asp Gly Val Ala Glu Leu Gly Val Arg Pro Gly Gly Gly Pro Val Ser Val Glu Leu Val Gly Pro Arg Ser His Arg Leu Arg Leu His Ser Leu Gly Pro Glu Asp Glu Gly Val Tyr His Cys Ala Pro Ser Ala Trp Val Gln His Ala Asp Tyr Ser Trp Tyr Gln Ala Gly Ser Ala Arg Ser Gly Pro Val Thr Val Tyr Pro Tyr Met His Ala Leu Asp Thr Leu Phe Val Pro Leu Leu Val Gly Thr Gly Val Ala Leu Val Thr Gly Ala Thr Val Leu Gly Thr Ile Thr Cys Cys Phe Met Lys Arg Leu Arg Lys Arg <210> 2 <211> 271 <212> PRT
<213> Homo Sapiens <220>
<221> misc feature <223> Incyte ID No.: 2751129CD1 <400> 2 Met Val Val Val Thr Gly Arg Glu Pro Asp Ser Arg Arg Gln Asp Gly Ala Met Ser Ser Ser Asp Ala Glu Asp Asp Phe Leu Glu Pro Ala Thr Pro Thr Ala Thr Gln Ala Gly His Ala Leu Pro Leu Leu Pro Gln Glu Phe Pro Glu Val Val Pro Leu Asn Ile Gly Gly Ala His Phe Thr Thr Arg Leu Ser Thr Leu Arg Cys Tyr Glu Asp Thr Met Leu Ala Ala Met Phe Ser Gly Arg His Tyr Ile Pro Thr Asp Ser Glu Gly Arg Tyr Phe Ile Asp Arg Asp Gly Thr His Phe Gly Asp Val Leu Asn Phe Leu Arg Ser Gly Asp Leu Pro Pro Arg Glu Arg Val Arg Ala Val Tyr Lys Glu Ala Gln Tyr Tyr Ala Ile Gly Pro Leu Leu Glu Gln Leu Glu Asn Met Gln Pro Leu Lys Gly Glu Lys Val Arg Gln Ala Phe Leu Gly Leu Met Pro Tyr Tyr Lys Asp His Leu Glu Arg Ile Val Glu Ile Ala Gly Cys Val Arg Ser Ser Gly Arg Pro Ala Leu Pro Ser Ser Arg Ser Val Ser Ser Arg Arg Arg Cys Pro Ser Pro Pro Met Ser Val Arg Ser Ser Thr Pro Cys Asp Leu Ser Gly Val Arg Val Thr Gly Ser Phe Leu Ser Thr Thr Val Lys Trp Met Cys Leu Leu Gly Pro Gly Arg Leu Trp Leu Met Phe Met Thr Cys Cys Thr Ala Trp Ser Arg Thr Ser Arg Pro Arg Val Ser Pro Trp Thr Thr Ser Ala Ser Gly Cys Val Thr Ser Thr Ser <210> 3 <211> 235 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 2818076CD1 <400> 3 Met Ala Ser Ser Thr Ser Leu Pro Ala Pro Gly Ser Arg Pro Lys Lys Pro Leu Gly Lys Met Ala Asp Trp Phe Arg Gln Thr Leu Leu Lys Lys Pro Lys Lys Arg Pro Asn Ser Pro Glu Ser Thr Ser Ser Asp Ala Ser Gln Pro Thr Ser Gln Asp Ser Pro Leu Pro Pro Ser Leu Ser Ser Val Thr Ser Pro Ser Leu Pro Pro Thr His Ala Ser Asp Ser Gly Ser Ser Arg Trp Ser Lys Asp Tyr Asp Val Cys Val Cys His Ser Glu Glu Asp Leu Val Ala Ala Gln Asp Leu Val Ser Tyr Leu Glu Gly Ser Thr Ala Ser Leu Arg Cys Phe Leu Gln Leu Arg Asp Ala Thr Pro Gly Gly Ala Ile Val Ser Glu Leu Cys Gln Ala Leu Ser Ser Ser His Cys Arg Val Leu Leu Ile Thr Pro Gly Phe Leu Gln Asp Pro Trp Cys Lys Tyr Gln Met Leu Gln Ala Leu Thr Glu Ala Pro Gly Ala Glu Gly Cys Thr Ile Pro Leu Leu Leu Gly Leu Ser Arg Ala Ala Tyr Pro Pro Glu Leu Arg Phe Met Tyr Tyr Val Asp Gly Arg Gly Pro Asp Gly Gly Phe Arg Gln Val Lys Glu Ala Val Met Arg Cys Lys Leu Leu Gln Glu Gly Glu Gly Glu Arg Asp Ser Ala Thr Val Ser Asp Leu Leu <210> 4 <211> 310 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No.: 2907049CD1 <400> 4 Met Gly Pro Gly Leu Leu His Trp Met Ala Leu Cys Leu Leu Gly Thr Gly His Gly Asp Ala Met Val Ile Gln Asn Pro Arg Tyr Gln Val Thr Gln Phe Gly Lys Pro Val Thr Leu Ser Cys Ser Gln Thr Leu Asn His Asn Val Met Tyr Trp Tyr Gln Gln Lys Ser Ser Gln Ala Pro Lys Leu Leu Phe His Tyr Tyr Asp Lys Asp Phe Asn Asn Glu Ala Asp Thr Pro Asp Asn Phe Gln Ser Arg Arg Pro Asn Thr Ser Phe Cys Phe Leu Asp Ile Arg Ser Pro Gly Leu Gly Asp Ala Ala Met Tyr Leu Cys Ala Thr Ser Lys Tyr Arg Asp Gly Glu Leu Phe Phe Gly Glu Gly Ser Arg Leu Thr Val Leu Glu Asp Leu Lys Asn Val Phe Pro Pro Glu Val Ala Val Phe Glu Pro Ser Glu Ala Glu Ile Ser His Thr Gln Lys Ala Thr Leu Val Cys Leu Ala Thr Gly Phe Tyr Pro Asp His Val Glu Leu Ser Trp Trp Val Asn Gly Lys Glu Val His Ser Gly Val Ser Thr Asp Pro Gln Pro Leu Lys Glu Gln Pro Ala Leu Asn Asp Ser Arg Tyr Cys Leu Ser Ser Arg Leu Arg Val Ser Ala Thr Phe Trp Gln Asn Pro Arg Asn His Phe Arg Cys Gln Val Gln Phe Tyr Gly Leu Ser Glu Asn Asp Glu Trp Thr Gln Asp Arg Ala Lys Pro Val Thr Gln Ile Val Ser Ala Glu Ala Trp Gly Arg Ala Asp Cys Gly Phe Thr Ser Glu Ser Tyr Gln Gln Gly Val Leu Ser Ala Thr Ile Leu Tyr Glu Ile Leu Leu Gly Lys Ala Thr Leu Tyr Ala Val Leu Val Ser Ala Leu Val Leu Met Ala Met Val Lys Arg Lys Asp Ser Arg Gly <210> 5 <211> 246 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3402252CD1 <400> 5 Met Ala Ala Ala Ala Ile Pro Ala Leu Leu Leu Cys Leu Pro Leu Leu Phe Leu Leu Phe Gly Trp Ser Arg Ala Arg Arg Asp Asp Pro His Ser Leu Cys Tyr Asp Ile Thr Val Ile Pro Lys Phe Arg Pro Gly Pro Arg Trp Cys Ala Val Gln Gly Gln Val Asp Glu Lys Thr Phe Leu His Tyr Asp Cys Gly Asn Lys Thr Val Thr Pro Val Ser Pro Leu Gly Lys Lys Leu Asn Val Thr Thr Ala Trp Lys Ala Gln Asn Pro Val Leu Arg Glu Val Val Asp Ile Leu Thr Glu Gln Leu Arg Asp Ile Gln Leu Glu Asn Tyr Thr Pro Lys Glu Pro Leu Thr Leu Gln Ala Arg Met Ser Cys Glu Gln Lys Ala Glu Gly His Ser Ser Gly Ser Trp Gln Phe Ser Phe Asp Gly Gln Ile Phe Leu Leu Phe Asp Ser Glu Lys Arg Met Trp Thr Thr Val His Pro Gly Ala Arg Lys Met Lys Glu Lys Trp Glu Asn Asp Lys Val Val Ala Met Ser Phe His Tyr Phe Ser Met Gly Asp Cys Ile Gly Trp Leu Glu Asp Phe Leu Met Gly Met Asp Ser Thr Leu Glu Pro Ser Ala Gly Ala Pro Leu Ala Met Ser Ser Gly Thr Thr Gln Leu Arg Ala Thr Ala Thr Thr Leu Ile Leu Cys Cys Leu Leu Ile Ile Leu Pro Cys Phe Ile Leu Pro Gly Ile <210> 6 <211> 180 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3577142CD1 <400> 6 Met Asp Trp Pro His Asn Leu Leu Phe Leu Leu Thr Ile Ser Ile Phe Leu Gly Leu Gly Gln Pro Arg Ser Pro Lys Ser Lys Arg Lys Gly Gln Gly Arg Pro Gly Pro Leu Val Pro Gly Pro His Gln Val Pro Leu Asp Leu Val Ser Arg Met Lys Pro Tyr Ala Arg Met Glu Glu Tyr Glu Arg Asn Ile Glu Glu Met Val Ala Gln Leu Arg Asn Ser Ser Glu Leu Ala Gln Arg Lys Cys Glu Val Asn Leu Gln Leu Trp Met Ser Asn Lys Arg Ser Leu Ser Pro Trp Gly Tyr Ser Ile Asn His Asp Pro Ser Arg Ile Pro Val Asp Leu Pro Glu Ala Arg Cys Leu Cys Leu Gly Cys Val Asn Pro Phe Thr Met Gln Glu Asp Arg Ser Met Val Ser Val Pro Val Phe Ser Gln Val Pro Val Arg Arg Arg Leu Cys Pro Pro Pro Pro Arg Thr Gly Pro Cys Arg Gln Arg Ala Val Met Glu Thr Ile Ala Val Gly Cys Thr Cys Ile Phe <210> 7 <211> 200 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3725986CD1 <400> 7 Met Glu Ser Gln Leu Gln Glu Arg Val Glu Ser Ser Arg Arg Ala Val Ser Gln Ile Val Thr Val Tyr Asp Lys Leu Gln Glu Lys Val Glu Leu Leu Ser Arg Lys Leu Asn Ser Gly Asp Asn Leu Ile Val Glu Glu Ala Val Gln Glu Leu Asn Ser Phe Leu Ala Gln Glu Asn Met Arg Leu Gln Glu Leu Thr Asp Leu Leu Gln Glu Lys His Arg Thr Met Ser Gln Glu Phe Ser Lys Leu Gln Ser Lys Val Glu Thr 80 ~ 85 90 Ala Glu Ser Arg Val Ser Val Leu Glu Ser Met Ile Asp Asp Leu Gln Trp Asp Ile Asp Lys Ile Arg Lys Arg Glu Gln Arg Leu Asn Arg His Leu Ala Glu Val Leu Glu Arg Val Asn Ser Lys Gly Tyr Lys Val Tyr Gly Ala Gly Ser Ser Leu Tyr Gly Gly Thr Ile Thr Ile Asn Ala Arg Lys Phe Glu Glu Met Asn Ala Glu Leu Glu Glu Asn Lys Glu Leu Ala Gln Asn Arg Leu Cys Glu Leu Glu Lys Leu Arg Gln Asp Phe Glu Glu Val Thr Thr Gln Asn Glu Lys Leu Lys Val Arg Thr His Pro <210> 8 <211> 211 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3799011CD1 <400> 8 Met Trp Leu Pro Trp Ala Leu Leu Leu Leu Trp Val Pro Gly Cys Phe Ala Leu Ser Lys Cys Arg Thr Val Ala Gly Pro Val Gly Gly Ser Leu Ser Val Gln Cys Pro Tyr Glu Lys Glu His Arg Thr Leu Asn Lys Tyr Trp Cys Arg Pro Pro Gln Ile Phe Leu Cys Asp Lys Ile Val Glu Thr Lys Gly Ser Ala Gly Lys Arg Asn Gly Arg Val Ser Ile Arg Asp Ser Pro Ala Asn Leu Ser Phe Thr Val Thr Leu Glu Asn Leu Thr Glu Glu Asp Ala Gly Thr Tyr Trp Cys Gly Val Asp Thr Pro Trp Leu Arg Asp Phe His Asp Pro Val Val Glu Val Glu Val Ser Val Phe Pro Ala Ser Thr Ser Met Thr Pro Ala Ser 125 _ 130 135 Ile Thr Ala Ala Lys Thr Ser Thr Ile Thr Thr Ala Phe Pro Pro Val Ser Ser Thr Thr Leu Phe Ala Val Gly Ala Thr His Ser Ala Ser Ile Gln Glu Glu Thr Glu Glu Val Val Asn Ser Gln Leu Pro Leu Thr Pro Leu Pro Ala Gly Ile Val Ala Ala Ser Val Gly Gly Gly Leu Pro Ala Ser Leu Glu Asp Val Ser Glu Met Asp Gln Ser Trp <210> 9 <211> 225 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3887384CD1 <400> 9 Met Ser Arg Pro Arg Lys Arg Leu Ala Gly Thr Ser Gly Ser Asp Lys Gly Leu Ser Gly Lys Arg Thr Lys Thr Glu Asn Ser Gly Glu Ala Leu Ala Lys Val Glu Asp Ser Asn Pro Gln Lys Thr Ser Ala Thr Lys Asn Cys Leu Lys Asn Leu Ser Ser His Trp Leu Met Lys Ser Glu Pro Glu Ser Arg Leu Glu Lys Gly Val Asp Val Lys Phe Ser Ile Glu Asp Leu Lys Ala Gln Pro Lys Gln Thr Thr Cys Trp Asp Gly Val Arg Asn Tyr Gln Ala Arg Asn Phe Leu Arg Ala Met Lys Leu Gly Glu Glu Ala Phe Phe Tyr His Ser Asn Cys Lys Glu Pro Gly Ile Ala Gly Leu Met Lys Ile Val Lys Glu Ala Tyr Pro Asp His Thr Gln Phe Glu Lys Asn Asn Pro His Tyr Asp Pro Ser Ser Lys Glu Asp Asn Pro Lys Trp Ser Met Val Asp Val Gln Phe Val Arg Met Met Lys Arg Phe Ile Pro Leu Ala Glu Leu Lys Ser Tyr His Gln Ala His Lys Ala Thr Gly Gly Pro Leu Lys Asn Met Val Leu Phe Thr Arg Gln Arg Leu Ser Ile Gln Pro Leu Thr Gln Glu Glu Phe Asp Phe Val Leu Ser Leu Glu Glu Lys Glu Pro Ser <210> 10 <211> 329 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 1352789CD1 <400> 10 Met Ala Ala Gln Tyr Gly Ser Met Ser Phe Asn Pro Ser Thr Pro Gly Ala Ser Tyr Gly Pro Gly Arg Gln Glu Pro Arg Asn Ser Gln Leu Arg Ile Val Leu Val Gly Lys Thr Gly Ala Gly Lys Ser Ala Thr Gly Asn Ser Ile Leu Gly Arg Lys Val Phe His Ser Gly Thr Ala Ala Lys Ser Ile Thr Lys Lys Cys Glu Lys Arg Ser Ser Ser Trp Lys Glu Thr Glu Leu Val Val Val Asp Thr Pro Gly Ile Phe Asp Thr Glu Val Pro Asn Ala Glu Thr Ser Lys Glu Ile Ile Arg Cys Ile Leu Leu Thr Ser Pro Gly Pro His Ala Leu Leu Leu Val Val Pro Leu Gly Arg Tyr Thr Glu Glu Glu His Lys Ala Thr Glu Lys Ile Leu Lys Met Phe Gly Glu Arg Ala Arg Ser Phe Met Ile Leu Ile Phe Thr Arg Lys Asp Asp Leu Gly Asp Thr Asn Leu His Asp Tyr Leu Arg Glu Ala Pro Glu Asp Ile Gln Asp Leu Met Asp Ile Phe Gly Asp Arg Tyr Cys Ala Leu Asn Asn Lys Ala Thr Gly Ala Glu Gln Glu Ala Gln Arg Ala Gln Leu Leu Gly Leu Ile Gln Arg Val Val Arg Glu Asn Lys Glu Gly Cys Tyr Thr Asn Arg Met Tyr Gln Arg Ala Glu Glu Glu Ile Gln Lys Gln Thr Gln Ala Met Gln Glu Leu His Arg Val Glu Leu Glu Arg Glu Lys Ala Arg Ile Arg Glu Glu Tyr Glu Glu Lys Ile Arg Lys Leu Glu Asp Lys Val Glu Gln Glu Lys Arg Lys Lys Gln Met Glu Lys Lys Leu Ala Glu Gln Glu Ala His Tyr Ala Val Arg Gln Gln Arg Ala Arg Thr Glu Val Glu Ser Lys Asp Gly Ile Leu Glu Leu Ile Met Thr Ala Leu Gln Ile Ala Ser Phe Ile Leu Leu Arg Leu Phe Ala Glu Asp <210> 11 <211> 237 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 1666486CD1 <400> 11 Met Asp Met Arg Val Pro Ala Gln Leu Leu Gly Leu Leu Leu Leu Trp Leu Pro Gly Ala Lys Cys Asp Ile Leu Leu Thr Gln Ser Pro Ser Thr Val Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr Cys Arg Ala Thr Gln Ser Ile Gly Ser Trp Val Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Gln Leu Leu Ile Tyr Lys Ala Ser Ser Leu Glu Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Glu Phe Thr Leu Ser Ile Asn Ser Leu Gln Pro Asp Asp Phe Ala Thr Tyr Phe Cys Gln Gln Tyr Asp Thr Tyr Pro Thr Trp Ser Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Leu Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 12 <211> 235 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 1706596CD1 <400> 12 Met Pro Trp Ala Leu Leu Leu Leu Thr Leu Leu Thr His Ser Ala Val Ser Val Val Gln Ala Gly Leu Thr Gln Pro Pro Ser Val Ser Arg Ala Leu Arg Gln Thr Ala Thr Leu Thr Cys Thr Gly Asn Asn Asn Asn Val Gly Asn Gln Gly Ala Ala Trp Leu Glri Gln His Gln Gly His Pro Pro Lys Leu Leu Ser Tyr Arg Asn Asn Asn Arg Pro Ser Gly Ile Ser Glu Arg Phe Ser Ala Ser Arg Ser Arg Asn Thr Ala Ser Leu Thr Ile Thr Gly Leu Gln Pro Glu Asp Glu Ala Asp Tyr Tyr Cys Ser Val Trp Asp Ser Ser Leu Ser Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Ser Gln Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Aia Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val Ala Pro Thr Glu Cys Ser <210> 13 <211> 246 <212> PRT

<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No.: 1890540CD1 <400> 13 Met Leu Trp Arg Gln Leu Ile Tyr Trp Gln Leu Leu Ala Leu Phe Phe Leu Pro Phe Cys Leu Cys Gln Asp Glu Tyr Met Glu Ser Pro Gln Thr Gly Gly Leu Pro Pro Asp Cys Ser Lys Cys Cys His Gly 35 ~ 40 45 Asp Tyr Ser Phe Arg Gly Tyr Gln Gly Pro Pro Gly Pro Pro Gly Pro Pro Gly Ile Pro Gly Asn His Gly Asn Asn Gly Asn Asn Gly Ala Thr Gly His Glu Gly Ala Lys Gly Glu Lys Gly Asp Lys Gly Asp Leu Gly Pro Arg Gly Glu Arg Gly Gln His Gly Pro Lys Gly Glu Lys Gly Tyr Pro Gly Ile Pro Pro Glu Leu Gln Ile Ala Phe Met Ala Ser Leu Ala Thr His Phe Ser Asn Gln Asn Ser Gly Ile Ile Phe Ser Ser Val Glu Thr Asn Ile Gly Asn Phe Phe Asp Val Met Thr Gly Arg Phe Gly Ala Pro Val Ser Gly Val Tyr Phe Phe Thr Phe Ser Met Met Lys His Glu Asp Val Glu Glu Val Tyr Val Tyr Leu Met His Asn Gly Asn Thr Val Phe Ser Met Tyr Ser Tyr Glu Met Lys Gly Lys Ser Asp Thr Ser Ser Asn His Ala Val Leu Lys Leu Ala Lys Gly Asp Glu Val Trp Leu Arg Met Gly Asn Gly Ala Leu His Gly Asp His Gln Arg Phe Ser Thr Phe Ala Gly Phe Leu Leu Phe Glu Thr Lys <210> 14 <211> 322 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No.: 2774913CD1 <400> 14 Met Pro Val Thr Val Thr Arg Thr Thr Ile Thr Thr Thr Thr Thr Ser Ser Ser Gly Leu Gly Ser Pro Met Ile Val Gly Ser Pro Arg Ala Leu Thr Gln Pro Leu Gly Leu Leu Arg Leu Leu Gln Leu Val Ser Thr Cys Val Ala Phe Ser Leu Val Ala Ser Val Gly Ala Trp Thr Gly Ser Met Gly Asn Trp Ser Met Phe Thr Trp Cys Phe Cys Phe Ser Val Thr Leu Ile Ile Leu Ile Val Glu Leu Cys Gly Leu Gln Ala Arg Phe Pro Leu Ser Trp Arg Asn Phe Pro Ile Thr Phe Ala Cys Tyr Ala Ala Leu Phe Cys Leu Ser Ala Ser Ile Ile Tyr Pro Thr Thr Tyr Val Gln Phe Leu Ser His Gly Arg Ser Arg Asp His Ala Ile Ala Ala Thr Phe Phe Ser Cys Ile Ala Cys Val Ala Tyr Ala Thr Glu Val Ala Trp Thr Arg Ala Arg Pro Gly Glu Ile Thr Gly Tyr Met Ala Thr Val Pro Gly Leu Leu Lys Val Leu Glu Thr Phe Val Ala Cys Ile Ile Phe Ala Phe Ile Ser Asp Pro Asn Leu Tyr Gln His Gln Pro Ala Leu Glu Trp Cys Val Ala Val Tyr Ala Ile Cys Phe Ile Leu Ala Ala Ile Ala Ile Leu Leu Asn Leu Gly Glu Cys Thr Asn Val Leu Pro Ile Pro Phe Pro Ser Phe Leu Ser Gly Leu Ala Leu Leu Ser Val Leu Leu Tyr Ala Thr Ala Leu Val Leu Trp Pro Leu Tyr Gln Phe Asp Glu Lys Tyr Gly Gly Gln Pro Arg Arg Ser Arg Asp Val Ser Cys Ser Arg Ser His Ala Tyr Tyr Val Cys Ala Trp Asp Arg Arg Leu Ala Val Ala Ile Leu Thr Ala Ile Asn Leu Leu Ala Tyr Val Ala Asp Leu Val His Ser Ala His Leu Val Phe Val Lys Val <210> 15 <211> 191 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No.: 5571291CD1 <400> 15 Met Trp Arg His Glu Arg Ile Lys Lys Thr Ser Phe Ser Thr Thr Thr Leu Leu Pro Pro Ile Lys Val Leu Val Val Tyr Pro Ser Glu Ile Cys Phe His His Thr Ile Cys Tyr Phe Thr Glu Phe Leu Gln Asn His Cys Arg Ser Glu Val Ile Leu Glu Lys Trp Gln Lys Lys Lys Ile Ala Glu Met Gly Pro Val Gln Trp Leu Ala Thr Gln Lys Lys Ala Ala Asp Lys Val Val Phe Leu Leu Ser Asn Asp Val Asn Ser Val Cys Asp Gly Thr Cys Gly Lys Ser Glu Gly Ser Pro Ser Glu Asn Ser Gln Asp Leu Phe Pro Leu Ala Phe Asn Leu Phe Cys Ser Asp Leu Arg Ser Gln I1e His Leu His Lys Tyr Val Val Val Tyr Phe Arg Glu Ile Asp Thr Lys Asp Asp Tyr Asn Ala Leu Ser Val Cys Pro Lys Tyr His Leu Met Lys Asp Ala Thr Ala Phe Cys Ala Glu Leu Leu His Val Lys Gln Gln Val Ser Ala Gly Lys Arg Ser Gln Ala Cys His Asp Gly Cys Cys Ser Leu <210> 16 <211> 2265 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 2705028CB1 <400> 16 cagttcagct cgctcggcgc acccacgcct cgctgccccg cttcctgccc tcaacctggg 60 catgctcccc ccacccttcc ggccccccag aacccgcgcc atcccccgga gcctccccag 120 agctggccgc gcaggatggg cgccctcagg cccacgctgc tgccgccttc gctgccgctg 180 ctgctgctgc taatgctagg aatgggatgc tgggcccggg aggtgctggt ccccgagggg 240 cccttgtacc gcgtggctgg cacagctgtc tccatctcct gcaatgtgac cggctatgag 300 ggccctgccc agcagaactt cgagtggttc ctgtataggc ccgaggcccc agatactgca 360 ctgggcattg tcagtaccaa ggatacccag ttctcctatg ctgtcttcaa gtcccgagtg 420 gtggcgggtg aggtgcaggt gcagcgccta caaggtgatg ccgtggtgct caagattgcc 480 cgcctgcagg cccaggatgc cggcatttat gagtgccaca ccccctccac tgatacccgc 540 tacctgggca gctacagcgg caaggtggag ctgagagttc ttccagatgt cctccaggtg 600 tctgctgccc ccccagggcc ccgaggccgc caggccccaa cctcaccccc acgcatgacg 660 gtgcatgagg ggcaggagct ggcactgggc tgcctggcga ggacaagcac acagaagcac 720 acacacctgg cagtgtcctt tgggcgatct gtgcccgagg caccagttgg gcggtcaact 780 ctgcaggaag tggtgggaat ccggtcagac ttggccgtgg aggctggagc tccctatgct 840 gagcgattgg ctgcagggga gcttcgtctg ggcaaggaag ggaccgatcg gtaccgcatg 900 gtagtagggg gtgcccaggc aggggacgca ggcacctacc actgcactgc cgctgagtgg 960 attcaggatc ctgatggcag ctgggcccag attgcagaga aaagggccgt cctggcccac 1020 gtggatgtgc agacgctgtc cagccagctg gcagtgacag tggggcctgg tgaacgtcgg 1080 atcggcccag gggagccctt ggaactgctg tgcaatgtgt caggggcact tcccccagca 1140 ggccgtcatg ctgcatactc tgtaggttgg gagatggcac ctgcgggggc acctgggccc 1200 ggccgcctgg tagcccagct ggacacagag ggtgtgggca gcctgggccc tggctatgag 1260 ggccgacaca ttgccatgga gaaggtggca tccagaacat accggctacg gctagaggct 1320 gccaggcctg gtgatgcggg cacctaccgc tgcctcgcca aagcctatgt tcgagggtct 1380 gggacccggc ttcgtgaagc agccagtgcc cgttcccggc ctctccctgt acacgtgcgg 1440 gaggaaggtg tggtgctgga ggctgtggca tggctagcag gaggcacagt gtaccgcggg 1500 gagactgcct ccctgctgtg caacatctct gtgcggggtg gccccccagg actgcggctg 1560 gccgccagct ggtgggtgga gcgaccagag gacggagagc tcagctctgt ccctgcccag 1620 ctggtgggtg gcgtaggcca ggatggtgtg gcagagctgg gagtccggcc tggaggaggc 1680 cctgtcagcg tagagctggt ggggccccga agccatcggc tgagactaca cagcttgggg 1740 cccgaggatg aaggcgtgta ccactgtgcc cccagcgcct gggtgcagca tgccgactac 1800 agctggtacc aggcgggcag tgcccgctca gggcctgtta cagtctaccc ctacatgcat 1860 gccctggaca ccctatttgt gcctctgctg gtgggtacag gggtggccct agtcactggt 1920 gccactgtcc ttggtaccat cacttgctgc ttcatgaaga ggcttcgaaa acggtgatcc 1980 cttactcccc aggtcttgca ggtgtcgact gtcttccggc ccagctccaa gccctcctct 2040 ggttgcctgg acaccctctc cctctgtcca ctcttccttt aatttatttg acctcccact 2100 acccagaatg ggagacgtgc ctccccttcc ccactccttc cctcccaagc ccctccctct 2160 ggccttctgt tcttgatctc ttagggatcc tatagggagg ccatttcctg tcctggaatt 2220 agtttttcta aaatgtgaat aaacttgttt tataaaaaaa aaaaa 2265 <210> 17 <211> 1124 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 2751129CB1 <400> 17 ccgcccccac tgcccagagc cagagggatg gtggtagtca cggggcggga gccagacagc 60 cgtcgtcagg acggtgccat gtccagctct gacgccgaag acgactttct ggagccggcc 120 acgccgacgg ccacgcaggc ggggcacgcg ctgcccctgc tgccacagga gtttcctgag 180 gttgttcccc ttaacatcgg aggggctcac ttcactacac gcctgtccac actgcggtgc 240 tacgaagaca ccatgttggc agccatgttc agtgggcggc actacatccc cacggactcc 300 gagggccggt acttcatcga ccgagatggc acacactttg gagatgtgct gaatttcctg 360 cgctcagggg acctcccacc cagggagcgt gttcgagctg tgtacaaaga ggcccagtac 420 tatgccatcg ggcccctcct ggagcagctg gagaacatgc agccactgaa gggcgagaag 480 gtgcgccaag cgtttctggg actcatgccc tattacaaag accacttgga gcggattgtg 540 gagatcgccg gctgcgtgcg gtccagcgga aggcccgctt tgccaagctc aaggtctgtg 600 tcttcaagga ggagatgccc atcaccccct atgagtgtcc gctcctcaac tccctgcgat 660 ttgagcggag tgagagtgac gggcagcttt ttgagcacca ctgtgaagtg gatgtgtctt 720 ttgggccctg ggaggctgtg gctgatgttt atgacctgct gcactgcctg gtcac tctcggccca gggtctcacc gtggaccacc agtgcatcgg ggtgtgtgac aagcacctcg 840 tgaaccacta ctact caa ct cccatct at a ttcaa ggacc 780 ctcc g g g g g gatcacatgg tggtgagtag 900 ggtagg cgagagtctc atcagggagg atgtccacct tgcttggtgg ctctgggagt 960 aagattcctg aaggggctgc tgactgccca gaatcctgcg aagtgagaac agcatcctga 1020 agcaaagctc ccagggacag aagtggtagt caatttcctg actgcactaa ggtttggctc 1080 aggtttcggc atgagantca ttcgngtaac tggctttctc agga 1124 <210> 18 <211> 1082 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 2818076CB1 <400> 18 gcccgcgcag tccgcgcagc ctcatcgcaa ctgggcccgc gcgcaggcct tacataggaa 60 gtccttctaa agagctgcct gccagctgcc cttccccaga tcccgaatat cctcctggcc 120 aggtggagca gagaacagtt cctcagctgg tcatgctgag ctcataccct gatggctgct 180 ccatgaggtc aagactgggt ctcctccctc ctcccccttc accaatgcct ggtctcacgg 240 ggctagtttt gacccccacg ctatggcatc atcgacctcc ctcccagctc ctggctctcg 300 gcctaagaag cctctaggca agatggctga ctggttcagg cagaccctgc tgaagaagcc 360 caagaagagg cccaactccc cagaaagcac ctccagcgat gcttcacagc ctacctcaca 420 ggacagccca ctacccccaa gcctcagctc agtcacgtct cccagcctgc cacccacaca 480 tgcgagtgac agtggcagta gtcgctggag caaagactat gacgtctgcg tgtgccacag 540 tgaggaagac ctggtggccg cccaagacct ggtctcctac ttggaaggca gcactgccag 600 cctgcgctgc ttcctgcaac tccgggatgc aaccccaggc ggcgctatag tgtccgagct 660 gtgccaggca ctgagcagta gtcactgccg ggtgctgctc atcacgccgg gcttccttca 720 ggacccctgg tgcaagtacc agatgctgca ggccctgacc gaggctccag gggccgaggg 780 ctgcaccatc cccctgctgt tgggcctcag cagagctgcc tacccacctg agctccgatt 840 catgtactac gtcgatggca ggggccctga tggtggcttt cgtcaagtca aagaagctgt 900 catgcgttgt aagctactac aggagggaga aggggaacgg gattcagcta cagtatctga 960 tctactttga cttttaggag acagccctgt agcctagtag ttcaaagcgc agcttctgga 1020 aaaggctgtc ggggtttgta tcctggctcc tgcacttatt aacccataaa aagtaacttg 1080 tg <210> 19 <211> 1180 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 2907049CB1 <400> 19 cagacagatg cttcattcct gtatggggtg gtattcctgc catgggtcct gggcttctcc 60 actggatggc cctttgtctc cttggaacag gtcatgggga tgccatggtc atccagaacc 120 caagatacca ggttacccag tttggaaagc cagtgaccct gagttgttct cagactttga 180 accataacgt catgtactgg taccagcaga agtcaagtca ggccccaaag ctgctgttcc 240 actactatga caaagatttt aacaatgaag cagacacccc tgataacttc caatccagga 300 ggccgaacac ttctttctgc tttcttgaca tccgctcacc aggcctgggg gacgcagcca 360 tgtacctgtg tgccaccagc aaatacaggg acggggagct gttttttgga gaaggctcta 420 ggctgaccgt actggaggac ctgaaaaacg tgttcccacc cgaggtcgct gtgtttgagc 480 catcagaagc agagatctcc cacacccaaa aggccacact ggtatgcctg gccacaggct 540 tctaccccga ccacgtggag ctgagctggt gggtgaatgg gaaggaggtg cacagtgggg 600 tcagcacaga cccgcagccc ctcaaggagc agcccgccct caatgactcc agatactgcc 660 tgagcagccg cctgagggtc tcggccacct tctggcagaa cccccgcaac cacttccgct 720 gtcaagtcca gttctacggg ctctcggaga atgacgagtg gacccaggat agggccaaac 780 ctgtcaccca gatcgtcagc gccgaggcct ggggtagagc agactgtggc ttcacctccg 840 agtcttacca gcaaggggtc ctgtctgcca ccatcctcta tgagatcttg ctagggaagg 900 ccaccttgta tgccgtgctg gtcagtgccc tcgtgctgat ggccatggtc aagagaaagg 960 attccagagg ctagctccaa aaccatccca ggtcattctt catcctcacc caggattctc 1020 ctgtacctgc tcccaatctg tgttcctaaa agtgattctc actctgcttc tcatctccta 1080 cttacatgaa tacttctctc ttttttctgt ttccctgaag attgagctcc caacccccaa 1140 gtacgaaata ggctaaacca ataaaaaatt gtgtgttgga 1180 <210> 20 <211> 1307 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3402252CB1 <400> 20 gatccacctt gattaaatct cttgtcccca gccctcctgg tccccaatgg cagcagccgc 60 catcccagct ttgcttctgt gcctcccgct tctgttcctg ctgttcggct ggtcccgggc 120 taggcgagac gaccctcact ctctttgcta tgacatcacc gtcatcccta agttcagacc 180 tggaccacgg tggtgtgcgg ttcaaggcca ggtggatgaa aagacttttc ttcactatga 240 ctgtggcaac aagacagtca cacctgtcag tcccctgggg aagaaactaa atgtcacaac 300 ggcctggaaa gcacagaacc cagtactgag agaggtggtg gacatactta cagagcaact 360 gcgtgacatt cagctggaga attacacacc caaggaaccc ctcaccctgc aggcaaggat 420 gtcttgtgag cagaaagctg aaggacacag cagtggatct tggcagttca gtttcgatgg 480 gcagatcttc ctcctctttg actcagagaa gagaatgtgg acaacggttc atcctggagc 540 cagaaagatg aaagaaaagt gggagaatga caaggttgtg gccatgtcct tccattactt 600 ctcaatggga gactgtatag gatggcttga ggacttcttg atgggcatgg acagcaccct 660 ggagccaagt gcaggagcac cactcgccat gtcctcaggc acaacccaac tcagggccac 720 agccaccacc ctcatccttt gctgcctcct catcatcctc ccctgcttca tcctccctgg 780 catctgagga gagtccttta gagtgacagg ttaaagctga taccaaaagg ctcctgtgag 840 cacggtcttg atcaaactcg cccttctgtc tggccagctg cccacgacct acggtgtatg 900 tccagtggcc tccagcagat catgatgaca tcatggaccc aatagctcat tcactgcctt 960 gattcctttt gccaacaatt ttaccagcag ttatacctaa catattatgc aattttctct 1020 tggtgctacc tgatggaatt cctgcactta aagttctggc tgactaaaca agatatatca 1080 ttttctttct tctctttttg tttggaaaat caagtacttc tttgaatgat gatctctttc 1140 ttgcaaatga tattgtcagt aaaataatca cgttagactt cagacctctg gggattcttt 1200 ccgtgtcctg aaagagaatt tttaaattat ttaataagaa aaaatttata ttaatgattg 1260 tttcctttag taatttattg ttctgtactg atatttaaat aacgcat 1307 <210> 21 <211> 689 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3577142CB1 <400> 21 cggcggcatc tggggttcca ggcgggcagc agctgcaggc tgaccttgca gcttggcgga 60 atggactggc ctcacaacct gctgtttctt cttaccattt ccatcttcct ggggctgggc 120 cagcccagga gccccaaaag caagaggaag gggcaagggc ggcctgggcc cctggtccct 180 ggccctcacc aggtgccact ggacctggtg tcacggatga aaccgtatgc ccgcatggag 240 gagtatgaga ggaacatcga ggagatggtg gcccagctga ggaacagctc agagctggcc 300 cagagaaagt gtgaggtcaa cttgcagctg tggatgtcca acaagaggag cctgtctccc 360 tggggctaca gcatcaacca cgaccccagc cgtatccccg tggacctgcc ggaggcacgg 420 tgcctgtgtc tgggctgtgt gaaccccttc accatgcagg aggaccgcag catggtgagc 480 gtgccggtgt tcagccaggt tcctgtgcgc cgccgcctct gcccgccacc gccccgcaca 540 gggccttgcc gccagcgcgc agtcatggag accatcgctg tgggctgcac ctgcatcttc 600 tgaatcacct ggcccagaag ccaggccagc agcccgagac catcctcctt gcacctttgt 660 gccaagaaag gcctatgaaa agtaaacac 689 <210> 22 <211> 818 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3725986CB1 <400> 22 aagggcaaga gccagctttc tctttccttg ctactttggc cagcagttcc agtgaagaga 60 tggagtctca gctgcaggaa cgtgtggagt cttcccgccg agccgtgtcc cagattgtga 120 ctgtttatga taaattgcaa gaaaaagtgg agctcttatc ccggaagcta aacagtggag 180 ataatctgat agtggaggaa gcagtgcagg agctgaactc tttcctcgca caggagaata 240 tgaggctaca ggaattgaca gatcttcttc aggaaaagca tcgcaccatg tctcaggagt 300 tctccaagtt gcagagtaaa gtggagacag ccgaatcacg agtgtctgtc ctggagtcca 360 tgattgatga cctgcagtgg gatattgaca aaattcgaaa gagggaacag cgactcaacc 420 ga:cacttagc agaagtccta gaacgggtga attccaaagg ttataaggtg tatggagcgg 480 ggagcagtct gtatggcggc acaatcacta tcaatgctcg gaagtttgag gaaatgaatg 540 cagagcttga ggagaacaaa gagttggctc agaaccgtct ctgtgagctg gagaaacttc 600 ggcaagactt tgaggaggtc actacacaaa atgaaaagct gaaggtacga acgcatccct 660 gaagggcagt aaaatcagac gttctgctga tcaactcacg tatatacata gttgtgaatc 720 tgcgtattca tgagggataa gaaaaatgta gacaaaatcc aacatccttt tatgataaaa 780 ctcttaacaa attaggtgta aagaagtgta cctcaaca 81g <210> 23 <211> 899 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3799011CB1 <400> 23 gccttggagg cgtgactttc ccctcgggtc caggtagggc ctggagctgc tgcaagtgcc 60 gcctgtgctg gggaagggac catgtggctg ccttgggctc tgttgcttct ctgggtccca 120 ggatgttttg ctctgagcaa atgcaggacc gtggcgggcc ccgtgggggg atccctgagt 180 gtgcagtgtc cctatgagaa ggaacacagg accctcaaca aatactggtg cagaccacca 240 cagattttcc tatgtgacaa gattgtggag accaaagggt cagcaggaaa aaggaacggc 300 cgagtgtcca tcagggacag tcctgcaaac ctcagcttca cagtgaccct ggagaatctc 360 acagaggagg atgcaggcac ctactggtgt ggggtggata caccatggct ccgagacttt 420 catgatcccg ttgtcgaggt tgaggtgtcc gtgttcccgg catcaacgtc aatgacacct 480 gcaagtatca ctgcggccaa gacctcaaca atcacaactg catttccacc tgtatcatcc 540 actaccctgt ttgcagtggg tgccacccac agtgccagca tccaggagga aactgaggag 600 gtggtgaact cacagctccc gctgactcct ctccctgctg gcattgttgc tgcttctgtt 660 ggtgggggcc tccctgctag cctggaggat gtttcagaaa tggatcaaag ctggtgacca 720 ttcagagctg tcccagaacc ccaagcaggc tgccacgcag agtgagctgc actacgcaaa 780 tctggagctg ctgatgtggc ctctgcagga aaagccagca ccaccaaggg aggtggaggt 840 ggaatacagc actgtggcct cccccaggga agaacttcac tatgcctcgg tggtgtttg 899 <210> 24 <211> 953 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 3887384CB1 <400> 24 cgcgggggtc gcgctgcaca gcctgcggcg cagcggaggc ggaccgcagt cgagtctgca 60 gagtgttgga atcgtccgca ctaaagtccc ctgcagcgtg accatgtcga gaccccggaa 120 gaggctggct gggacttctg gttcagacaa gggactatca ggaaaacgca ccaaaactga 180 gaactcaggt gaggcattag ctaaagtgga ggactccaac cctcagaaga cttcagccac 240 taaaaactgt ttgaagaatc taagcagcca ctggctgatg aagtcagagc cagagagccg 300 cctagagaaa ggtgtagatg tgaagttcag cattgaggat ctcaaagcac agcccaaaca 360 gacaacatgc tgggatggtg ttcgtaacta ccaggctcgg aacttcctta gagccatgaa 420 gctgggagaa gaagccttct tctaccatag caactgcaaa gagccaggca tcgcaggact 480 catgaagatc gtgaaagagg cttacccaga ccacacacag tttgagaaaa acaatcccca 540 ttatgaccca tctagcaaag aggacaaccc taagtggtcc atggtggatg tacagtttgt 600 tcggatgatg aaacgtttca ttcccctggc tgagctcaaa tcctatcatc aagctcacaa 660 agctactggt ggccccttaa aaaatatggt tctcttcact cgccagagat tatcaatcca 720 gcccctgacc caggaagagt ttgattttgt tttgagcctg gaggaaaagg aaccaagtta 780 actgagatac tgctgctgga atgggcgaga cattgctgca aagaagtcaa gcttttttca 840 gacaaaaggt gtgagggggc ttgcttggta tgcttacctg ggcttgtgta cctcagtggt 900 ttttgtgtac ttttttcaat aaaatatcaa agttgaagaa aaaaaaaaaa aaa 953 <210> 25 <211> 1979 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 1352789CB1 <400> 25 ttctcaacat cctggcttag tattgtgtgc aaaatcagag aggggtgcaa gatcctgatt 60 tttcaggagt tcaagcgaca atggcagccc aatacggcag tatgagcttc aaccccagca 120 caccaggggc cagttatggg cctggaaggc aagagcccag aaattcccaa ttgagaattg 180 tgttagtggg taaaaccgga gcaggaaaaa gtgcaacagg aaacagcatc cttggccgga 240 aagtgtttca ttctggcact gcagcaaaat ccattaccaa gaagtgtgag aaacgcagca 300 gctcatggaa ggaaacagaa cttgtcgtag ttgacacacc aggcattttc gacacagagg 360 tgcccaatgc tgaaacgtcc aaggagatta ttcgctgcat tcttctgacc tccccagggc 420 ctcatgctct gcttctggtg gttccactgg gccgttacac tgaggaagag cacaaagcca 480 cagagaagat cctgaaaatg tttggagaga gggctagaag tttcatgatt ctcatattca 540 cccggaaaga tgacttaggt gacaccaatt tgcatgacta cttaagggaa gctccagaag 600 acattcaaga cttgatggac attttcggtg accgctactg tgcgttaaac aacaaggcaa 660 caggcgctga gcaggaggcc cagagggcac agttgctggg cctgatccag cgcgtggtga 720 gggagaacaa ggaaggctgc tacactaata ggatgtacca aagggcggag gaggagatcc 780 agaagcaaac acaagcaatg caagaactcc acagagtgga gctggagaga gagaaagcgc 840 ggataagaga ggagtatgaa gagaaaatca gaaagctgga agataaagtg gagcaggaaa 900 agagaaagaa gcaaatggag aagaaactag cagaacagga ggctcactat gctgtaaggc 960 agcaaagggc aagaacggaa gtggagagta aggatgggat acttgaatta atcatgacag 1020 cgttacagat tgcttccttt attttgttac gtctgttcgc ggaagattaa acttaatgaa 1080 aatctgtttg tattttctgc atattctctg gcaaccttgc cccatactta cttatttagc 1140 atagtcgagt gctctagttt ctgtctctca ggcactcgta actaaggacc accattggcc 1200 attggtagat gtttgattga cttaacaaga gagggacaaa ttttcaattt gtgaaactcc 1260 aaagcagaaa gtattggtgc ttgctacctt gtgaattctt ccttagacat gcagagaaaa 1320 tgtatgcaag agaccaaaaa gatggctcca agctatgtca tgttacctgt aataaaatct 1380 tttcttctag attctttcta tgttggcaga taatctcccc ttgtagcttc cactcactta 1440 ttcttgcatt cagagtcaca atgatcatct tacccatgtg gtttttgaga aagaaagatc 1500 aattctttgt ttgcagtagg taatcttaga gatggagatg attgtagaat tattcctaga 1560 tgagtgtcaa tttatttaat tccattgtca tataaggagt caaattgttt cttatcattt 1620 gttcattgaa gaacagagac ctgtctggaa aatcgatctc tacaaattca attaaataat 1680 gatccccaaa tgctgaaaaa gtgaaataca gcaattcaac agataataga gcaatgttta 1740 gtatattcag ctgtatctgt agaaactctt tgacgaacct caatttaacc aatttgatga 1800 atacccagtt ctcttctttt ctagagaaag atagttgcaa cctcacctcc ctcactcaac 1860 actttgaata cttattgttt ggcaggtcat ccacacactt ctgcccccac tgcattgaat 1920 tttttgctta tgttgtttat aataaaactt ttcaattatc tcataaaaaa aaaaaaaaa 1979 <210> 26 <211> 923 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No.: 1666486CT1 <400> 26 cccagtcagg acacagcatg gacatgaggg tccccgctca gctcctgggg ctcctgctgc 60 tctggctccc aggtgccaaa tgtgacatcc tgctgaccca gtctccttcc accgtctctg 120 catctgtggg cgacagagtc accattactt gccgggccac tcagagtatt ggtagctggg 180 tggcctggta tcagcaaaaa ccagggaaag cccctcagct cctgatctat aaggcgtcca 240 gtttagaaag tggagtccca tcaaggttta gtggcagtgg atctgggaca gaattcactc 300 tcagcatcaa cagcctgcag cctgatgatt ttgcaactta tttctgccag cagtatgaca 360 cttaccccac gtggtcgttc ggccagggga ccaagctgga gatcaaacga actgtggctg 420 caccatctgt cttcatcttc ccgccatctg atgagcagtt gaaatctgga actgcctctg 480 ttgtgtgcct gctgaataac ttctatccca gagaggccaa agtacagtgg aaggtggata 540 acgccctcca atcgggtaac tcccaggaga gtgtcacaga gcaggacagc aaggacagca 600 cctacagcct cagcagcacc ctgacgctga gcaaagcaga ctacgagaaa cacaaactct 660 acgcctgcga agtcacccat cagggcctga gctcgcccgt cacaaagagc ttcaacaggg 720 gagagtgtta gagggagaag tgcccccacc tgctcctcag ttccagcctg accccctccc 780 atcctttggc ctctgaccct ttttccacag gggacctacc cctattgcgg tcctccagct 840 catctttcac ctcacccccc tcctcctcct tggctttaat tatgctaatg ttggaggaga 900 atgaataaat aaagtgaatc ttc 923 <210> 27 <211> 888 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No.: 1706596CB1 <400> 27 cagcaagcat agtgggaatc tgcaccatgc cctgggctct gctcctcctg accctcctca 60 ctcactctgc agtgtcagtg gtccaggcag ggctgactca gccaccctcg gtgtccaggg 120 ccttgagaca gaccgccaca ctcacctgca ctgggaacaa caacaatgtt ggcaaccaag 180 gagcagcttg gctgcagcag caccagggcc accctcccaa actcctgtcc tacaggaata 240 acaaccggcc ctcagggatc tcagagagat tctctgcatc caggtcgaga aacacagcct 300 ccctgaccat tactggactc cagcctgagg acgaggctga ctattactgc tcagtatggg 360 acagcagcct cagtgcttgg gtgttcggcg gagggaccaa gctgaccgtc ctaagtcagc 420 ccaaggctgc cccctcggtc actctgttcc caccctcctc tgaggagctt caagccaaca 480 aggccacact ggtgtgtctc ataagtgact tctacccggg agccgtgaca gtggcctgga 540 aggcagatag cagccccgtc aaggcgggag tggagaccac cacaccctcc aaacaaagca 600 acaacaagta cgcggccagc agctacctga gcctgacgcc tgagcagtgg aagtcccaca 660 gaagctacag ctgccaggtc acgcatgaag ggagcaccgt ggagaagaca gtggccccta 720 cagaatgttc ataggttctc aaccctcacc ccccaccacg ggagactaga gctgcaggat 780 cccaggggag gggtctctcc tcccacccca aggcatcaag cccttctccc tgcactcaat 840 aaaccctcaa taaatattct cattgtcaat cagaaaaaaa aaaaaaaa 888 <210> 28 <211> 1760 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No.: 1890540CB1 <400> 28 ccaaagtata aaaccactcc gccgctgcag ctctcagctc cagtcctggc atctgcccga 60 ggagaccacg ctcctggagc tctgctgtct tctcagggag actctgaggc tctgttgaga 120 atcatgcttt ggaggcagct catctattgg caactgctgg ctttgttttt cctccctttt 180 tgcctgtgtc aagatgaata catggagtct ccacaaaccg gaggactacc cccagactgc 240 agtaagtgtt gtcatggaga ctacagcttt cgaggctacc aaggcccccc tgggccaccg 300 ggccctcctg gcattccagg aaaccatgga aacaatggca acaatggagc cactggtcat 360 gaaggagcca aaggtgagaa gggcgacaaa ggtgacctgg ggcctcgagg ggagcggggg 420 cagcatggcc ccaaaggaga gaagggctac ccggggattc caccagaact tcagattgca 480 ttcatggctt ctctggcaac ccacttcagc aatcagaaca gtgggattat cttcagcagt 540 gttgagacca acattggaaa cttctttgat gtcatgactg gtagatttgg ggccccagta 600 tcaggtgtgt atttcttcac cttcagcatg atgaagcatg aggatgttga ggaagtgtat 660 gtgtacctta tgcacaatgg caacacagtc ttcagcatgt acagctatga aatgaagggc 720 aaatcagata catccagcaa tcatgctgtg ctgaagctag ccaaagggga tgaggtttgg 780 ctgcgaatgg gcaatggcgc tctccatggg gaccaccaac gcttctccac ctttgcagga 840 ttcctgctct ttgaaactaa gtaaatatat gactagaata gctccacttt ggggaagact 900 tgtagctgag ctgatttgtt acgatctgag gaacattaaa gttgagggtt ttacattgct 960 gtattcaaaa aattattggt tgcaatgttg ttcacgctac aggtacacca ataatgttgg 1020 acaattcagg ggctcagaag aatcaaccac aaaatagtct tctcagatga ccttgactaa 1080 tatactcagc atctttatca ctctttcctt ggcacctaaa agataattct cctctgacgc 1140 aggttggaaa tatttttttc tatcacagaa gtcatttgca aagaattttg actactctgc 1200 ttttaattta ataccagttt tcaggaaccc ctgaagtttt aagttcatta ttctttataa 1260 catttgagag aatcagatgt agtgatatga cagggctggg gcaagaacag gggcactagc 1320 tgccttatta gctaatttag tgccctccgt gttcagctta gcctttgacc ctttcctttt 1380 gatccacaaa atacattaaa actctgaatt cacatacaat gctattttaa agtcaataga 1440 ttttagctat aaagtgcttg accagtaatg tggttgtaat tttgtgtatg ttcccccaca 1500 tcgcccccaa cttcggatgt ggggtcagga ggttgaggtt cactattaac aaatgtcata 1560 aatatctcat agaggtacag tgccaataga tattcaaatg ttgcatgttg accagaggga 1620 ttttatatct gaagaacata cactattaat aaatacctta gagaaagatt ttgacctggc 1680 tttagataaa actgtggcaa gaaaaatgta atgagcaata tatggaaata aacacacctt 1740 tgttaaagat aaaaaaaaaa 1760 <210> 29 <211> 2015 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No.: 2774913CB1 <400> 29 gcttcagccg cagtcgccac tggctgcctg aggtgctctt acagcctgtt ccaagtgtgg 60 cttaatccgt ctccaccacc agatctttct ccgtggattc ctctgctaag accgctgcca 120 tgccagtgac ggtaacccgc accaccatca caaccaccac gacgtcatct tcgggcctgg 180 ggtcccccat gatcgtgggg tcccctcggg ccctgacaca gcccctgggt ctccttcgcc 240 tgctgcagct ggtgtctacc tgcgtggcct tctcgctggt ggctagcgtg ggcgcctgga 300 cggggtccat gggcaactgg tccatgttca cctggtgctt ctgcttctcc gtgaccctga 360 tcatcctcat cgtggagctg tgcgggctcc aggcccgctt ccccctgtct tggcgcaact 420 tccccatcac cttcgcctgc tatgcggccc tcttctgcct ctcggcctcc atcatctacc 480 ccaccaccta tgtccagttc ctgtcccacg gccgttcgcg ggaccacgcc atcgccgcca 540 ccttcttctc ctgcatcgcg tgtgtggctt acgccaccga agtggcctgg acccgggccc 600 ggcccggcga gatcactggc tatatggcca ccgtacccgg gctgctgaag gtgctggaga 660 ccttcgttgc ctgcatcatc ttcgcgttca tcagcgaccc caacctgtac cagcaccagc 720 cggccctgga gtggtgcgtg gcggtgtacg ccatctgctt catcctagcg gccatcgcca 780 tcctgctgaa cctgggggag tgcaccaacg tgctacccat ccccttcccc agcttcctgt 840 cggggctggc cttgctgtct gtcctcctct atgccaccgc ccttgttctc tggcccctct 900 accagttcga tgagaagtat ggcggccagc ctcggcgctc gagagatgta agctgcagcc 960 gcagccatgc ctactacgtg tgtgcctggg accgccgact ggctgtggcc atcctgacgg 1020 ccatcaacct actggcgtat gtggctgacc tggtgcactc tgcccacctg gtttttgtca 1080 aggtctaaga ctctcccaag aggctcccgt tccctctcca acctctttgt tcttcttgcc 1140 cgagttttct ttatggagta cttctttcct ccgcctttcc tctgttttcc tcttcctgtc 1200 tcccctccct cccacctttt tctttccttc ccaattcctt gcactctaac cagttcttgg 1260 atgcatcttc ttccttccct ttcctcttgc tgtttccttc ctgtgttgtt ttgttgccca 1320 catcctgttt tcacccctga gctgtttctc tttttctttt ctttcttttt tttttttttt 1380 tttaagacgg attctcacca ctgtgctcca gcctggggga cagagcgaga ctccatctca 1440 aaaaaaaaaa ggaatcggac gaagaaccac aggatgttga agacaactgt ctgaagtatt 1500 tgtgagggac agcgatgtgg ccctctgtgt taagaataac gtgtcctgct ttggcagaga 1560 gaagaaaata gccactgccc gctttcaagg caagatcgac cttttctgtt ttgttttgtt 1620 tttctttctt tttcctggcc atgaggacaa aaattactga gtggccctta aagagggaag 1680 tttgttttca gctgttctct tttgcccgta ggtgggaggg tggggattgc tgcgtcctag 1740 ctagaggaat ggctttgctt gaatgtgtag tgcacacgca cgggtgtttc tgtgtgctag 1800 ttgcttcttg ctgctgcttc ctgcttgtct gggactcaca tacataacgt gatatatata 1860 tatatatata aatgtataaa tatatatttt attttttttt aaatccttgg agcttctggt 1920 tcctatcagt tcctgttgtt aatcgtagaa ccgttgtccc ttcccccatt cccgtatcca 1980 tcatgttctt tttcttttaa atatcaatat aaaaa 2015 <210> 30 <211> 2080 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No.: 5571291CB1 <400> 30 gctgcagtga accgagattg cgccactgca ctccagccta ggcaacaaag cgagactctg 60 tctcaaaact aaaaaataat aaaaaataaa taaacctcta taaagtatac caagtcttag 120 tttttaaatt aagagataag tgtggatttg ttttccaaag gtgaataagc tttgtttttt 180 ccagacaaaa gcaagccggg aggctggctg cctctcctcc tgctgtctct gctggtggcc 240 acatgggtgc tggtggcagg gatctatcta atgtggaggc acgaaaggat caagaagact 300 tccttttcta ccaccacact actgcccccc attaaggttc ttgtggttta cccatctgaa 360 atatgtttcc atcacacaat ttgttacttc actgaatttc ttcaaaacca ttgcagaagt 420 gaggtcatcc ttgaaaagtg gcagaaaaag aaaatagcag agatgggtcc agtgcagtgg 480 cttgccactc aaaagaaggc agcagacaaa gtcgtcttcc ttctttccaa tgacgtcaac 540 agtgtgtgcg atggtacctg tggcaagagc gagggcagtc ccagtgagaa ctctcaagac 600 ctcttccccc ttgcctttaa ccttttctgc agtgatctaa gaagccagat tcatctgcac 660 aaatacgtgg tggtctactt tagagagatt gatacaaaag acgattacaa tgctctcagt 720 gtctgcccca agtaccacct catgaaggat gccactgctt tctgtgcaga acttctccat 780 gtcaagcagc aggtgtcagc aggaaaaaga tcacaagcct gccacgatgg ctgctgctcc 840 ttgtagccca cccatgagaa gcaagagacc ttaaaggctt cctatcccac caattacagg 900 gaaaaaacgt gtgatgatcc tgaagcttac tatgcagcct acaaacagcc ttagtaatta 960 aaacatttta taccaataaa attttcaaat attgctaact aatgtagcat taactaacga 1020 ttggaaacta catttacaac ttcaaagctg ttttatacat agaaatcaat-tacagtttta 1080 attgaaaact ataaccattt tgataatgca acaataaagc atcttcagcc aaacatctag 1140 tcttccatag accatgcatt gcagtgtacc cagaactgtt tagctaatat tctatgttta 1200 attaatgaat actaactcta agaacccctc actgattcac tcaatagcat cttaagtgaa 1260 aaaccttcta ttacatgcaa aaaatcattg tttttaagat aacaaaagta gggaataaac 1320 aagctgaacc cacttttact ggaccaaatg atctattata tgtgtaacca cttgtatgat 1380 ttggtatttg cataagacct tccctctaca aactagattc atatcttgat tcttgtacag 1440 gtgcctttta acatgaacaa caaaataccc acaaacttgt ctacttttgc ctaaagttac 1500 ctattagagg tcactgtcag agttctcagt ttcttagtta ctatttaact tttcatgttc 1560 aaaatgaaaa taattcttaa gttgaaagcc ctcttgaagt aaccttttta taaatgagtt 1620 attataatgg tttacttaaa taaaaaacag gggtgggtgc agtggctcat gcctccaatc 1680 ccagcacttt ggcaaggcca aggcaaaagg atcgctcaag accaggctac gtcacaaagc 1740 gagacctcca tctctacaaa agatttaaaa aattagctga gtgtgatggt gtaagcctgt 1800 ggtcccagct actagggagg ctgagatggg aggatcactt gagccctgga ggtcaagggt 1860 gcagtaaacg gtgattgtgc cactgcactc catcctgggt gagagcagac cctgtctaaa 1920 acaaacaaac gaaaaaaccc ccacagaatg acagaacata aaagatgcac attttgtctt 1980 cccacttttt tactcttcta aaggcatctt tttttaaatt ttcttaaatt tttttttttt 2040 tgagacagag gttcacccgg tcacccaggc tggaggtgcg 2080

Claims (23)

What is claimed is:

1. An isolated polypeptide comprising an amino acid sequence selected from the group consisting of:
a) an amino acid sequence selected from the group consisting of SEQ ID NO:1-15, b) a naturally occurring amino acid sequence having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO:1-15, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-15, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-15.

2. An isolated polypeptide of claim 1 selected from the group consisting of SEQ ID NO:1-15.

3. An isolated polynucleotide encoding a polypeptide of claim 1.

4. An isolated polynucleotide of claim 3 selected from the group consisting of SEQ ID
NO:16-30.

5. A recombinant polynucleotide comprising a promoter sequence operably linked to a polynucleotide of claim 3.

6. A cell transformed with a recombinant polynucleotide of claim 5.

7. A transgenic organism comprising a recombinant polynucleotide of claim 5.

8. A method for producing a polypeptide of claim 1, the method comprising:
a) culturing a cell under conditions suitable for expression of the polypeptide, wherein said cell is transformed with a recombinant polynucleotide, and said recombinant polynucleotide comprises a promoter sequence operably linked to a polynucleotide encoding the polypeptide of claim 1, and b) recovering the polypeptide so expressed.

9. An isolated antibody which specifically binds to a polypeptide of claim 1.

10. An isolated polynucleotide comprising a polynucleotide sequence selected from the group consisting of:
a) a polynucleotide sequence selected from the group consisting of SEQ ID
NO:16-30, b) a naturally occurring polynucleotide sequence having at least 90% sequence identity to a polynucleotide sequence selected from the group consisting of SEQ ID NO:16-30, c) a polynucleotide sequence complementary to a), d) a polynucleotide sequence complementary to b), and e) an RNA equivalent of a)-d).

11. An isolated polynucleotide comprising at least 60 contiguous nucleotides of a polynucleotide of claim 10.

12. A method for detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide of claim 10, the method comprising:
a) hybridizing the sample with a probe comprising at least 16 contiguous nucleotides comprising a sequence complementary to said target polynucleotide in the sample, and which probe specifically hybridizes to said target polynucleotide, under conditions whereby a hybridization complex is formed between said probe and said target polynucleotide, and b) detecting the presence or absence of said hybridization complex, and, optionally, if present, the amount thereof.

13. A method of claim 12, wherein the probe comprises at least 30 contiguous nucleotides.

14. A method of claim 12, wherein the probe comprises at least 60 contiguous nucleotides.

15. A pharmaceutical composition comprising an effective amount of a polypeptide of claim 1 and a pharmaceutically acceptable excipient.

16. A method for treating a disease or condition associated with decreased expression of functional IMOL, comprising administering to a patient in need of such treatment the pharmaceutical composition of claim 15.

17. A method for screening a compound for effectiveness as an agonist of a polypeptide of claim 1, the method comprising:
a) exposing a sample comprising a polypeptide of claim 1 to a compound, and b) detecting agonist activity in the sample.

18. A pharmaceutical composition comprising an agonist compound identified by a method of claim 17 and a pharmaceutically acceptable excipient.

19. A method for treating a disease or condition associated with decreased expression of functional IMOL, comprising administering to a patient in need of such treatment a pharmaceutical composition of claim 18.

20. A method for screening a compound for effectiveness as an antagonist of a polypeptide of claim 1, the method comprising:
a) exposing a sample comprising a polypeptide of claim 1 to a compound, and b) detecting antagonist activity in the sample.

21. A pharmaceutical composition comprising an antagonist compound identified by a method of claim 20 and a pharmaceutically acceptable excipient.

22. A method for treating a disease or condition associated with overexpression of functional IMOL, comprising administering to a patient in need of such treatment a pharmaceutical composition of claim 21.

23. A method for screening a compound for effectiveness in altering expression of a target polynucleotide, wherein said target polynucleotide comprises a sequence of claim 4, the method comprising:
a) exposing a sample comprising the target polynucleotide to a compound, and b) detecting altered expression of the target polynucleotide.