CA2395007A1 - Transporters and ion channels - Google Patents

Abstract

The invention provides human transporters and ion channels (TRICH) and polynucleotides which identify and encode TRICH. 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 aberrant expression of TRICH.

Description

TRANSPORTERS AND ION CHANNELS
TECHNICAL FIELD
This invention relates to nucleic acid and amino acid sequences of transporters and ion channels and to the use of these sequences in the diagnosis, treatment, and prevention of transport, neurological, muscle, and immunological disorders, and in the assessment of the effects of exogenous compounds on the expression of nucleic acid and amino acid sequences of transporters and ion channels.
BACKGROUND OF THE INVENTION
Eukaryotic cells are surrounded and subdivided into functionally distlrict organelles by hydrophobic lipid bilayer membranes which are highly impermeable to most polar molecules. Cells and organelles require transport proteins to import and export essential nutrients and metal ions including K+, NH4+, P;, S042-, sugars, and vitamins, as well as various metabolic waste products. Transport proteins also play roles in antibiotic resistance, toxin secretion, ion balance, synaptic neurotransmission, kidney function, intestinal absorption, tumor growth, and other diverse cell functions (Griffith, J. and C.
Sansom (1998) The TransporterFacts Book, Academic Press, San Diego CA, pp. 3-29). Transport can occur by a passive concentration-dependent mechanism, or can be linked to an energy source such as ATP hydrolysis or an ion gradient. Proteins that function in transport include carrier proteins, which bind to a specific solute and undergo a conformational change that translocates the bound solute across the membrane, and channel proteins, which form hydrophilic pores that allow specific solutes to diffuse through the membrane down an electrochemical solute gradient.
Carrier proteins which transport a single solute from one side of the membrane to the other are called uniporters. In contrast, coupled transporters link the transfer of one solute with simultaneous or sequential transfer of a second solute, either in the same direction (symport) or in the opposite direction (antiport). For example, intestinal and kidney epithelium contains a variety of symporter systems driven by the sodium gradient that exists across the plasma membrane. Sodium moves into the cell down its electrochemical gradient and brings the solute into the cell with it. The sodium gradient that provides the driving force for solute uptake is maintained by the ubiquitous Na+lK+ ATPase system. Sodium-coupled transporters include the mammalian glucose transporter (SGLT1), iodide transporter (NIS), and multivitamin transporter (SMVT). All three transporters have twelve putative transmembrane segments, extracellular glycosylation sites, and cytoplasmically-oriented N- and C-termini. NIS plays a crucial role in the evaluation, diagnosis, and treatment of various thyroid pathologies because it is the molecular basis for radioiodide thyroid-imaging techniques and for specific targeting of radioisotopes to the thyroid gland (Levy, O: et al. (1997) Proc.

Natl. Acad. Sci, USA 94:5568-5573). SMVT is expressed in the intestinal mucosa, kidney, and placenta, and is implicated in the transport of the water-soluble vitamins, e.g., biotin and pantothenate (Prasad, P.D. et al. (1998) J. Biol. Chem. 273:7501-7506).
One of the largest families of transporters is the major facilitator superfamily (MFS), also called the uniporter-symporter-antiporter family. MFS transporters are single polypeptide carriers that transport small solutes in response to ion gradients. Members of the MFS
are found in all classes of living organisms, and include transporters for sugars, oligosaccharides, phosphates, nitrates, nucleosides, monocarboxylates, and drugs. MFS transporters found in eukaryotes all have a structure comprising 12 transmembrane segments (Pao, S.S. et al. (1998) Microbiol.
Molec. Biol. Rev. 62:1-34). The largest family of MFS transporters is the sugar transporter family, which includes the seven glucose transporters (GLUTI-GLUT7) found in humans that are required for the transport of glucose and other hexose sugars.. These glucose transport proteins have unique tissue distributions! and physiological functions. GLUT1 provides many, cell types with their basal glucose requirements and transports glucose across epithelial and endothelial barrier tissues; GLUT2 facilitates glucose uptake or efflux from the liver; GLUT3 regulates glucose supply to neurons; GLUT4 is responsible.for insulin-regulated glucose disposal; and GLUTS regulates fructose uptake into skeletal muscle.
Defects in glucose transporters are involved in a recently identified neurological syndrome causing infantile seizures and developmental delay, as well as glycogen storage disease, Fanconi-Bickel syndrome, and non-insulin-dependent diabetes mellitus (Mueckler, M. (1994) Eur. J. Biochem.
219:713-725; Longo, N. and L.J. Elsas (1998) Adv. Pediatr. 45:293-313).
Monocarboxylate anion transporters are proton-coupled symporters with a broad substrate specificity that includes L-lactate, pyruvate, and the ketone bodies acetate, acetoacetate, and beta-hydroxybutyrate. At least seven isoforms have been identified to date.
The isoforms are predicted to have twelve transmembrane (TM) helical domains with a large intracellular loop between TM6 and TM7, and play a critical role in maintaining intracellular pH by removing the protons that axe produced stoichiometrically with lactate during glycolysis. The best characterized H+-monocarboxylate transporter is that of the erythrocyte membrane, which transports L-lactate and a wide range of other aliphatic monocarboxylates. Other cells possess H+-linked monocarboxylate transporters with differing substrate and inhibitor selectivities. In particular, cardiac muscle and tumor cells have transporters that differ in their Km values for certain substrates, including stereoselectivity for L- over D-lactate, and in their sensitivity to inhibitors. There are Na+-monocarboxylate cotransporters on the luminal surface of intestinal and kidney epithelia, which allow the uptake of lactate, pyruvate, and ketone bodies in these tissues. In addition, there are specific and selective transporters for organic cations and organic anions in organs including the kidney, intestine and liver. Organic anion transporters are selective for hydrophobic, charged molecules with electron-attracting side groups. Organic canon transporters, such as the ammonium transporter, mediate the secretion of a variety of drugs and endogenous metabolites, and contribute to the maintenance of intercellular pH (Poole, R.C. and A.P.
Halestrap (1993) Am. J.
Physiol. 264:C761-C782; Price, N.T. et al. (1998) Biochem. J. 329:321-328; and Martinelle, K. and I.
Haggstrom (1993) J. Biotechnol. 30:339-3.50).
ATP-binding cassette (ABC) transporters are members of a superfamily of membrane proteins that transport substances ranging from small molecules such as ions, sugars, amino acids, peptides, and phospholipids, to lipopeptides, large proteins, and complex hydrophobic drugs.
ABC transporters consist of four modules: two nucleotide-binding domains (NBD), which hydrolyze ATP to supply the energy required for transport, and two membrane-spanning domains (MSD), each containing six putative transmembrane segments. These four modules may be encoded by a single gene, as is the case for the cystic fibrosis transmembrane regulator (CFTR), or by separate genes.
When encoded by separate genes, each gene product contains a single NBD and MSD. These "half molecules" form homo- and heterodimers, such as Tapl and Tap2, the endoplasmic reticulum based major histocompatibility (MHC) peptide transport system. Several genetic diseases are attributed to defects in ABC transporters, such as the following diseases and their corresponding proteins: cystic fibrosis (CFTR, an ion channel), adrenoleukodystrophy (adrenoleukodystrophy protein, ALDP), 2ellweger syndrome (peroxisomal membrane protein-70, PMP70), and hyperinsulinemic hypoglycemia (sulfonylurea receptor, SUR). Overexpression of the multidrug resistance (MDR) protein, another ABC transporter, in human cancer cells makes the cells resistant to a variety of cytotoxic drugs used in chemotherapy (Taglicht, D. and S. Michaelis (1998) Meth. Enzymol. 292:130-162).
A number of metal ions such as iron, zinc, copper, cobalt, manganese, molybdenum, selenium, nickel, and chromium are important as cofactors for a number of enzymes. For example, copper is involved in hemoglobin synthesis, connective tissue metabolism, and bone development, by acting as a cofactor in oxidoreductases such as superoxide dismutase, ferroxidase (ceruloplasmin), and lysyl oxidase. Copper and other metal ions must be provided in the diet, and are absorbed by transporters in the gastrointestinal tract. Plasma proteins transport the metal ions to the liver and other target organs, where specific transporters move the ions into cells and cellular organelles as needed. Imbalances in metal ion metabolism have been associated with a number of disease states (banks, D.M. (1986) J.
Med. Genet. 23:99-106).
Transport of fatty acids across the plasma membrane can occur by diffusion, a high capacity, low affinity process. However, under normal physiological conditions a significant fraction of fatty acid transport appears to occur via a high affinity, low capacity protein-mediated transport process.
Fatty acid transport protein (FATP), an integral membrane protein with four transmembrane segments, is expressed in tissues exhibiting high levels of plasma membrane fatty acid flux, such as muscle, heart, and adipose. Expression of FATP is upregulated in 3T3-L1 cells during adipose conversion, and expression in COS7 fibroblasts elevates uptake of long-chain fatty acids (Hui, T.Y. et al. (1998) J.
Biol. Chem. 273:27420-27429).
Mitochondrial carrier proteins are transmembrane-spanning proteins which transport ions and charged metabolites between the cytosol and the mitochondrial matrix. Examples include the ADP, ATP carrier protein; the 2-oxoglutaratelmalate carrier; the phosphate carrier protein; the pyruvate carrier; the dicarboxylate carrier which transports malate, succinate, fumarate, and phosphate; the tricarboxylate carrier which transports citrate and malate; and the Grave's disease carrier protein, a protein recognized by IgG in patients with active Grave's disease, an autoimmune disorder resulting in hyperthyroidism. Proteins in this family consist of three tandem repeats of an approximately 100 amino acid domain, each of which contains two transmembrane regions (Stryer, L.
(1995) Biochemistry, W.H.
Freeman and Company, New York NY, p. 551; PROSITE PDOC00189 Mitochotidrial energy transfer proteins signature; Online Mendelian Inheritance in Man (OMIM) *275000 Graves Disease).
This class of transporters also includes the mitochondrial uncoupling proteins, which create proton leaks across the inner mitochondrial membrane, thus uncoupling oxidative phosphorylation from ATP synthesis. The result is energy dissipation in the form of heat.
Mitochondrial uncoupling proteins have been implicated as modulators of thermoregulation and metabolic rate, and have been proposed as potential targets for drugs against metabolic diseases such as obesity (Ricquier, D. et al. (1999) J. Int.
Med.245:637-642).
Ion Channels The electrical potential of a cell is generated and maintained by controlling the movement of ions across the plasma membrane. The movement of ions requires ion channels, which form ion-selective pores within the membrane. There are two basic types of ion channels, ion transporters and gated ion channels. Ion transporters utilize the energy obtained from ATP
hydrolysis to actively transport an ion against the ion's concentration gradient. Gated ion channels allow passive flow of an ion down the ion's electrochemical gradient under restricted conditions.
Together, these types of ion channels generate, maintain, and utilize an electrochemical gradient that is used in 1) electrical impulse conduction down the axon of a nerve cell, 2) transport of molecules into cells against concentration gradients, 3) initiation of muscle contraction, and 4) endocrine cell secretion.
Ion Transporters Ion transporters generate and maintain the resting electrical potential of a cell. Utilizing the energy derived from ATP hydrolysis, they transport ions against the ion's concentration gradient.
These transmembrane ATPases are divided into three families. The phosphorylated (P) class ion transporters, including Na+-K+ ATPase, Ca2+-ATPase, and H+-ATPase, are activated by a phosphorylation event. P-class ion transporters are responsible for maintaining resting potential distributions such that cytosolic concentrations of Nay and Ca2+ are low and cytosolic concentration of K+ is high. The vacuolar (V) class of ion transporters includes H+ pumps on intracellular organelles, such as lysosomes and Golgi. V-class ion transporters are responsible for generating the low pH within the lumen of these organelles that is required for function. The coupling factor (F) class consists of H+
pumps in the mitochondria. F-class ion transporters utilize a proton gradient to generate ATP from ADP and inorganic phosphate (Pi).
The P-ATPases are hexamers of a 100 kD subunit with ten transmembrane domains and several large cytoplasmic regions that may play a role in ion binding (Scarborough, G.A. (1999) Curr.
Opin. Cell Biol. 11:517-522). The V-ATPases are composed of two functional domains: the V 1 domain, a peripheral complex responsible for ATP hydrolysis; and the Vo domain, an integral .
complex responsible for proton translocation across the membrane. The F-ATPases are structurally and evolutionarily related to the V-ATPases. The F-ATPase Fo domain contains 12 copies of the c subunit, a highly hydrophobic protein composed of two transmembrane domains and containing a single buried carboxyl group in TM2 that is essential for proton transport.
The V-ATPase V o domain contains three types of homologous c subunits with four or five transmembrane domains and the essential carboxyl group in TM4 or TM3. Both types of complex also contain a single a subunit that may be involved in regulating the pH dependence of activity (Forgac, M. (1999) J. Biol. Chem.
274:12951-12954).
The resting potential of the cell is utilized in many processes involving carrier proteins and gated ion channels. Carrier proteins utilize the resting potential to transport molecules into and out of the cell. Amino acid and glucose transport into many cells is linked to sodium ion co-transport (symport) so that the movement of Na+ down an electrochemical gradient drives transport of the other molecule up a concentration gradient. Similarly, cardiac muscle links transfer of Ca2+ out of the cell with transport of Na+ into the cell (antiport).
Gated Ion Channels Gated ion channels control ion flow by regulating the opening and closing of pores. The ability to control ion flux through various gating mechanisms allows ion channels to mediate such diverse signaling and homeostatic functions as neuronal and endocrine signaling, muscle contraction, fertilization, and regulation of ion and pH balance. Gated ion channels are categorized according to the manner of regulating the gating function. Mechanically-gated channels open their pores in response to mechanical stress; voltage-gated channels (e.g., Na+, K+, Ca2+, and Cl-channels) open their pores in response to changes in membrane potential; and ligand-gated channels (e.g., acetylcholine-, serotonin-, and glutamate-gated cation channels, and GABA- and glycine-gated chloride channels) open their pores in the presence of a specific ion, nucleotide, or neurotransmitter.
The gating properties of a particular ion channel (i.e., its threshold for and duration of opening and closing) are sometimes modulated by association with auxiliary channel proteins and/or post translational modifications, such as phosphorylation.
Mechanically-gated or mechanosensitive ion channels act as transducers for the senses of touch, hearing, and balance, and also play important roles in cell volume regulation, smooth muscle contraction, and cardiac rhythm generation. A stretch-inactivated channel (SIC) was recently cloned from rat kidney. The SIC channel belongs to a group of channels which are activated by pressure or stress on the cell membrane and conduct both Ca2+ and Na''- (Suzuki, M. et al.
(1999) J. Biol. Chem.
274:6330-6335).
The pore-forming subunits of the voltage-gated canon channels form a superfamily of ion channel proteins. The characteristic domain of these channel proteins comprises six transmembrane domains (S 1-S6), a pore-forming region (P) located between SS and S6, and intracellular amino and carboxy termini. In the Na+ and Ca2+ subfamilies, this domain is repeated four times, while in the K+
channel subfamily, each channel is formed from a tetramer of either identical or dissimilar subunits.
The P region contains information specifying the ion selectivity for the channel. In the case of K+
channels, a GYG tripeptide is involved in this selectivity (Ishii, T.M. et al.
(1997) Proc. Natl. Acad.
Sci. USA 94:11651-11656).
Voltage-gated Na+ and K+ channels are necessary for the function of electrically excitable cells, such as nerve arid muscle cells. Action potentials, which lead to neurotransmitter release and muscle contraction, arise from large, transient changes in the permeability of the membrane to Na+ and K+
ions. Depolarization of the membrane beyond the threshold level opens voltage-gated Na+ channels.
Sodium ions flow into the cell, further depolarizing the membrane and opening more voltage-gated Na+
channels, which propagates the depolarization down the length of the cell.
Depolarization also opens voltage-gated potassium channels. Consequently, potassium ions flow outward, which leads to repolarization of the membrane. Voltage-gated channels utilize charged residues in the fourth transmembrane segment (S4) to sense voltage change. The open state lasts only about 1 millisecond, at which time the channel spontaneously converts into an inactive state that cannot be opened irrespective of the membrane potential. Inactivation is mediated by the channel's N-terminus, which acts as a plug that closes the pore. The transition from an inactive to a closed state requires a return to resting potential.
Voltage-gated Na+ channels are heterotrimeric complexes composed of a 260 kDa pore-forming a subunit that associates with two smaller auxiliary subunits, (31 and (32.
The X32 subunit is a integral membrane glycoprotein that contains an extracellular Ig domain, and its association with a and (31 subunits correlates with increased functional expression of the channel, a change in its gating properties, as well as an increase in whole cell capacitance due to an increase in membrane surface area (Isom, L.L. et al. (1995) Cell 83:433-442).
Non voltage-gated Na+ channels include the members of the amiloride-sensitive Na+
channel/degenerin (NaC/DEG) family. Channel subunits of this family are thought to consist of two transmembrane domains flanking a long extracellular loop, with the amino and carboxyl termini located within the cell. The NaC/DEG family includes the epithelial Na+ channel (ENaC) involved in Na+
reabsorption in epithelia including the airway, distal colon, cortical collecting duct of the kidney, and exocrine duct glands. Mutations in ENaC result in pseudohypoaldosteronism type 1 and Liddle's syndrome (pseudohyperaldosteronism). The NaCIDEG family also includes the recently characterized H+-gated canon channels or acid-sensing ion channels (ASIC). ASIC subunits are expressed in the brain and form heteromultimeric Na+-permeable channels. These channels require acid pH fluctuations for activation. ASIC subunits show homology to the degenerins, a family of mechanically-gated channels originally isolated from C. elegans. Mutations in the degenerins cause neurodegeneration.
ASIC subunits may also have a role in neuronal funtion, or in pain perception, since tissue acidosis causes pain (Waldmann, R, and M. Lazdunski (1998) Curr. Opin: Neurobiol. 8:418-424; Eglen, R.M.
et al. (1999) Trends Pharmacol. Sci. 20:337-342).
K+ channels are located in all cell types, and may be regulated by voltage, ATP concentration, or second messengers such as Ca2+ and cAMP. In non-excitable tissue, K+
channels are involved in protein synthesis, control of endocrine secretions, and the maintenance of osmotic equilibrium across membranes. In neurons and other excitable cells, in addition to regulating action potentials and repolarizing membranes, K+ channels are responsible for setting resting membrane potential. The cytosol contains non-diffusible anions and, to balance this net negative charge, the cell contains a Na+-K+ pump and ion channels that provide the redistribution of Na+, K+, and Cl-.
The pump actively transports Na+ out of the cell and K+ into the cell in a 3:2 ratio. Ion channels in the plasma membrane allow K+ and Cl- to flow by passive diffusion. Because of the high negative charge within the cytosol, Cl - flows out of the cell. The flow of K+ is balanced by an electromotive force pulling K+ into the cell, and a K+ concentration gradient pushing K+ out of the cell. Thus, the resting membrane potential is primarily regulated by K+flow (SalkofF, L. and T. Jegla (1995) Neuron 15:489-492).
Potassium channel subunits of the Shaker-like superfamily all have the characteristic six transmembrane/1 pore domain structure. Four subunits combine as homo- or heterotetramers to form functional K channels. These pore-forming subunits also associate with various cytoplasmic J3 subunits that alter channel inactivation kinetics. The Shaker-like channel family includes the voltage-gated K+ channels as well as the delayed rectifier type channels such as the human ether-a-go-go related gene (HERD) associated with long QT, a cardiac dysrythmia syndrome (Curran, M.E. (1998) Curr. Opin. Biotechnol. 9:565-572; Kaczarowski, G.J. and M.L. Garcia (1999) Curr. Opin. Chem.
Biol. 3:448-458).
A second superfamily of K+ channels is composed of the inward rectifying channels (Kir).
Kir channels have the property of preferentially conducting I~ currents in the inward direction. These proteins consist of a single potassium selective pore domain and two transmembrane domains, which correspond to the fifth and sixth transmembrane domains of voltage-gated K+
channels. Kir subunits also associate as tetramers. The Kir family includes ROMK1, mutations in which lead to B artter syndrome, a renal tubular disorder. Kir channels are also involved in regulation of cardiac pacemaker activity, seizures and epilepsy, and insulin regulation (Doupnik, C.A. et al.
(1995) Curr. Opin.
Neurobiol. 5:268-277; Curran, supra).
The recently recognized TWIK K+ channel family includes the mammalian TWIK-l, and TASK proteins. Members of this family possess an overall structure with four transmembrane domains and two P domains. These proteins are probably involved in controlling the resting potential in a large set of cell types (Duprat, F. et al. (1997) EMBO J 16:5464-5471).
The voltage-gated Ca 2+ channels have been classified into several subtypes based upon their electrophysiological and pharmacological characteristics. L-type Ca 2+
channels are predominantly expressed in heart and skeletal muscle where they play an essential role in excitation-contraction coupling. T-type channels are important for cardiac pacemaker activity, while N-type and P/Q-type channels are involved in the control of neurotransmitter release in the central and peripheral nervous system. The L-type and N-type voltage-gated C a 2+ channels have been purified and, though their functions differ dramatically, they have similar subunit compositions. The channels are composed of three subunits. The al subunit forms the membrane pore and voltage sensor, while the a28 and (3 subunits modulate the voltage-dependence, gating properties, and the current amplitude of the channel.
These subunits are encoded by at least six al, one a~~, and four (3 genes. A
fourth subunit, 'y, has been identified in skeletal muscle (Walker, D. et al. (1998) J. Biol. Chem.
273:2361-2367; McCleskey, E.W.
(1994) Curr. Opin. Neurobiol. 4:304-312).
Chloride channels are necessary in endocrine secretion and in regulation of cytosolic and organelle pH. In secretory epithelial cells, Cl- enters the cell across a basolateral membrane through an Na+, K+/Cl- cotransporter, accumulating in the cell above its electrochemical equilibrium concentration.
Secretion of Cl- from the apical surface, in response to hormonal stimulation, leads to flow of Na+ and water into the secretory lumen. The cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride channel encoded by the gene for cystic fibrosis, a common fatal genetic disorder in humans.
CFTR is a member of the ABC transporter family, and is composed of two domains each consisting of six transmembrane domains followed by a nucleotide-binding site. Loss of CFTR
function decreases transepithelial water secretion and, as a result, the layers of mucus that coat the respiratory tree, pancreatic ducts, and intestine are dehydrated and difficult to clear. The resulting blockage of these sites leads to pancreatic insufficiency, "meconium ileus", and devastating "chronic obstructive pulmonary disease" (Al-Awqati, Q. et al. (1992) J. Exp. Biol. 172:245-266).
The voltage-gated chloride channels (CLC) are characterized by 10-12 transmembrane domains, as well as two small globular domains known as CBS domains. The CLC
subunits probably function as homotetramers. CLC proteins are involved in regulation of cell volume, membrane potential stabilization, signal transduction, and transepithelial transport.
Mutations in CLC-1, expressed predominantly in skeletal muscle, are responsible for autosomal recessive generalized myotonia and autosomal dominant myotonia congenita, while mutations in the kidney channel CLC-5 lead to kidney stones (Jentsch, T.J. (1996) Curr. Opin. Neurobiol. 6:303-310).
Ligand-gated channels open their pores when an extracellular or intracellular mediator binds to the channel. Neurotransmitter-gated channels are channels that open when a neurotransmitter binds to their extracellular domain. These channels exist in the postsynaptic membrane of nerve or muscle cells.
There are two types of neurotransmitter-gated channels. Sodium channels open in response to excitatory neurotransmitters, such as acetylcholine, glutamate, and serotonin.
This opening causes an influx of Na+ and produces the initial localized depolarization that activates the voltage-gated channels and starts the action potential. Chloride channels open in response to inhibitory neurotransmitters, such as y-aminobutyric acid (GABA) and glycine, leading to hyperpolarization of the membrane and the subsequent generation of an action potential. Neurotransmitter-gated ion channels have four transmembrane domains and probably function as pentamers (Jentsch, supra).
Amino acids in the second transmembrane domain appear to be important in determining channel permeation and selectivity (Sather, W.A. et al. (1994) Curr. Opin. Neurobiol. 4:313-323).
Ligand-gated channels can be regulated by intracellular second messengers. For example, calcium-activated K+ channels are gated by internal calcium ions. In nerve cells, an influx of calcium during depolarization opens K+ channels to modulate the magnitude of the action potential (Ishi et al., supra). The large conductance (BK) channel has been purified from brain and its subunit composition determined. The a subunit of the BK channel has seven rather than six transmembrane domains in contrast to voltage-gated K+ channels. The extra transmembrane domain is located at the subunit N-terminus. A 28-amino-acid stretch in the C-terminal region of the subunit (the "calcium bowl" region) contains many negatively charged residues and is thought to be the region responsible for calcium binding. The (3 subunit consists of two transmembrane domains connected by a glycosylated extracellular loop, with intracellular N- and C-termini (Kaczorowski, supra;
Vergara, C. et al. (1998) Curr. Opin. Neurobiol. 8:321-329).
Cyclic nucleotide-gated (CNG) channels are gated by cytosolic cyclic nucleotides. The best examples of these are the cAMP-gated Na+ channels involved in olfaction and the cGMP-gated cation channels involved in vision. Both systems involve ligand-mediated activation of a G-protein coupled receptor which then alters the level of cyclic nucleotide within the cell. CNG
channels also represent a major pathway for Ca2+ entry into neurons, and play roles in neuronal development and plasticity.
CNG channels are tetramers containing at least two types of subunits, an a subunit which can form functional homomeric channels, and a [3 subunit, which modulates the channel properties. All CNG
subunits have six transmembrane domains and a pore forming region between the fifth and sixth transmembrane domains, similar to voltage-gated K+ channels. A large C-terminal domain contains a cyclic nucleotide binding domain, while the N-terminal domain confers variation among channel subtypes (Zufall, F. et al. (1997) Curr. Opin. Neurobiol. 7:404-412).
The activity of other types of ion channel proteins may also be modulated by a variety of intracellular signalling proteins. Many channels have sites for phosphorylation by one or more protein kinases including protein kinase A, protein kinase C, tyrosine kinase, and casein kinase II, all of which regulate ion channel activity in cells. Kir channels are activated by the binding of the G~i~y subunits of heterotrimeric G-proteins (Reimann, F. and F.M. Ashcroft (1999) Curr. Opin. Cell. Biol.
11:503-508). Other proteins are involved in the localization of ion channels to specific sites in the cell membrane. Such proteins include the PDZ domain proteins known as MAGUKs (membrane-associated guanylate kinases) which regulate the clustering of ion channels at neuronal synapses (Craven, S.E. and D.S. Bredt (1998) Cell 93:495-498).
Disease Correlation The etiology of numerous human diseases and disorders can be attributed to defects in the transport of molecules across membranes. Defects in the trafficking of membrane-bound transporters and ion channels are associated with several disorders, e.g., cystic fibrosis, glucose-galactose malabsorption syndrome, hypercholesterolemia, von Gierke disease, and certain forms of diabetes mellitus. Single-gene defect diseases resulting in an inability to transport small molecules across membranes include, e.g., cystinuria, iminoglycinuria, Hartup disease, and Fanconi disease (van't Hoff, W.G. (1996) Exp. Nephrol. 4:253-262; Talente, G.M. et al. (1994) Ann. Intern.
Med. 120:218-226;
and Chillon, M. et al. (1995) New Engl. J. Med. 332:1475-1480).
Human diseases caused by mutations in ion channel genes include disorders of skeletal muscle, cardiac muscle, and the central nervous system. Mutations in the pore-forming subunits of sodium and chloride channels cause myotonia, a muscle disorder in which relaxation after voluntary contraction is delayed. Sodium channel myotonias have been treated with channel blockers.
Mutations in muscle sodium and calcium channels cause forms of periodic paralysis, while mutations in the sarcoplasmic calcium release channel, T-tubule calcium channel, and muscle sodium channel cause malignant hyperthermia. Cardiac arrytlunia disorders such as the long QT syndromes and idiopathic ventricular fibrillation are caused by mutations in potassium and sodium channels (Cooper, E.C. and L.Y. Jan (1998) Proc. Natl. Acad. Sci. USA 96:4759-4766). All four known human idiopathic epilepsy genes code for ion channel proteins (Berkovic, S.F. and LE. Scheffer (1999) Curr.
Opin. Neurology 12:177-182). Other neurological disorders such as ataxias, hemiplegic migraine and hereditary deafness can also result from mutations in ion channel genes (Jen, J. (1999) Curr. Opin.
Neurobiol. 9:274-280;
Cooper, supra).
Ion channels have been the target for many drug therapies. Neurotransmitter-gated channels have been targeted in therapies for treatment of insomnia, anxiety, depression, and schizophrenia.
Voltage-gated channels have been targeted in therapies for arrhythmia, ischemic stroke, head trauma, and neurodegenerative disease (Taylor, C.P. and L.S. Narasimhan (1997) Adv.
Pharmacol. 39:47-98).
Various classes of ion channels also play an important role in the perception of pain, and thus are potential targets for new analgesics. These include the vanilloid-gated ion channels, which are activated by the vanilloid capsaicin, as well as by noxious heat. Local anesthetics such as lidocaine and mexiletine which blockade voltage-gated Na+ channels have been useful in the treatment of neuropathic pain (Eglen, supra).
Ion channels in the immune system have recently been suggested as targets for immunomodulation. T-cell activation depends upon calcium signaling, and a diverse set of T-cell specific ion channels has been characterized that affect this signaling process. Channel blocking agents can inhibit secretion of lymphokines, cell proliferation, and killing of target cells. A peptide antagonist of the T-cell potassium channel I~vl.3 was found to suppress delayed-type hypersensitivity and allogenic responses in pigs, validating the idea of channel blockers as safe and efficacious immunosuppressants (Calahan, M.D. and K.G. Chandy (1997) Curr. Opin.
Biotechnol. 8:749-756).
The discovery of new transporters and ion channels 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 transport, neurological, muscle, and immunological disorders, and in the assessment of the effects of exogenous compounds on the expression of nucleic acid and amino acid sequences of transporters and ion channels.
SUMMARY OF THE INVENTION
The invention features purified polypeptides, transporters and ion channels, referred to collectively as "TRICH" and individually as "TRICH-1," "TRICH-2," "TRICH-3,"
"TRICH-4,"

"TRICH-5," "TRICH-6," "TRICH-7," "TRICH-8," "TRICH-9," "TRICH-10," "TRICH-11,"
"TRICH-12," "TRICH-13," "TRICH-14," "TRICH-15," "TRICH-16," "TRICH-17," "TRICH-18,"
"TRICH-19," "TRICH-20," "TRICH-21," "TRICH-22," "TRICH-23," "TRICH-24," "TRICH-25,"
"TRICH-26," and "TRICH-27." In one aspect, the invention provides 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 N0:1-27, 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 N0:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27. In one alternative, the invention provides an isolated polypeptide comprising the amino acid sequence of SEQ ID NO:1-27.
The invention further provides an isolated polynucleotide encoding a 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 N0:1-27, 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 N0:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27. In one alternative, the polynucleotide encodes a polypeptide selected from the group consisting of SEQ ID NO:1-27. In another alternative, the polynucleotide is selected from the group consisting of SEQ ID N0:28-54.
Additionally, the invention provides a recombinant polynucleotide comprising a promoter sequence operably linked to a polynucleotide encoding a 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 N0:1-27, 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 N0:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID
N0:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27. 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 an amino acid sequence selected from the group consisting of a) an amino acid sequence selected from the group consisting of SEQ ID N0:1-27, 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 N0:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID
NO:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27. 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 linkedto 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 an amino acid sequence selected from the group consisting of a) an amino acid sequence selected from the group consisting of SEQ ID N0:1-27, 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 N0:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27.
The invention further provides 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 N0:28-54, b) a naturally occurring polynucleotide sequence having at least 90% sequence identity to a polynucleotide sequence selected from the group consisting of SEQ ID N0:28-54, c) a polynucleotide sequence complementary to a), d) a polynucleotide sequence complementary to b), and e) an RNA equivalent of a)-d). 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 polynucleotide sequence selected from the group consisting of a) a polynucleotide sequence selected from the group consisting of SEQ ID N0:28-54, b) a naturally occurring polynucleotide sequence having at least 90% sequence identity to a polynucleotide sequence selected from the group consisting of SEQ ID N0:28-54, c) a polynucleotide sequence complementary to a), d) a polynucleotide sequence complementary to b), and e) an RNA equivalent of a)-d). The method comprises a) hybridizing the sample with a probe comprising at least 20 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 or fragments thereof, 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 60 contiguous nucleotides.
The invention further provides a method for detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide comprising a polynucleotide sequence selected from the group consisting of a) a polynucleotide sequence selected from the group consisting of SEQ ID N0:28-54, b) a naturally occurring polynucleotide sequence having at least 90% sequence identity to a polynucleotide sequence selected from the group consisting of SEQ ID N0:28-54, c) a polynucleotide sequence complementary to a), d) a polynucleotide sequence complementary to b), and e) an RNA equivalent of a)-d). The method comprises a) amplifying said target polynucleotide or fragment thereof using polymerase chain reaction amplification, and b) detecting the presence or absence of said amplified target polynucleotide or fragment thereof, and, optionally, if present, the amount thereof.
The invention further provides a composition comprising an effective amount of a 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:1-27, 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 N0:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27, and a pharmaceutically acceptable excipient In one embodiment, the composition comprises an amino acid sequence selected from the group consisting of SEQ ID NO:1-27. The invention additionally provides a method of treating a disease or condition associated with decreased expression of functional TRICH, comprising administering to a patient in need of such treatment the composition.
The invention also provides a method for screening a compound for effectiveness as an agonist of a 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 N0:1-27, 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 N0:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-27. 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 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 TRICH, comprising administering to a patient in need of such treatment the composition.
Additionally, the invention provides a method for screening a compound for effectiveness as an antagonist of a 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 N0:1-27, 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 N0:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27. 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 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 TRICH, comprising administering to a patient in need of such treatment the composition.
The invention further provides a method of screening for a compound that specifically binds to a 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:I-27, 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 N0:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-27. The method comprises a) combining the polypeptide with at least one test compound under suitable conditions, and b) detecting binding of the polypeptide to the test compound, thereby identifying a compound that specifically binds to the polypeptide.
The invention further provides a method of screening for a compound that modulates the activity of a 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 N0:1-27, 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:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID N0:1-27. The method comprises a) combining the polypeptide with at least one test compound under conditions permissive for the activity of the polypeptide, b) assessing the activity of the polypeptide in the presence of the test compound, and c) comparing the activity of the polypeptide in the presence of the test compound with the activity of the polypeptide in the absence of the test compound, wherein a change in the activity of the polypeptide in the presence of the test compound is indicative of a compound that modulates the activity of the polypeptide.

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 N0:28-54, the method comprising a) exposing a sample comprising the target polynucleotide to a compound, and b) detecting altered expression of the target polynucleotide.
The invention further provides a method for assessing toxicity of a test compound, said method comprising a) treating a biological sample containing nucleic acids with the test compound;
b) hybridizing the nucleic acids of the treated biological sample with a probe comprising at least 20 contiguous nucleotides of a polynucleotide comprising a polynucleotide sequence selected from the group consisting of i) a polynucleotide sequence selected from the group consisting of SEQ ID
N0:28-54, ii) a naturally occurring polynucleotide sequence having at least 90% sequence identity to a polynucleotide sequence selected from the group consisting of SEQ ID N0:28-54, iii) a polynucleotide sequence complementary to i), iv) a polynucleotide sequence complementary to ii), and v) an RNA equivalent of i)-iv). Hybridization occurs under conditions whereby a specific hybridization complex is formed between said probe and a target polynucleotide in the biological sample, said target polynucleotide comprising a polynucleotide sequence selected from the group consisting of i) a polynucleotide sequence selected from the group consisting of SEQ ID N0:28-54, ii) a naturally occurring polynucleotide sequence having at least 90% sequence identity to a polynucleotide sequence selected from the group consisting of SEQ ID N0:28-54, iii) a polynucleotide sequence complementary to i), iv) a polynucleotide sequence complementary to ii), and v) an RNA equivalent of i)-iv). Alternatively, the target polynucleotide comprises a fragment of a polynucleotide sequence selected from the group consisting of i)-v) above;
c) quantifying the amount of hybridization complex; and d) comparing the amount of hybridization complex in the treated biological sample with the amount of hybridization complex in an untreated biological sample, wherein a difference in the amount of hybridization complex in the treated biological sample is indicative of toxicity of the test compound.
BRIEF DESCRIPTION OF THE TABLES
Table 1 summarizes the nomenclature for the full length polynucleotide and polypeptide sequences of the present invention.
Table 2 shows the GenBank identification number and annotation of the nearest GenBank homolog for each polypeptide of the invention. The probability score for the match between each polypeptide and its GenBank homolog is also shown.
Table 3 shows structural features of each polypeptide sequence, including predicted motifs and domains, along with the methods, algorittuns, and searchable databases used for analysis of each polypeptide.
Table 4 lists the cDNA and genomic DNA fragments which were used to assemble each polynucleotide sequence, along with selected fragments of the polynucleotide sequences.
Table 5 shows the representative cDNA library for each polynucleotide of the invention.
Table 6 provides an appendix which describes the tissues and vectors used for construction of the cDNA libraries shown in Table 5.
Table 7 shows the tools, programs, and algorithms used to analyze the polynucleotides and polypeptides of the invention, 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
"TRICH" refers to the amino acid sequences of substantially purified TRICH
obtained from any species, particularly a mammalian species, including bovine, ovine, porcine, marine, 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 TRICH. Agonists may include proteins, nucleic acids, carbohydrates, small molecules, or any other compound or composition which modulates the activity of TRICH either by directly interacting with TRICH or by acting on components of the biological pathway in which TRICH
participates.
An "allelic variant" is an alternative form of the gene encoding TRICH.
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 TRICH include those sequences with deletions, insertions, or substitutions of different nucleotides, resulting in a polypeptide the same as TRICH or a polypeptide with at least one functional characteristic of TRICH. Included within this definition are polymorphisms which may or may not be readily detectable using a particular oligonucleotide probe of the polynucleotide encoding TRICH, and improper or unexpected hybridization to allelic variants, with a locus other than the normal chromosomal locus for the polynucleotide sequence encoding TRICH.
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 TRICH.
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 TRICH 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 a 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 TRICH. Antagonists may include proteins such as antibodies, nucleic acids, carbohydrates, small molecules, or any other compound or composition which modulates the activity of TRICH either by directly interacting with TRICH or by acting on components of the biological pathway in which TRICH
participates.
The term "antibody" refers to intact immunoglobulin molecules as well as to fragments thereof, such as Fab, F(ab')2, and Fv fragments, which are capable of binding an epitopic determinant.
Antibodies that bind TRICH 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"
(coding) 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 sugaxs 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" or "immunogenic"
refers to the capability of the natural, recombinant, or synthetic TRICH, or of any oligopeptide thereof, to induce a specific immune response in appropriate animals or cells and to bind with specific antibodies.

"Complementary" describes the relationship between two single-stranded nucleic acid sequences that anneal by base-pairing. For example, 5'-AGT-3' pairs with its complement, 3'-TCA-5'.
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 TRICH or fragments of TRICH 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., NaCl), 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 subjected tb repeated DNA sequence analysis to resolve uncalled bases, extended using the XL-PCR kit (PE Biosystems, Foster City CA) in the 5' and/or the 3' direction, and resequenced, or which has been assembled from one or more overlapping cDNA, EST, or genomic DNA fragments using a computer program for fragment assembly, such as the GELVIEW fragment assembly system (GCG, Madison WI) or Phrap (University of Washington, Seattle WA). Some sequences have been both extended and assembled to produce the consensus sequence.
"Conservative amino acid substitutions" are those substitutions that are predicted to 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 ~n Asp, Gln, His psp 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 T~' 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 a chemically modified polynucleotide or polypeptide. Chemical modifications of a polynucleotide can include, for example, replacement of hydrogen by an alkyl, aryl, 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 "detectable label" refers to a reporter molecule or enzyme that is capable of generating a measurable signal and is covalently or noncovalently joined to a polynucleotide or polypeptide.
A "fragment" is a unique portion of TRICH or the polynucleotide encoding TRICH
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 N0:28-54 comprises a region of unique polynucleotide sequence that specifically identifies SEQ ID N0:28-54, for example, as distinct from any other sequence in the genome from which the fragment was obtained. A fragment of SEQ ID N0:28-54 is useful, for example, in hybridization and amplification technologies and in analogous methods that distinguish SEQ ID N0:28-54 from related polynucleotide sequences. The precise length of a fragment of SEQ

ID N0:28-54 and the region of SEQ ID N0:28-54 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 N0:1-27 is encoded by a fragment of SEQ ID N0:28-54. A
fragment of SEQ ID N0:1-27 comprises a region of unique amino acid sequence that specifically identifies SEQ ID NO:1-27. For example, a fragment of SEQ ID N0:1-27 is useful as an immunogenic peptide for the development of antibodies that specifically recognize SEQ ID N0:1-27.
The precise length of a fragment of SEQ ID N0:1-27 and the region of SEQ ID NO:l-27 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 "full length" polynucleotide sequence is one containing at least a translation initiation codon (e.g., methionine) followed by an open reading frame and a translation termination codon. A "full length" polynucleotide sequence encodes a "full length" polypeptide sequence.
"Homology" refers to sequence similarity or, interchangeably, sequence identity, between two or more polynucleotide sequences or two or more polypeptide sequences.
The terms "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 WI). 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:
Ktupl~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 sequences.
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 2Ø12 (April-21-2000) set at default parameters. Such default parameters may be, for example:
Matrix: BLOSUM62 Reward for match: 1 Penalty for mismatch: -2 Opera Gap: 5 and Extension Gap: 2 penalties Gap x drop-off.- SO -Expect: 10 Word Size: Il Filter: ort 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 charge and hydrophobicity 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 2Ø12 (April-21-2000) with blastp set at default parameters. Such default parameters may be, for example:
Matrix: BLOSUM62 Opera Gap: 11 arad Extension Gap: 1 penalties Gap x drop-off. 50 Expect: 1 D
Word Size: 3 Filter: ora 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 chromosome replication, segregation and maintenance.
The term "humanized antibody" refers to an antibody molecule 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 complementarity.
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 ~ g/ml sheared, 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. Such wash temperatures are typically selected to be about 5°C to 20°C lower than the thermal melting point (T"~ for the specific sequence at a de~'med 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, J.
et al. (1989) Molecular Cloning: A Laboratory Manual, 2nd 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, sheared and 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 TRICH
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 TRICH 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 a plurality of polynucleotides, polypeptides, or other chemical compounds on a substrate.
The terms "element" and "array element" refer to a polynucleotide, polypeptide, or other chemical compound having a unique and defined position on a microarray.
The term "modulate" refers to a change in the activity of TRICH. For example, modulation may cause an increase or a decrease in protein activity, binding characteristics, or any other biological, functional, or immunological properties of TRICH.
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 a 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. 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.
"Post-translational modification" of an TRICH may involve lipidation, glycosylation, phosphorylation, acetylation, racemization, proteolytic cleavage, and other modifications known in the art. These processes may occur synthetically or biochemically. Biochemical modifications will vary by cell type depending on the enzymatic milieu of TRICH.
"Probe" refers to nucleic acid sequences encoding TRICH, 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
polymerase enzyme. Primer pairs can be used for amplification (and identification) of a nucleic acid sequence, e.g., by the polymerase 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, J. et al. (1989) Molecular Cloning: A Laboratory Manual, 2"d ed., vol. 1-3, Cold Spring Harbor Press, Plainview NY; Ausubel, F.M. et al. (1987) Current Protocols in Molecular Biolo~v, Greene Publ. Assoc. & Wiley-Intersciences, New York NY; Innis, M. et al. (f990) 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/MIT 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.
A "regulatory element" refers to a nucleic acid sequence usually derived from untranslated regions of a gene and includes enhancers, promoters, introns, and 5' and 3' untranslated regions (UTRs).
Regulatory elements interact with host or viral proteins which control transcription, translation, or RNA
stability.
"Reporter molecules" are chemical or biochemical moieties used for labeling a nucleic acid, amino acid, or antibody. Reporter molecules include radionuclides; enzymes;
fluorescent, chemiluminescent, or chromogenic agents; substrates; cofactors; inhibitors;
magnetic particles; and other moieties known in the art.
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 TRICH, nucleic acids encoding TRICH, or fragments thereof 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 comprising 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 acid residues or nucleotides by different amino acid residues 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.
A "transcript image" refers to the collective pattern of gene expression by a particular cell type or tissue under given conditions at a given time.
"Transformation" describes a process by which exogenous DNA is introduced into 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, bacteriophage or 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, 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), supra.
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 alternative 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 will generally 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 transporters and ion channels (TRICH), the polynucleotides encoding TRICH, and the use of these compositions for the diagnosis, treatment, or prevention of transport, neurological, muscle, and immunological disorders.

Table 1 sununarizes the nomenclature for the full length polynucleotide and polypeptide sequences of the invention. Each polynucleotide and its corresponding polypeptide are correlated to a single Incyte project identification number (Incyte Project ID). Each polypeptide sequence is denoted by both a polypeptide sequence identification number (Polypeptide SEQ ID NO:) and an Incyte polypeptide sequence number (Incyte Polypeptide ID) as shown. Each polynucleotide sequence is denoted by both a polynucleotide sequence identification number (Polynucleotide SEQ ID NO:) and an Incyte polynucleotide consensus sequence number (Incyte Polynucleotide ID) as shown.
Table 2 shows sequences with homology to the polypeptides of the invention as identified by BLAST analysis against the GenBank protein (genpept) database. Columns 1 and 2 show the polypeptide sequence identification number (Polypeptide SEQ ID NO:) and the corresponding Incyte polypeptide sequence number (Incyte Polypeptide ID) for each polypeptide of the invention. Column 3 shows the GenBank identification number (Genbank ID NO:) of the nearest GenBank homolog.
Column 4 shows the probability score for the match between each polypeptide and its GenBank homolog. Column 5 shows the annotation of the GenBank homolog along with relevant citations where applicable, all of which are expressly incorporated by reference herein.
Table 3 shows various structural features of each of the polypeptides of the invention. Columns 1 and 2 show the polypeptide sequence identification number (SEQ ID NO:) and the corresponding Incyte polypeptide sequence number (Incyte Polypeptide ID) for each polypeptide of the invention.
Column 3 shows the number of amino acid residues in each polypeptide. Column 4 shows potential phosphorylation sites, and column 5 shows potential glycosylation sites, as determined by the MOTIFS
program of the GCG sequence analysis software package (Genetics Computer Group, Madison WI).
Column 6 shows amino acid residues comprising signature sequences, domains, and motifs. Column 7 shows analytical methods for protein structurelfunction analysis and in some cases, searchable databases to which the analytical methods were applied.
As shown in Table 4, the full length polynucleotide sequences of the present invention were assembled using cDNA sequences or coding (exon) sequences derived from genomic DNA, or any combination of these two types of sequences. Columns 1 and 2 list the polynucleotide sequence identification number (Polynucleotide SEQ ID NO:) and the corresponding Incyte polynucleotide consensus sequence number (Incyte Polynucleotide ID) for each polynucleotide of the invention.
Column 3 shows the length of each polynucleotide sequence in basepairs. Column 4 lists fragments of the polynucleotide sequences which are useful, for example, in hybridization or amplification technologies that identify SEQ ID N0:28-54 or that distinguish between SEQ ID
N0:28-54 and related polynucleotide sequences. Column 5 shows identification numbers corresponding to cDNA
sequences, coding sequences (exons) predicted from genomic DNA, and/or sequence assemblages comprised of both cDNA and genomic DNA. These sequences were used to assemble the full length polynucleotide sequences of the invention. Columns 6 and 7 of Table 4 show the nucleotide start (5') and stop (3') positions of the cDNA and genomic sequences in column 5 relative to their respective full length sequences.
The identification numbers in Column 5 of Table 4 may refer specifically, for example, to Incyte cDNAs along with their corresponding cDNA libraries. For example, 6813453H1 is the identification number of an Incyte cDNA sequence, and ADRETUR01 is the cDNA
library from which it is derived. Incyte cDNAs for which cDNA libraries are not indicated were derived from pooled cDNA libraries (e.g., 70207988V1). Alternatively, the identification numbers in column 5 may refer to GenBank cDNAs or ESTs (e.g., g1947104) which contributed to the assembly of'the full length polynucleotide sequences. Alternatively, the identification numbers in column 5 may refer to coding regions predicted by Genscan analysis of genomic DNA. For example, GNN.g6554406_006 is the identification number of a Genscan-predicted coding sequence, with g6554406 being the GeriBank identification number of the sequence to which Genscan was applied. The Genscan-predicted coding sequences may have been edited prior to assembly. (See Example IV.) Alternatively, the identification numbers in column 5 may refer to assemblages of both cDNA and Genscan-predicted exons brought together by an "exon stitching" algorithm. (See Example V.) Alternatively, the identification numbers in column 5 may refer to assemblages of both cDNA and Genscan-predicted exons brought together by an "exon-stretching" algorithm. (See Example V.) In some cases, Incyte cDNA
coverage redundant with the sequence coverage shown in column 5 was obtained to confirm the final consensus polynucleotide sequence, but the relevant Incyte cDNA identification numbers are not shown.
Table 5 shows the representative cDNA libraries for those full length polynucleotide sequences which were assembled using Incyte cDNA sequences. The representative cDNA
library is the Incyte cDNA library which is most frequently represented by the Incyte cDNA sequences which were used to assemble and conf'~rm the above polynucleotide sequences. The tissues and vectors which were used to construct the cDNA libraries shown in Table 5 are described in Table 6.
The invention also encompasses TRICH variants. A preferred TRICH 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 TRICH amino acid sequence, and which contains at least one functional or structural characteristic of TRICH.
The invention also encompasses polynucleotides which encode TRICH. In a particular embodiment, the invention encompasses a polynucleotide sequence comprising a sequence selected from the group consisting of SEQ ID N0:28-54, which encodes TRICH. The polynucleotide sequences of SEQ ID N0:28-54, 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 TRICH. In particular, such a variant polynucleotide sequence will have at least about 70%, or alternatively at least about 85 %, or even at least about 95 % polynucleotide sequence identity to the polynucleotide sequence encoding TRICH. A particular aspect of the invention encompasses a variant of a polynucleotide sequence comprising a sequence selected from the group consisting of SEQ ID
N0:28-54 which has at least about 70%, or alternatively at least about 85%, or even at least about 95% polynucleotide sequence identity to a nucleic acid sequence selected from the group consisting of SEQ ID N0:28-54.
Any one of the polynucleotide variants described above can encode an amino acid sequence which contains at least one functional or structural characteristic of TRICH.
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 TRICH, some bearing minimal similarity to the polynucleotide sequences of any known and naturally occurring 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 TRICH, and all such variations are to be considered as being specifically disclosed.
Although nucleotide sequences which encode TRICH and its variants are generally capable of hybridizing to the nucleotide sequence of the naturally occurring TRICH under appropriately selected conditions of stringency, it may be advantageous to produce nucleotide sequences encoding TRICH 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 TRICH 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 TRICH
and TRICH 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 TRICH 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 ID
NO:28-54 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.
152:507-511.) 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 (PE
Biosystems), 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 (PE
Biosystems). Sequencing is then carried out using either the ABI 373 or 377 DNA sequencing system (PE Biosystems), 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 Biology and Biotechnolo~y> Wiley VCH, New York NY, pp. 856-853.) The nucleic acid sequences encoding TRICH 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 a1. (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, PE Biosystems), 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 TRICH may be cloned in recombinant DNA molecules that direct expression of TRICH, 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 TRICH.
The nucleotide sequences of the present invention can be engineered using methods generally known in the art in order to alter TRICH-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 Clara 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 TRICH, 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 TRICH 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 a1. (1980) Nucleic Acids Symp. Ser.
7:225-232.) Alternatively, TRICH itself or a fragment thereof may be synthesized using chemical methods.
For example, peptide synthesis can be performed using various solution-phase or solid-phase techniques. (See, e.g., Creighton, T. (1984) Proteins, Structures and Molecular Properties, WH
Freeman, New York NY, pp.55-60; and Roberge, J.Y. et al. (1995) Science 269:202-204.) Automated synthesis may be achieved using the ABI 431A peptide synthesizer (PE Biosystems). Additionally, the amino acid sequence of TRICH, 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 or a polypeptide having a sequence of a naturally occurring polypeptide.
The peptide may be substantially purified by preparative high performance liquid chromatography. (See, e.g., Chiez, R.M. and F.2. Regnier (1990) Methods Enzymol. 182:392-421.) The composition of the synthetic peptides may be conf'~rmed by amino acid analysis or by sequencing.
(See, e.g., Creighton, su ra, pp. 28-53.) In order to express a biologically active TRICH, the nucleotide sequences encoding TRICH 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 TRICH. Such elements may vary in their strength and specificity.
Specific initiation signals may also be used to achieve more efficient translation of sequences encoding TRICH. Such signals include the ATG initiation codon and adjacent sequences, e.g. the Kozak sequence. In cases where sequences encoding TRICH 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 TRICH 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 TRICH. 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. (See, e.g., Sambrook, su ra; Ausubel, supra; Van Heeke, G. and S.M. Schuster (1989) J. Biol. Chem. 264:5503-5509; 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; Takamatsu, N. (1987) EMBO
J. 6:307-311; The McGraw Hill Yearbook of Science and Technolo~y (1992) McGraw Hill, New York NY, pp. 191-196; Logan, J. and T. Shenk (1984) Proc. Natl. Acad. Sci. USA
81:3655-3659; and Harrington, J.J. et al. (1997) Nat. Genet. 15:345-355.) 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. (See, e.g., Di Nicola, M. et a1. (1998) Cancer Gen. Ther. 5(6):350-356; Yu, M. et al. (1993) Proc. Natl. Acad. Sci.
USA 90(13):6340-6344; Buller, R.M. et al. (1985) Nature 317(6040):813-815;
McGregor, D.P. et al.
(1994) Mol. Immunol. 31(3):219-226; and Verma, LM. and N. Somia (1997) Nature 389:239-242.) 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 TRICH. For example, routine cloning, subcloning, and propagation of polynucleotide sequences encoding TRICH can be achieved using a multifunctional E. coli vector such as PBLLTESCRIPT (Stratagene, La Jolla CA) or PSPORT1 plasmid (Life Technologies). Ligation of sequences encoding TRICH 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 TRICH are needed, e.g. for the production of antibodies, vectors which direct high level expression of TRICH may be used. For example, vectors containing the strong, inducible SP6 or T7 bacteriophage promoter may be used.
Yeast expression systems may be used for production of TRICH. 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 uastoris. 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 TRICH. Transcription of sequences encoding TRICH may be driven by 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 Technolo~y (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 TRICH
may be ligated into an adenovirus transcription/translation complex consisting of the late promoter and tripartite leader sequence. Insertion in a non-essential E1 or E3 region of the viral genome may be used to obtain infective virus which expresses TRICH 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 TRICH in cell lines is preferred. For example, sequences encoding TRICH 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 apY 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),13 glucuronidase and its substrate 13-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 presencelabsence 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 TRICH is inserted within a marker gene sequence, transformed cells containing sequences encoding TRICH can be identified by the absence of marker gene function. Alternatively, a marker gene can be placed in tandem with a sequence encoding TRICH 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 TRICH
and that express TRICH 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 TRICH
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 TRICH 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 Immunolo~y, 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 TRICH include oligolabeling, nick translation, end-labeling, or PCR amplification using a labeled nucleotide.
Alternatively, the sequences encoding TRICH, 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 WI), 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 TRICH 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 TRICH may be designed to contain signal sequences which direct secretion of TRICH 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 TRICH 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 TRICH protein containing a heterologous moiety that can be recognized by a commercially available antibody may facilitate the screening of peptide libraries for inhibitors of TRICH
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-rrayc, 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 TRICH encoding sequence and the heterologous protein sequence, so that TRICH
may be cleaved away from the heterologous moiety following purification.
Methods for fusion protein expression and purification are discussed in Ausubel (1995, sera, 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 TRICH 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.
TRICH of the present invention or fragments thereof may be used to screen for compounds that specifically bind to TRICH. At least one and up to a plurality of test compounds may be screened for specific binding to TRICH. Examples of test compounds include antibodies, oligonucleotides, proteins (e.g., receptors), or small molecules.
In one embodiment, the compound thus identified is closely related to the natural ligand of TRICH, e.g., a ligand or fragment thereof, a natural substrate, a structural or functional mimetic, or a natural binding partner. (See, e.g., Coligan, J.E, et al. (1991) Current Protocols in Immunolo~y 1(2):
Chapter 5.) Similarly, the compound can be closely related to the natural receptor to which TRICH
binds, or to at least a fragment of the receptor, e.g., the ligand binding site. In either case, the compound can be rationally designed using known techniques. In one embodiment, screening for these compounds involves producing appropriate cells which express TRICH, either as a secreted protein or on the cell membrane. Preferred cells include cells from mammals, yeast, Drosophila, or E.
coli. Cells expressing TRICH or cell membrane fractions which contain TRICH
are then contacted with a test compound and binding, stimulation, or inhibition of activity of either TRICH or the compound is analyzed.
An assay may simply test binding of a test compound to the polypeptide, wherein binding is detected by a fluorophore, radioisotope, enzyme conjugate, or other detectable label. For example, the assay may comprise the steps of combining at least one test compound with TRICH, either in solution or affixed to a solid support, and detecting the binding of TRICH to the compound.
Alternatively, the assay may detect or measure binding of a test compound in the presence of a labeled competitor. Additionally, the assay may be carried out using cell-free preparations, chemical libraries, or natural product mixtures, and the test compounds) may be free in solution or affixed to a solid support.
TRICH of the present invention or fragments thereof may be used to screen for compounds that modulate the activity of TRICH. Such compounds may include agonists, antagonists, or partial or inverse agonists. In one embodiment, an assay is performed under conditions permissive for TRICH
activity, wherein TRICH is combined with at least one test compound, and the activity of TRICH in the presence of a test compound is compared with the activity of TRICH in the absence of the test compound. A change in the activity of TRICH in the presence of the test compound is indicative of a compound that modulates the activity of TRICH. Alternatively, a test compound is combined with an in vitro or cell-free system comprising TRICH under conditions suitable for TRICH activity, and the assay is performed. In either of these assays, a test compound which modulates the activity of TRICH
may do so indirectly and need not come in direct contact with the test compound. At least one and up to a plurality of test compounds may be screened.
In another embodiment, polynucleotides encoding TRICH or their mammalian homologs may be "knocked out" in an animal model system using homologous recombination in embryonic stem (ES) cells. Such techniques are well known in the art and are useful for the generation of animal models of human disease. (See, e.g., U.S. Patent Number 5,175,383 and U.S. Patent Number 5,767,337.) For example, mouse ES cells, such as the mouse 129/SvJ cell line, are derived from the early mouse embryo and grown in culture. The ES cells are transformed with a vector containing the gene of interest disrupted by a marker gene, e.g., the neomycin phosphotransferase gene (neo;
Capecchi, M.R. (1989) Science 244:1288-1292). The vector integrates into the corresponding region of the host genome by homologous recombination. Alternatively, homologous recombination takes place using the Cre-loxP
system to knockout a gene of interest in a tissue- or developmental stage-specific manner (Marth, J.D.
(1996) Clin. Invest. 97:1999-2002; Wagner, K.U. et al. (1997) Nucleic Acids Res. 25:4323-4330).
Transformed ES cells are identified and microinjected into mouse cell blastocysts such as those from the C57BL/6 mouse strain. The blastocysts are surgically transferred to pseudopregnant dams, and the resulting chimeric progeny are genotyped and bred to produce heterozygous or homozygous strains.
Transgenic animals thus generated may be tested with potential therapeutic or toxic agents.
Polynucleotides encoding TRICH may also be manipulated in vitro in ES cells derived from human blastocysts. Human ES cells have the potential to differentiate into at least eight separate cell lineages including endoderm, mesoderm, and ectodermal cell types. These cell lineages differentiate into, for example, neural cells, hematopoietic lineages, and cardiomyocytes (Thomson, J.A. et al. (1998) Science 282:1145-1147).
Polynucleotides encoding TRICH can also be used to create ''knockin" humanized animals (pigs) or transgenic animals (mice or rats) to model human disease. With knockin technology, a region of a polynucleotide encoding TRICH is injected into animal ES cells, and the injected sequence integrates into the animal cell genome. Transformed cells are injected into blastulae, and the blastulae are implanted as described above. Transgenic progeny or inbred lines are studied and treated with potential pharmaceutical agents to obtain information on treatment of a human disease. Alternatively, a mammal inbred to overexpress TRICH, e.g., by secreting TRICH in its milk, may also serve as a convenient source of that protein (Janne, J. et al. (1998) Biotechnol. Annu.
Rev. 4:55-74).
THERAPEUTICS
Chemical and structural similarity, e.g., in the context of sequences and motifs, exists between regions of TRICH and transporters and ion channels. Therefore, TRICH
appears to play a role in transport, neurological, muscle, and immunological disorders. In the treatment of disorders associated with increased TRICH expression or activity, it is desirable to decrease the expression or activity of TRICH. In the treatment of disorders associated with decreased TRICH expression or activity, it is desirable to increase the expression or activity of TRICH.
Therefore, in one embodiment, TRICH 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 TRICH. Examples of such disorders include, but are not limited to, a transport disorder such as akinesia, amyotrophic lateral sclerosis, ataxiatelangiectasia, cystic fibrosis, Becker's muscular dystrophy, Bell's palsy, Charcot-Marie Tooth disease, diabetes mellitus, diabetes insipidus, diabetic neuropathy, Duchenne muscular dystrophy, hyperkalemic periodic paralysis, normokalemic periodic paralysis, Parkinson's disease, malignant hyperthermia, multidrug resistance, myasthenia gravis, myotonic dystrophy, catatonia, tardive dyskinesia, dystonias, peripheral neuropathy, cerebral neoplasms, prostate cancer, cardiac disorders associated with transport, e.g., angina, bradyarrythmia, tachyartythrnia, hypertension, Long QT syndrome, myocarditis, cardiomyopathy, nemaline myopathy, centronuclear myopathy, lipid myopathy, nutochondrial myopathy, thyrotoxic myopathy, ,ethanol myopathy, dermatomyositis, inclusion body myositis, infectious myositis, polymyositis, neurological disorders associated with transport, e.g., Alzheimer's disease, amnesia, bipolar disorder, dementia, depression, epilepsy, Tourette's disorder, paranoid psychoses, and schizophrenia, and other disorders associated with transport, e.g., neurofibromatosis, postherpetic neuralgia, trigeminal neuropathy, sarcoidosis, sickle cell anemia, Wilson's disease, cataracts, infertility, pulmonary artery stenosis, sensorineural autosomal deafness, hyperglycemia, hypoglycemia, Grave's disease, goiter, Cushing's disease, Addison's disease, glucose-galactose malabsorption syndrome, hypercholesterolemia, adrenoleukodystrophy, Zellweger syndrome, Menkes disease, occipital horn syndrome, von Gierke disease, cystinuria, iminoglycinuria, Hartup disease, and Fanconi disease; a neurological disorder such as epilepsy, ischemic cerebrovascular disease, stroke, cerebral neoplasms, Alzheimer's disease, Pick's disease, Huntington's disease, dementia, Parkinson's disease and other extrapyramidal disorders, amyotrophic lateral sclerosis and other motor neuron disorders, progressive neural muscular atrophy, retinitis pigmentosa, hereditary ataxias, multiple sclerosis and other demyelinating diseases, bacterial and viral meningitis, brain abscess, subdural empyema, epidural abscess, suppurative intracranial thrombophlebitis, myelitis and radiculitis, viral central nervous system disease, prion diseases including kuru, Creutzfeldt-Jakob disease, and Gerstmann-Straussler-Scheinker syndrome, fatal familial insomnia, nutritional and metabolic diseases of the nervous system, neurofibromatosis, tuberous sclerosis, cerebelloretinal hemangioblastomatosis, encephalotrigeminal syndrome, mental retardation and other developmental disorders of the central nervous system including Down syndrome, cerebral palsy, neuroskeletal disorders, autonomic nervous system disorders, cranial nerve disorders, spinal cord diseases, muscular dystrophy and other neuromuscular disorders, peripheral nervous system disorders, dermatomyositis and polymyositis, inherited, metabolic, endocrine, and toxic myopathies, myasthenia gravis, periodic paralysis, mental disorders including mood, anxiety, and schizophrenic disorders, seasonal affective disorder (SAD), akathesia, amnesia, catatonia, diabetic neuropathy, tardive dyskinesia, dystonias, paranoid psychoses, postherpetic neuralgia, Tourette's disorder, progressive supranuclear palsy, corticobasal degeneration, and familial frontotemporal dementia; a muscle disorder such as cardiomyopathy, myocarditis, Duchenne's muscular dystrophy, Becker's muscular dystrophy, myotonic dystrophy, central core disease, nemaline myopathy, centronuclear myopathy, lipid myopathy, mitochondria) myopathy, infectious myositis, polymyositis, dermatomyositis, inclusion body myositis, thyrotoxic myopathy, ethanol myopathy, angina, anaphylactic shock, arrhythmias, asthma, cardiovascular shock, Cushing's syndrome, hypertension, hypoglycemia, myocardial infarction, migraine, pheochromocytoma, and myopathies including encephalopathy, epilepsy, Kearns-Sayre syndrome, lactic acidosis, myoclonic disorder, ophthalmoplegia, and acid maltase deficiency (AMD, also known as Pompe's disease); and an irnmunological disorder such as acquired immunodeficiency syndrome (AIDS), Addison's disease, adult respiratory distress syndrome, allergies, ankylosing spondylitis, amyloidosis, anemia, asthma, atherosclerosis, autoimmune hemolytic anemia, autoimmune thyroiditis, autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED), bronchitis, cholecystitis, contact dermatitis, Crohn's disease, atopic dermatitis, dermatomyositis, diabetes mellitus, emphysema, episodic lymphopenia with lymphocytotoxins, erythroblastosis fetalis, erythema nodosum, atrophic gastritis, glomerulonephritis, Goodpasture's syndrome, gout, Graves' disease, Hashimoto's thyroiditis, hypereosinophilia, irritable bowel syndrome, multiple sclerosis, myasthenia gravis, myocardial or pericardial inflammation, osteoarthritis, osteoporosis, pancreatitis, polymyositis, psoriasis, Reiter's syndrome, rheumatoid arthritis, scleroderma, Sjogren's syndrome, systemic anaphylaxis, systemic lupus erythematosus, systemic sclerosis, thrombocytopenic purpura, ulcerative colitis, uveitis, Werner syndrome, complications of cancer, hemodialysis, and extracorporeal circulation, viral, bacterial, fungal, parasitic, protozoa), and helminthic infections, and trauma.
In another embodiment, a vector capable of expressing TRICH 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 TRICH including, but not limited to, those described above.
In a further embodiment, a composition comprising a substantially purified TRICH in conjunction with a suitable pharmaceutical carrier may be administered to a subject to treat or prevent a disoxder associated with decreased expression or activity of TRICH including, but not limited to, those provided above.
In still another embodiment, an agonist which modulates the activity of TRICH
may be administered to a subject to treat or prevent a disorder associated with decreased expression or activity of TRICH including, but not limited to, those listed above.
In a further embodiment, an antagonist of TRICH may be administered to a subject to treat or prevent a disorder associated with increased expression or activity of TRICH.
Examples of such disorders include, but are not limited to, those transport, neurological, muscle, and immunological disorders described above. In one aspect, an antibody which specifically binds TRICH 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 TRICH.
In an additional embodiment, a vector expressing the complement of the polynucleotide encoding TRICH may be administered to a subject to treat or prevent a disorder associated with increased expression or activity of TRICH 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 TRICH may be produced using methods which are generally known in the art.
In particular, purified TRICH may be used to produce antibodies or to screen libraries of pharmaceutical agents to identify those which specifically bind TRICH.
Antibodies to TRICH 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 TRICH 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, KLH, and dinitrophenol. Among adjuvants used in humans, BCG
(bacilli Calmette-Guerin) and Corynebacterium parvum are especially preferable.
It is preferred that the oligopeptides, peptides, or fragments used to induce antibodies to TRICH 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. Short stretches of TRICH 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 TRICH 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 EB
V-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 a1. (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 TRICH-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 TRICH 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 TRICH and its specific antibody. A two-site, monoclonal-based immunoassay utilizing monoclonal antibodies reactive to two non-interfering TRICH epitopes is generally used, but a competitive binding assay may also be employed (Pound, supra).
Various methods such as Scatchard analysis in conjunction with radioimmunoassay techniques may be used to assess the affinity of antibodies for TRICH. Affinity is expressed as an association constant, Ka, which is defined as the molar concentration of TRICH-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 TRICH epitopes, represents the average affinity, or avidity, of the antibodies for TRICH. The Ka determined for a preparation of monoclonal antibodies, which are monospecific for a particular TRICH
epitope, represents a true measure of affinity. High-affinity antibody preparations with Ka ranging from about 109 to 1012 L/mole are preferred for use in immunoassays in which the TRICH-antibody complex must withstand rigorous manipulations. Low-affinity antibody preparations with Ka ranging from about 106 to 10' L/mole are preferred for use in immunopurification and similar procedures which ultimately require dissociation of TRICH, preferably in active form, from the antibody (Catty, D.
(1988) Antibodies, Volume I: A Practical Approach, IRL Press, Washington DC;
Liddell, J.E. and A.
Cryer (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 TRICH-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 TRICH, or any fragment or complement thereof, may be used for therapeutic purposes. In one aspect, modifications of gene expression can be achieved by designing complementary sequences or antisense molecules (DNA, RNA, PNA, or modified oligonucleotides) to the coding or regulatory regions of the gene encoding TRICH.
Such technology is well known in the art, and antisense oligonucleotides or larger fragments can be designed from various locations along the coding or control regions of sequences encoding TRICH.
(See, e.g., Agrawal, S., ed. (1996) Antisense Therapeutics, Humana Press Inc., Totawa NJ.) In therapeutic use, any gene delivery system suitable for introduction of the antisense sequences into appropriate target cells can be used. Antisense sequences can be delivered intracellularly in the form of an expression plasmid which, upon transcription, produces a sequence complementary to at least a portion of the cellular sequence encoding the target protein. (See, e.g., Slater, J.E. et al. (1998) J. Allergy Cli. Immunol. 102(3):469-475; and Scanlon, K.J. et al. (1995) 9(13):1288-1296.) Antisense sequences can also be introduced intracellularly through the use of viral vectors, such as retrovirus and adeno-associated virus vectors. (See, e.g., Miller, A.D. (1990) Blood 76:271; Ausubel, supra; Uckert, W. and W. Walther (1994) Pharmacol. Ther.
63(3):323-347.) Other gene delivery mechanisms include liposome-derived systems, artificial viral envelopes, and other systems known in the art. (See, e.g., Rossi, J.J. (1995) Br. Med. Bull.
51(1):217-225; Boado, R.J. et al. (1998) J. Pharm. Sci. 87(11):1308-1315; and Morris, M.C. et al. (1997) Nucleic Acids Res.
25(14):2730-2736.) In another embodiment of the invention, polynucleotides encoding TRICH may be used for somatic or germline gene therapy. Gene therapy may be performed to (i) correct a genetic deficiency (e.g., in the cases of severe combined immunodeficiency (SCID)-X1 disease characterized by X-linked inheritance (Cavazzana-Calvo, M. et al. (2000) Science 288:669-672), severe combined immunodeficiency syndrome associated with an inherited adenosine deaminase (ADA) deficiency (Blaese, R.M. et al. (1995) Science 270:475-480; Bordignon, C. et al. (1995) Science 270:470-475), cystic fibrosis (Zabner, J. et al. (1993) Cel175:207-216; Crystal, R.G. et al.
(1995) Hum. Gene Therapy 6:643-666; Crystal, R.G. et al. (1995) Hum. Gene Therapy 6:667-703), thalassamias, familial hypercholesterolemia, and hemophilia resulting from Factor VIII or Factor IX
deficiencies (Crystal, R.G. (1995) Science 270:404-410; Verma, LM. and N. Somia (1997) Nature 389:239-242)), (ii) express a conditionally lethal gene product (e.g., in the case of cancers which result from unregulated cell proliferation), or (iii) express a protein which affords protection against intracellular parasites (e.g., against human retroviruses, such as human immunodeficiency virus (HIV) (Baltimore, D. (1988) Nature 335:395-396; Poeschla, E. et al. (1996) Proc. Natl. Acad. Sci. USA.
93:11395-11399), hepatitis B or C virus (HBV, HCV); fungal parasites, such as Candida albicans and Paracoccidioides brasiliensis; and protozoan.parasites such as Plasmodium falciparum and Trypanosoma cruzi). In the case where a genetic deficiency in TRICH expression or regulation causes disease, the expression of TRICH from an appropriate population of transduced cells may alleviate the clinical manifestations caused by the genetic deficiency.
In a further embodiment of the invention, diseases or disorders caused by deficiencies in TRICH are treated by constructing mammalian expression vectors encoding TRICH
and introducing these vectors by mechanical means into TRICH-deficient cells. Mechanical transfer technologies for use with cells in vivo or ex vitro include (i) direct DNA microinjection into individual cells, (ii) ballistic gold particle delivery, (iii) liposome-mediated transfection, (iv) receptor-mediated gene transfer, and (v) the use of DNA transposons (Morgan, R.A. and W.F. Anderson (1993) Annu. Rev.
Biochem. 62:191-217; Ivics, Z. (1997) Cell 91:501-510; Boulay, J-L. and H. Recipon (1998) Curr. Opin. Biotechnol.
9:445-450).
Expression vectors that may be effective for the expression of TRICH include, but are not limited to, the PCDNA 3.1, EPITAG, PRCCMV2, PREP, PVAX vectors (Invitrogen, Carlsbad CA), PCMV-SCRIPT, PCMV-TAG, PEGSH/PERV (Stratagene, La Jolla CA), and PTET-OFF, PTET-ON, PTRE2, PTRE2-LUC, PTK HYG (Clontech, Palo Alto CA). TRICH may be expressed using (i) a constitutively active promoter, (e.g., from cytomegalovirus (CMV), Rous sarcoma virus (RSV), SV40 virus, thymidine kinase (TK), or /3-actin genes), (ii) an inducible promoter (e.g., the tetracycline-regulated promoter (Gossen, M. and H. Bujard (1992) Proc. Natl.
Acad. Sci. U.S.A.
89:5547-5551; Gossen, M. et al. (1995) Science 268:1766-1769; Rossi, F.M.V.
and H.M. Blau (1998) Curr. Opin. Biotechnol. 9:451-456), commercially available in the T-REX
plasmid (Invitrogen)); the ecdysone-inducible promoter (available in the plasmids PVGRXR and PIND;
Invitrogen); the FK506lrapamycin inducible promoter; or the RU486/mifepristone inducible promoter (Rossi, F.M. V.
and Blau, H.M. supra)), or (iii) a tissue-specific promoter or the native promoter of the endogenous gene encoding TRICH from a normal individual.
Commercially available liposome transformation kits (e.g., the PERFECT LIPID
TRANSFECTION KIT, available from Invitrogen) allow one with ordinary skill in the art to deliver polynucleotides to target cells in culture and require minimal effort to optimize experimental parameters. In the alternative, transformation is performed using the calcium phosphate method (Graham, F.L. and A.J. Eb (1973) Virology 52:456-467), or by electroporation (Neumann, E. et al.
(1982) EMBO J. 1:841-845). The introduction of DNA to primary cells requires modification of these standardized mammalian transfection protocols.
In another embodiment of the invention, diseases or disorders caused by genetic defects with respect to TRICH expression are treated by constructing a retrovirus vector consisting of (i) the polynucleotide encoding TRICH under the control of an independent promoter or the retrovirus long terminal repeat (LTR) promoter, (ii) appropriate RNA packaging signals, and (iii) a Rev-responsive element (RRE) along with additional retrovirus cis-acting RNA sequences and coding sequences required for efficient vector propagation. Retrovirus vectors (e.g., PFB and PFBNEO) are commercially available (Stratagene) and are based on published data (Riviere, I. et al. (1995) Proc.
Natl. Acad. Sci. U.S.A. 92:6733-6737), incorporated by reference herein. The vector is propagated in an appropriate vector producing cell line (VPCL) that expresses an envelope gene with a tropism for receptors on the target cells or a promiscuous envelope protein such as VSVg (Armentano, D. et al.
(1987) J. Virol. 61:1647-1650; Bender, M.A. et al. (1987) J. Virol. 61:1639-1646; Adam, M.A. and A.D. Miller (1988) J. Virol. 62:3802-3806; Dull, T. et al. (1998) J. Virol.
72:8463-8471; Zufferey, R.
et al. (1998) J. Virol. 72:9873-9880). U.S. Patent Number 5,910,434 to Rigg ("Method for obtaining retrovirus packaging cell lines producing high transducing efficiency retroviral supernatant") discloses a method for obtaining retrovirus packaging cell lines and is hereby incorporated by reference.
Propagation of retrovirus vectors, transduction of a population of cells (e.g., CD4+ T-cells), and the return of transduced cells to a patient are procedures well known to persons skilled in the art of gene therapy and have been well documented (Ranga, U. et al. (1997) J. Virol.
71:7020-7029; Bauer, G. et al. (1997) Blood 89:2259-2267; Bonyhadi, M.L. (1997) J. Virol. 71:4707-4716;
Ranga, U. et al.
(1998) Proc. Natl. Acad. Sci. U.S.A. 95:1201-1206; Su, L. (1997) Blood 89:2283-2290).
In the alternative, an adenovirus-based gene therapy delivery system is used to deliver polynucleotides encoding TRICH to cells which have one or more genetic abnormalities with respect to the expression of TRICH. The construction and packaging of adenovirus-based vectors are well known to those with ordinary skill in the art. Replication defective adenovirus vectors have proven to be versatile for importing genes encoding immunoregulatory proteins into intact islets in the pancreas (Csete, M.E. et al. (1995) Transplantation 27:263-268). Potentially useful adenoviral vectors are described in U.S. Patent Number 5,707,618 to Armentano ("Adenovirus vectors for gene therapy"), hereby incorporated by reference. For adenoviral vectors, see also Antinozzi, P.A. et al. (1999) Annu.
Rev. Nutr. 19:511-544 and Verma, LM. and N. Somia (1997) Nature 18:389:239-242, both incorporated by reference herein.
In another alternative, a herpes-based, gene therapy delivery system is used to deliver polynucleotides encoding TRICH to target cells which have one or more genetic abnormalities with respect to the expression of TRICH. The use of herpes simplex virus (HSV)-based vectors may be especially valuable for introducing TRICH to cells of the central nervous system, for which HSV has a tropism. The construction and packaging of herpes-based vectors are well known to those with ordinary skill in the art. A replication-competent herpes simplex virus (HSV) type 1-based vector has been used to deliver a reporter gene to the eyes of primates (Liu, X. et al.
(1999) Exp. Eye Res.169:385-395). The construction of a HSV-1 virus vector has also been disclosed in detail in U.S.
Patent Number 5,804,413 to DeLuca ("Herpes simplex virus strains for gene transfer"), which is hereby incorporated by reference. U.S. Patent Number 5,804,413 teaches the use of recombinant HSV
d92 which consists of a genome containing at least one exogenous gene to be transferred to a cell under the control of the appropriate promoter for purposes including human gene therapy. Also taught by this patent are the construction and use of recombinant HSV strains deleted for ICP4, ICP27 and ICP22.
For HSV vectors, see also Goins, W.F. et al. (1999) J. Virol. 73:519-532 and Xu, H. et al. (1994) Dev.
Biol. 163:152-161, hereby incorporated by reference. The manipulation of cloned herpesvirus sequences, the generation of recombinant virus following the transfection of multiple plasmids containing different segments of the large herpesvirus genomes, the growth and propagation of herpesvirus, and the infection of cells with herpesvirus are techniques well known to those of ordinary skill in the art.
In another alternative, an alphavirus (positive, single-stranded RNA virus) vector is used to deliver polynucleotides encoding TRICH to target cells. The biology of the prototypic alphavirus, Semliki Forest Virus (SFV), has been studied extensively and gene transfer vectors have been based on the SFV genome (Garoff, H. and K.-J. Li (1998) Curr. Opin. Biotechnol. 9:464-469). During alphavirus RNA replication, a subgenomic RNA is generated that normally encodes the viral capsid proteins. This subgenomic RNA replicates to higher levels than the full length genomic RNA, resulting in the overproduction of capsid proteins relative to the viral proteins with enzymatic activity (e.g., protease and polymerase). Similarly, inserting the coding sequence for TRICH
into the alphavirus genome in place of the capsid-coding region results in the production of a large number of TRICH-coding RNAs and the synthesis of high levels of TRICH in vector transduced cells. While alphavirus infection is typically associated with cell lysis within a few days, the ability to establish a persistent infection in hamster normal kidney cells (BHK 21) with a variant of Sindbis virus (SIN) indicates that the lytic replication of alphaviruses can be altered to suit the needs of the gene therapy application (Dryga, S.A. et al. (1997) Virology 228:74-83). The wide host range of alphaviruses will allow the introduction of TRICH into a variety of cell types. The specific transduction of a subset of cells in a population may require the sorting of cells prior to transduction. The methods of manipulating infectious cDNA clones of alphaviruses, performing alphavirus cDNA and RNA
transfections, and performing alphavirus infections, are well known to those with ordinary skill in the art.
Oligonucleotides derived from the transcription initiation site, e.g., between about positions -10 and +10 from the start site, may also be employed to inhibit gene expression.
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~ic 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 TRICH.
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 TRICH. 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' andlor 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.
An additional embodiment of the invention encompasses a method for screening for a compound which is effective in altering expression of a polynucleotide encoding TRICH.
Compounds which may be effective in altering expression of a specific polynucleotide may include, but are not limited to, oligonucleotides, antisense oligonucleotides, triple helix-forming oligonucleotides, transcription factors and other polypeptide transcriptional regulators, and non-macromolecular chemical entities which are capable of interacting with specific polynucleotide sequences. Effective compounds may alter polynucleotide expression by acting as either inhibitors or promoters of polynucleotide expression. Thus, in the treatment of disorders associated with increased TRICH expression or activity, a compound which specifically inhibits expression of the polynucleotide encoding TRICH may be therapeutically useful, and in the treament of disorders associated with decreased TRICH expression or activity, a compound which specifically promotes expression of the polynucleotide encoding TRICH may be therapeutically useful.
At least one, and up to a plurality, of test compounds may be screened for effectiveness in altering expression of a specific polynucleotide. A test compound may be obtained by any method commonly known in the art, including chemical modification of a compound known to be effective in altering polynucleotide expression; selection from an existing, commercially-available or proprietary library of naturally-occurring or non-natural chemical compounds; rational design of a compound based on chemical and/or structural properties of the target polynucleotide;
and selection from a library of chemical compounds created combinatorially or randomly. A sample comprising a polynucleotide encoding TRICH is exposed to at least one test compound thus obtained. The sample may comprise, for example, an intact or permeabilized cell, or an in vitro cell-free or reconstituted biochemical system. Alterations in the expression of a polynucleotide encoding TRICH are assayed by any method commonly known in the art. Typically, the expression of a specific nucleotide is detected by hybridization with a probe having a nucleotide sequence complementary to the sequence of the polynucleotide encoding TRICH. The amount of hybridization may be quantified, thus forming the basis for a comparison of the expression of the polynucleotide both with and without exposure to one or more test compounds. Detection of a change in the expression of a polynucleotide exposed to a test compound indicates that the test compound is effective in altering the expression of the polynucleotide. A screen for a compound effective in altering expression of a specific polynucleotide can be carried out, for example, using a Schizosaccharomyces pombe gene expression system (Atkins, D. et al. (1999) U.S. Patent No. 5,932,435; Arndt, G.M. et al.
(2000) Nucleic Acids Res. 28:E15) or a human cell line such as HeLa cell (Clarke, M.L. et al.
(2000) Biochem. Biophys.
Res. Commun. 268:8-13). A particular embodiment of the present invention involves screening a combinatorial library of oligonucleotides (such as deoxyribonucleotides, ribonucleotides, peptide nucleic acids, and modified oligonucleotides) for antisense activity against a specific polynucleotide sequence (Bruice, T.W. et al. (1997) U.S. Patent No. 5,686,242; Bruice, T.W.
et al. (2000) U.S.
Patent No. 6,022,691).
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.I~. et al. (1997) Nat.
Biotechnol. 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 composition which generally comprises an active ingredient formulated with a pharmaceutically acceptable excipient.
Excipients may include, for example, sugars, starches, celluloses, gums, and proteins. Various formulations are commonly known and are thoroughly discussed in the latest edition of Remin.~ton's Pharmaceutical Sciences (Maack Publishing, Easton PA). Such compositions may consist of TRICH, antibodies to TRICH, and mimetics, agonists, antagonists, or inhibitors of TRICH.
The compositions utilized in this invention may be administered by any number of routes including, but not limited to, oral, intravenous, intramuscular, infra-arterial, intramedullary, intrathecal, intraventricular, pulmonary, transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical, sublingual, or rectal means.
Compositions for pulmonary administration may be prepared in liquid or dry powder form.
These compositions are generally aerosolized immediately prior to inhalation by the patient. In the case of small molecules (e.g. traditional low molecular weight organic drugs), aerosol delivery of fast-acting formulations is well-known in the art. In the case of macromolecules (e.g.
larger peptides and proteins), recent developments in the field of pulmonary delivery via the alveolar region of the lung have enabled the practical delivery of.drugs such as insulin to blood circulation (see, e.g., Patton. J.S. et al., U.S.
Patent No. 5,997,848). Pulmonary delivery has the advantage of administration without needle injection, and obviates the need for potentially toxic penetration enhancers.
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.
Specialized forms of compositions may be prepared for direct intracellular delivery of macromolecules comprising TRICH or fragments thereof. For example, liposome preparations containing a cell-impermeable macromolecule may promote cell fusion and intracellular delivery of the macromolecule. Alternatively, TRICH or a fragment thereof may be joined to a short cationic N-terminal portion from the HIV Tat-1 protein. Fusion proteins thus generated have been found to transduce into the cells of all tissues, including the brain, in a mouse model system (Schwarze, S.R. et al. (1999) Science 285:1569-1572).
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, monkeys, 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 TRICH
or fragments thereof, antibodies of TRICH, and agonists, antagonists or inhibitors of TRICH, 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. 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 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 ~cg to 100,000 ~.g, 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 TRICH may be used for the diagnosis of disorders characterized by expression of TRICH, or in assays to monitor patients being treated with TRICH or agonists, antagonists, or inhibitors of TRICH.
Antibodies useful for diagnostic purposes may be prepared in the same manner as described above for therapeutics. Diagnostic assays for TRICH include methods which utilize the antibody and a label to detect TRICH 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 TRICH, including ELISAs, RIAs, and FACS, are known in the art and provide a basis for diagnosing altered or abnormal levels of TRICH expression. Normal or standard values for TRICH expression are established by combining body fluids or cell extracts taken from normal mammalian subjects, for example, human subjects, with antibodies to TRICH under conditions suitable for complex formation. The amount of standard complex formation may be quantitated by various methods, such as photometric means. Quantities of TRICH
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 TRICH may be used for diagnostic purposes. The polynucleotades 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 TRICH may be correlated with disease. The diagnostic assay may be used to determine absence, presence, and excess expression of TRICH, and to monitor regulation of TRICH levels during therapeutic intervention.
In one aspect, hybridization with PCR probes which are capable of detecting polynucleotide sequences, including genomic sequences, encoding TRICH or closely related molecules may be used to identify nucleic acid sequences which encode TRICH. 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 TRICH, 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 TRICH encoding sequences. The hybridization probes of the subject invention may be DNA or RNA and may be derived from the sequence of SEQ ID
N0:28-54 or from genomic sequences including promoters, enhancers, and introns of the TRICH
gene.
Means for producing specific hybridization probes for DNAs encoding TRICH
include the cloning of polynucleotide sequences encoding TRICH or TRICH 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 35S, or by enzymatic labels, such as alkaline phosphatase coupled to the probe via avidin/biotin coupling systems, and the like.
Polynucleotide sequences encoding TRICH may be used for the diagnosis of disorders associated with expression of TRICH. Examples of such disorders include, but are not limited to, a transport disorder such as akinesia, amyotrophic lateral sclerosis, ataxia telangiectasia, cystic fibrosis, Becker's muscular dystrophy, Bell's palsy, Charcot-Marie Tooth disease, diabetes mellitus, diabetes insipidus, diabetic neuropathy, Duchenne muscular dystrophy, hyperkalemic periodic paralysis, normokalemic periodic paralysis, Parkinson's disease, malignant hyperthermia, multidrug resistance, myasthenia gravis, myotonic dystrophy, catatonia, tardive dyskinesia, dystonias, peripheral neuropathy, cerebral neoplasms, prostate cancer, cardiac disorders associated with transport, e.g., angina, bradyarrythmia, tachyarrythmia, hypertension, Long QT syndrome, myocarditis, cardiomyopathy, nemaline myopathy, centronuclear myopathy, lipid myopathy, mitochondria) myopathy, thyrotoxic myopathy, ethanol myopathy, dermatomyositis, inclusion body myositis, infectious myositis, polymyositis, neurological disorders associated with transport, e.g., Alzheimer's disease, amnesia, bipolar disorder, dementia, depression, epilepsy, Tourette's disorder, paranoid psychoses, and schizophrenia, and other disorders associated with transport, e.g., neurofibromatosis, postherpetic neuralgia, trigeminal neuropathy, sarcoidosis, sickle cell anemia, Wilson's disease, cataracts, infertility, pulmonary artery stenosis, sensorineural autosomal deafness, hyperglycemia, hypoglycemia, Grave's disease, goiter, Cushing's disease, Addison's disease, glucose-galactose malabsorption syndrome, hypercholesterolemia, adrenoleukodystrophy, Zellweger syndrome, Menkes disease, occipital horn syndrome, von Gierke disease, cystinuria, iminoglycinuria, Hartup disease, and Fanconi disease; a neurological disorder such as epilepsy, ischemic cerebrovascular disease, stroke, cerebral neoplasms, Alzheimer's disease, Pick's disease, Huntington's disease, dementia, Parkinson's disease and other extrapyramidal disorders, amyotrophic lateral sclerosis and other motor neuron disorders, progressive neural muscular atrophy, retinitis pigmentosa, hereditary ataxias, multiple sclerosis and other demyelinating diseases, bacterial and viral meningitis, brain abscess, subdural empyema, epidural abscess, suppurative intracranial thrombophlebitis, myelitis and radiculitis, viral central nervous system disease, prion diseases including kuru, Creutzfeldt-Jakob disease, and Gerstmann-Straussler-Scheinker syndrome, fatal familial insomnia, nutritional and metabolic diseases of the nervous system, neurofibromatosis, tuberous sclerosis, cerebelloretinal hemangioblastomatosis, encephalotrigeminal syndrome, mental retardation and other developmental disorders of the central nervous system including Down syndrome, cerebral palsy, neuroskeletal disorders, autonomic nervous system disorders, cranial nerve disorders, spinal cord diseases, muscular dystrophy and other neuromuscular disorders, peripheral nervous system disorders, dermatomyositis and polymyositis, inherited, metabolic, endocrine, and toxic myopathies, myasthenia gravis, periodic paralysis, mental disorders including mood, anxiety, and schizophrenic disorders, seasonal affective disorder (SAD), akathesia, amnesia, catatonia, diabetic neuropathy, tardive dyskinesia, dystonias, paranoid psychoses, postherpetic neuralgia, Tourette's disorder, progressive supranuclear palsy, corticobasal degeneration, and familial frontotemporal dementia; a muscle disorder such as cardiomyopathy, myocarditis, Duchenne's muscular dystrophy, Becker's muscular dystrophy, myotonic dystrophy, central core disease, nemaline myopathy, centronuclear myopathy, lipid myopathy, mitochondria) myopathy, infectious myositis, polymyositis, dermatomyositis, inclusion body myositis, thyrotoxic myopathy, ethanol myopathy, angina, anaphylactic shock, arrhythmias, asthma, cardiovascular shock, Cushing's syndrome, hypertension, hypoglycemia, myocardial infarction, migraine, pheochromocytoma, and myopathies including encephalopathy, epilepsy, Kearns-Sayre syndrome, lactic acidosis, myoclonic disorder, ophthalmoplegia, and acid maltase deficiency (AMD, also known as Pompe's disease); and an immunological disorder such as acquired immunodeficiency syndrome (AIDS), Addison's disease, adult respiratory distress syndrome, allergies, ankylosing spondylitis, amyloidosis, anemia, asthma, atherosclerosis, autoimmune hemolytic anemia, autoimmune thyroiditis, autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED), bronchitis, cholecystitis, contact dermatitis, Crohn's disease, atopic dermatitis, dermatomyositis, diabetes mellitus, emphysema, episodic lymphopenia with lymphocytotoxins, erythroblastosis fetalis, erythema nodosum, atrophic gastritis, glomerulonephritis, Goodpasture's syndrome, gout, Graves' disease, Hashimoto's thyroiditis, hypereosinophilia, irritable bowel syndrome, multiple sclerosis, myasthenia gravis, myocardial or pericardial inflammation, osteoarthritis, osteoporosis, pancreatitis, polymyositis, psoriasis, Reiter's syndrome, rheumatoid arthritis, scleroderma, Sjogren's syndrome, systemic anaphylaxis, systemic lupus erythematosus, systemic sclerosis, thrombocytopenic purpura, ulcerative colitis, uveitis, Werner syndrome, complications of cancer, hemodialysis, and extracorporeal circulation, viral, bacterial, fungal, parasitic, protozoal, and helminthic infections, and trauma. The polynucleotide sequences encoding TRICH 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 TRICH expression.
Such qualitative or quantitative methods are well known in the art.
a In a particular aspect, the nucleotide sequences encoding TRICH may be useful in assays that detect the presence of associated disorders, particularly those mentioned above. The nucleotide sequences encoding TRICH 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 TRICH 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 TRICH, a normal or standard profile for expression is established. This may be accomplished by combining body fluids or Bell extracts taken from normal subjects, either animal or human, with a sequence, or a fragment thereof, encoding TRICH, 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 TRICH
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 TRICH, or a fragment of a polynucleotide complementary to the polynucleotide encoding TRICH, 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.
In a particular aspect, oligonucleotide primers derived from the polynucleotide sequences encoding TRICH may be used to detect single nucleotide polymorphisms (SNPs).
SNPs are substitutions, insertions and deletions that are a frequent cause of inherited or acquired genetic disease in humans. Methods of SNP detection include, but are not limited to, single-stranded conformation polymorphism (SSCP) and fluorescent SSCP (fSSCP) methods. In SSCP, oligonucleotide primers derived from the polynucleotide sequences encoding TRICH are used to amplify DNA using the polymerase chain reaction (PCR). The DNA may be derived, for example, from diseased or normal tissue, biopsy samples, bodily fluids, and the like. SNPs in the DNA cause differences in the secondary and tertiary structures of PCR products in single-stranded form, and these differences are detectable using gel electrophoresis in non-denaturing gels. In fSCCP, the oligonucleotide primers are fiuorescently labeled, which allows detection of the amplimers in high-throughput equipment such as DNA sequencing machines. Additionally, sequence database analysis methods, termed in silico SNP

(isSNP), are capable of identifying polymorphisms by comparing the sequence of individual overlapping DNA fragments which assemble into a common consensus sequence.
These computer-based methods filter out sequence variations due to laboratory preparation of DNA and sequencing errors using statistical models and automated analyses of DNA sequence chromatograms. In the alternative, SNPs may be detected and characterized by mass spectrometry using, for example, the high throughput MASSARRAY system (Sequenom, Inc., San Diego CA).
Methods which may also be used to quantify the expression of TRICH 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 or polynucleotide 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 elements on a microarray. The microarray can be used in transcript imaging techniques which monitor the relative expression levels of large numbers of genes simultaneously as described below. The microarray may also be used 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, to monitor progression/regression of disease as a function of gene expression, and to develop and monitor the activities of therapeutic agents in the treatment of disease. In particular, this information may be used to develop a pharmacogenomic profile of a patient in order to select the most appropriate and effective treatment regimen for that patient. For example, therapeutic agents which are highly effective and display the fewest side effects may be selected for a patient based on his/her pharmacogenomic profile.
In another embodiment, TRICH, fragments of TRICH, or antibodies specific for TRICH may be used as elements on a microarray. The microarray may be used to monitor or measure protein-protein interactions, drug-target interactions, and gene expression profiles, as described above.
A particular embodiment relates to the use of the polynucleotides of the present invention to generate a transcript image of a tissue or cell type. A transcript image represents the global pattern of gene expression by a particular tissue or cell type. Global gene expression patterns are analyzed by quantifying the number of expressed genes and their relative abundance under given conditions and at a given time. (See Seilhamer et al., "Comparative Gene Transcript Analysis,"
U.S. Patent Number 5,840,484, expressly incorporated by reference herein.) Thus a transcript image may be generated by hybridizing the polynucleotides of the present invention or their complements to the totality of transcripts or reverse transcripts of a particular tissue or cell type. In one embodiment, the hybridization takes place in high-throughput format, wherein the polynucleotides of the present invention or their complements comprise a subset of a plurality of elements on a microarray. The resultant transcript image would provide a profile of gene activity.
Transcript images may be generated using transcripts isolated from tissues, cell lines, biopsies, or other biological samples. The transcript image may thus reflect gene expression in vivo, as in the case of a tissue or biopsy sample, or in vitro, as in the case of a cell line.
Transcript images which profile the expression of the polynucleotides of the present invention may also be used in conjunction with in vitro model systems and preclinical evaluation of pharmaceuticals, as well as toxicological testing of industrial and naturally-occurring environmental compounds. All compounds induce characteristic gene expression patterns, frequently termed molecular fingerprints or toxicant signatures, which are indicative of mechanisms of action and toxicity (Nuwaysir, E.F. et al. (1999) Mol. Carcinog. 24:153-159; Steiner, S. and N.L.
Anderson (2000) Toxicol. Lett. 112-113:467-471, expressly incorporated by reference herein).
If a test compound has a signature similar to that of a compound with known toxicity, it is likely to share those toxic properties.
These fingerprints or signatures are most useful and refined when they contain expression information from a large number of genes and gene families. Ideally, a genome-wide measurement of expression provides the highest quality signature. Even genes whose expression is not altered by any tested compounds are important as well, as the levels of expression of these genes are used to normalize the rest of the expression data. The normalization procedure is useful for comparison of expression data after treatment with different compounds. While the assignment of gene function to elements of a toxicant signature aids in interpretation of toxicity mechanisms, knowledge of gene function is not necessary for the statistical matching of signatures which leads to prediction of toxicity. (See, for example, Press Release 00-02 from the National Institute of Environmental Health Sciences, released February 29, 2000, available at http://www.niehs.nih.gov/oc/news/toxchip.htm.) Therefore, it is important and desirable in toxicological screening using toxicant signatures to include all expressed gene sequences.
In one embodiment, the toxicity of a test compound is assessed by treating a biological sample containing nucleic acids with the test compound. Nucleic acids that are expressed in the treated biological sample are hybridized with one or more probes specific to the polynucleotides of the present invention, so that transcript levels corresponding to the polynucleotides of the present invention may be quantified. The transcript levels in the treated biological sample are compared with levels in an untreated biological sample. Differences in the transcript levels between the two samples are indicative of a toxic response caused by the test compound in the treated sample.

Another particular embodiment relates to the use of the polypeptide sequences of the present invention to analyze the proteome of a tissue or cell type. The term proteome refers to the global pattern of protein expression in a particular tissue or cell type. Each protein component of a proteome can be subjected individually to further analysis. Proteome expression patterns, or profiles, are analyzed by quantifying the number of expressed proteins and their relative abundance under given conditions and at a given time. A profile of a cell's proteome may thus be generated by separating and analyzing the polypeptides of a particular tissue or cell type. In one embodiment, the separation is achieved using two-dimensional gel electrophoresis, in which proteins from a sample are separated by isoelectric focusing in the first dimension, and then according to molecular weight by sodium dodecyl sulfate slab gel electrophoresis in the second dimension (Steiner and Anderson, supra). The proteins are visualized in the gel as discrete and uniquely positioned spots, typically by staining the gel with an agent such as Coomassie Blue or silver or fluorescent stains. The optical density of each protein spot is generally proportional to the level of the protein in the sample. The optical densities of equivalently positioned protein spots from different samples, for example, from biological samples either treated or untreated with a test compound or therapeutic agent, are compared to identify any changes in protein spot density related to the treatment. The proteins in the spots are partially sequenced using, for example, standard methods employing chemical or enzymatic cleavage followed by mass spectrometry.
The identity of the protein in a spot may be determined by comparing its partial sequence, preferably of at least 5 contiguous amino acid residues, to the polypeptide sequences of the present invention. In some cases, further sequence data may be obtained for definitive protein identification.
A proteomic profile may also be generated using antibodies specific for TRICH
to quantify the levels of TRICH expression. In one embodiment, the antibodies are used as elements on a microarray, and protein expression levels are quantified by exposing the microarray to the sample and detecting the levels of protein bound to each array element (Lueking, A. et al. (1999) Anal.
Biochem. 270:103-111;
Mendoze, L. G. et al. (1999) Biotechniques 27:778-788). Detection may be performed by a variety of methods known in the art, for example, by reacting the proteins in the sample with a thiol- or amino-reactive fluorescent compound and detecting the amount of fluorescence bound at each array element.
Toxicant signatures at the proteome level are also useful for toxicological screening, and should be analyzed in parallel with toxicant signatures at the transcript level.
There is a poor correlation between transcript and protein abundances for some proteins in some tissues (Anderson, N.L. and J.
Seilhamer (1997) Electrophoresis 18:533-537), so proteome toxicant signatures may be useful in the analysis of compounds which do not significantly affect the transcript image, but which alter the proteomic profile. In addition, the analysis of transcripts in body fluids is difficult, due to rapid degradation of mRNA, so proteomic profiling may be more reliable and informative in such cases.

In another embodiment, the toxicity of a test compound is assessed by treating a biological sample containing proteins with the test compound. Proteins that are expressed in the treated biological sample are separated so that the amount of each protein can be quantified. The amount of each protein is compared to the amount of the corresponding protein in an untreated biological sample. A difference in the amount of protein between the two samples is indicative of a toxic response to the test compound in the treated sample. Individual proteins are identified by sequencing the amino acid residues of the individual proteins and comparing these partial sequences to the polypeptides of the present invention.
In another embodiment, the toxicity of a test compound is assessed by treating a biological sample containing proteins with the test compound. Proteins from the biological sample are incubated with antibodies specific to the polypeptides of the present invention. The amount of protein recognized by the antibodies is quantified. The amount of protein in the treated biological sample is compared with the amount in an untreated biological sample. A difference in the amount of protein between the two samples is indicative of a toxic response to the test compound in the treated sample.
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.) Various types of microarrays are well known and thoroughly described in DNA Microarrays: A Practical Approach, M.
Schena, ed. (1999) Oxford University Press, London, hereby expressly incorporated by reference.
In another embodiment of the invention, nucleic acid sequences encoding TRICH
may be used to generate hybridization probes useful in mapping the naturally occurring genomic sequence. Either coding or noncoding sequences may be used, and in some instances, noncoding sequences may be preferable over coding sequences. For example, conservation of a coding sequence among members of a multi-gene family may potentially cause undesired cross hybridization during chromosomal mapping. 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.) Once mapped, the nucleic acid sequences of the invention may be used to develop genetic linkage maps, for example, which correlate the inheritance of a disease state with the inheritance of a particular chromosome region or restriction fragment length polymorphism (RFLP). (See, for example, Larder, E.S. and D. Botstein (1986) Proc. Natl. Acad. Sci. USA
83:7353-7357.) Fluorescent in situ hybridization (FISH) may be correlated with other physical and genetic map data. (See, e.g., Heinz-ITlrich, et al. (1995) in Meyers, supra, 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 TRICH on a physical map and a specific disorder, or a predisposition to a specific disorder, may help define the region of DNA associated with that disorder and thus may further positional cloning efforts.
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 exact chromosomal locus is not known.
This information is valuable to investigators searching for disease genes using positional cloning or other gene discovery techniques. Once the gene or genes responsible for a disease or syndrome have 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 ftirther investigation. (See, e.g., Gatti, R.A. et al. (1988) Nature 336:577-580.) The nucleotide sequence of the instant invention may also be used to detect differences in the chromosomal location due to translocatian, inversion, etc., among normal, carrier, or affected individuals.
In another embodiment of the invention, TRICH, 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 TRICH 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 TRICH, or fragments thereof, and washed. Bound TRICH is then detected by methods well known in the art.
Purified TRICH 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 TRICH specifically compete with a test compound for binding TRICH.
In this manner, antibodies can be used to detect the presence of any peptide which shares one or more antigenic determinants with TRICH.

In additional embodiments, the nucleotide sequences which encode TRICH 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 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. 60/172,000, U.S. Ser. No. 60/176,083, U.S. Ser. No.
60/177,332, U.S. Ser.
No. 60/178,572, U.S. Ser. No. 60/179,758, and U.S. Ser. No. 60/181,625, are expressly incorporated by reference herein.
EXAMPLES
I. Construction of cDNA Libraries Incyte cDNAs were derived from cDNA libraries described in the LIFESEQ GOLD
database (Incyte Genomics, Palo Alto CA) and shown in Table 4, column 5. 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 CsCl 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 inthe art. (See, e.g., Ausubel, 1997, su ra, 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., PBLUESCRIPT plasmid (Stratagene), PSPORT1 plasmid (Life Technologies), PCDNA2.1 plasmid (Invitrogen, Carlsbad CA), or pINCY (Incyte Genomics, Palo Alto CA).
Recombinant plasmids were transformed into competent E. coli cells including XL1-Blue, XL1-BlueMRF, or SOLR from Stratagene or DHSa, DH10B, or ElectroMAX DH10B from Life Technologies.
II. Isolation of cDNA Clones Plasmids obtained as described in Example I 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, QIAWELL 8 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 carried 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 Incyte cDNA recovered in plasmids as described in Example II were sequenced as follows.
Sequencing reactions were processed using standard methods or high-throughput instrumentation such as the ABI CATALYST 800 (PE Biosystems) 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 (PE Biosystems).
Electrophoretic separation of cDNA sequencing reactions and detection of labeled polynucleotides were carried out using the MEGABACE 1000 DNA sequencing system (Molecular Dynamics); the ABI
PRISM 373 or 377 sequencing system (PE Biosystems) 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, su ra, unit 7.7). Some of the cDNA sequences were selected for extension using the techniques disclosed in Example VIII.
The polynucleotide sequences derived from Incyte cDNAs were validated by removing vector, linker, and poly(A) sequences and by masking ambiguous bases, using algorithms and programs based on BLAST, dynamic programming, and dinucleotide nearest neighbor analysis. The Incyte cDNA
sequences or translations thereof 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 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, for example, Eddy, S.R. (1996) Curr. Opin. Struct. Biol. 6:361-365.) The queries were performed using programs based on BLAST, FASTA, BLIMPS, and HMMR. The Incyte cDNA
sequences were assembled to produce full length polynucleotide sequences.
Alternatively, GenBank cDNAs, GenBankESTs, stitched sequences, stretched sequences, or Genscan-predicted coding sequences (see Examples IV and V) were used to extend Incyte cDNA assemblages to full length.
Assembly was performed using programs based on Phred, Phrap, and Conned, and cDNA assemblages 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 polypeptide sequences which were subsequently analyzed by querying against databases such as the GenBank protein databases (genpept), SwissProt, BLOCKS, PRINTS, DOMO, PRODOM, Prosite, and hidden Markov model (HMM)-based protein family databases such as PFAM.
Full length polynucleotide sequences are also analyzed using MACDNASIS PRO software (Hitachi Software Engineering, South San Francisco CA) and LASERGENE software (DNASTAR).
Polynucleotide and polypeptide sequence alignments are generated using 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.
Table 7 summarizes the tools, programs, and algorithms used for the analysis and assembly of Incyte cDNA and full length sequences and provides applicable descriptions, references, and threshold parameters. The first column of Table 7 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 or the lower the probability value, the greater the identity between two sequences).
The programs described above for the assembly and analysis of full length polynucleotide and polypeptide sequences were also used to identify polynucleotide sequence fragments from SEQ ID
N0:28-54. Fragments from about 20 to about 4000 nucleotides which are useful in hybridization and amplification technologies are described in Table 4, column 4.
IV. Identification and Editing of Coding Sequences from Genomic DNA
Putative transporters and ion channels were initially identified by running the Genscan gene identification program against public genomic sequence databases (e.g., gbpri and gbhtg). Genscan is a general-purpose gene identification program which analyzes genomic DNA
sequences from a variety of organisms (See Burge, C. and S. Karlin (1997) J. Mol. Biol. 268:78-94, and Burge, C. and S. Karlin (1998) Curr. Opin. Struct. Biol. 8:346-354). The program concatenates predicted exons to form an assembled cDNA sequence extending from a methionine to a stop codon. The output of Genscan is a FASTA database of polynucleotide and polypeptide sequences. The maximum range of sequence for Genscan to analyze at once was set to 30 kb. To determine which of these Genscan predicted cDNA
sequences encode transporters and ion channels, the encoded polypeptides were analyzed by querying against PFAM models for transporters and ion channels. Potential transporters and ion channels were also identified by homology to Incyte cDNA sequences that had been annotated as transporters and ion channels. These selected Genscan-predicted sequences were then compared by BLAST analysis to the genpept and gbpri public databases. Where necessary, the Genscan-predicted sequences were then edited by comparison to the top BLAST hit from genpept to correct errors in the sequence predicted by Genscan, such as extra or omitted exons. BLAST analysis was also used to find any Incyte cDNA or public cDNA coverage of the Genscan-predicted sequences, thus providing evidence for transcription.
When Incyte cDNA coverage was available, this information was used to correct or confirm the Genscan predicted sequence. Full length polynucleotide sequences were obtained by assembling Genscan-predicted coding sequences with Incyte cDNA sequences andlor public cDNA sequences using the assembly process described in Example III. Alternatively, full length polynucleotide sequences were derived entirely from edited or unedited Genscan-predicted coding sequences.
V. Assembly of Genomic Sequence Data with cDNA Sequence Data "Stitched" Seguences Partial cDNA sequences were extended with exons predicted by the Genscan gene identification program described in Example IV. Partial cDNAs assembled as described in Example III were mapped to genomic DNA and parsed into clusters containing related cDNAs and Genscan exon predictions from one or more genomic sequences. Each cluster was analyzed using an algorithm based on graph theory and dynamic programming to integrate cDNA and genomic information, generating possible splice variants that were subsequently conf'~rmed, edited, or extended to create a full length sequence.
Sequence intervals in which the entire length of the interval was present on more than one sequence in the cluster were identified, and intervals thus identified were considered to be equivalent by transitivity.
For example, if an interval was present on a cDNA and two genomic sequences, then all three intervals were considered to be equivalent. This process allows unrelated but consecutive genomic sequences to be brought together, bridged by cDNA sequence. Intervals thus identified were then "stitched" together by the stitching algorithm in the order that they appear along their parent sequences to generate the longest possible sequence, as well as sequence variants. Linkages between intervals which proceed along one type of parent sequence (cDNA to cDNA or genomic sequence to genomic sequence) were given preference over linkages which change parent type (cDNA to genomic sequence). The resultant stitched sequences were translated and compared by BLAST analysis to the genpept and gbpri public databases. Incorrect exons predicted by Genscan were corrected by comparison to the top BLAST hit from genpept. Sequences were further extended with additional cDNA sequences, or by inspection of genomic DNA, when necessary.
"Stretched" Sequences Partial DNA sequences were extended to full length with an algorithm based on BLAST
analysis. First, partial cDNAs assembled as described in Example III were queried against public databases such as the GenBank primate, rodent, mammalian, vertebrate, and eukaryote databases using the BLAST program. The nearest GenBank protein homolog was then compared by BLAST analysis to either Incyte cDNA sequences or GenScan exon predicted sequences described in Example IV. A
chimeric protein was generated by using the resultant high-scoring segment pairs (HSPs) to map the translated sequences onto the GenBank protein homolog. Insertions or deletions may occur in the chimeric protein with respect to the original GenBank protein homolog. The GenBank protein homolog, the chimeric protein, or both were used as probes to search for homologous genomic sequences from the public human genome databases. Partial DNA sequences were therefore "stretched" or extended by the addition of homologous genomic sequences. The resultant stretched sequences were examined to determine whether it contained a complete gene.
VI. Chromosomal Mapping of TRICH Encoding Polynucleotides The sequences which were used to assemble SEQ ID N0:28-54 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 N0:28-54 were assembled into clusters of contiguous and overlapping sequences using assembly algorithms such as Phrap (Table 7). 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.
Map locations are represented by ranges, or intervals, or human chromosomes.
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. Human genome maps and other resources available to the public, such as the NCBI "GeneMap'99" World Wide Web site (http://www.ncbi.nlm.nih.gov/genemap~, can be employed to determine if previously identified disease genes map within or in proximity to the intervals indicated above.
VII. Analysis of Polynucleotide Expression 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 cDNA databases such as GenBank or LIFESEQ (Incyte Genomics). 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:
BLAST Score x Percent Identity 5 x minimum {length(Seq. 1), length(Seq. 2)}
The product score takes into account both the degree of similarity between two sequences and the length of the sequence match. The product score is a normalized value between 0 and 100, and is calculated as follows: the BLAST score is multiplied by the percent nucleotide identity and the product is divided by (5 times the length of the shorter of the two sequences). The BLAST score is calculated by assigning a score of +5 for every base that matches in a high-scoring segment pair (HSP), and -4 for every mismatch. Two sequences may share more than one HSP (separated by gaps).
If there is more than one HSP, then the pair with the highest BLAST score is used to calculate the product score. The product score represents a balance between fractional overlap and quality in a BLAST alignment. For example, a product score of 100 is produced only for 100% identity over the entire length of the shorter of the two sequences being compared. A product score of 70 is produced either by 100% identity and 70% overlap at one end, or by 88% identity and 100% overlap at the other. A
product score of 50 is produced either by 100% identity and 50% overlap at one end, or 79% identity and 100% overlap:
Alternatively, polynucleotide sequences encoding TRICH are analyzed with respect to the tissue sources from which they were derived. For example, some full length sequences are assembled, at least in part, with overlapping Incyte cDNA sequences (see Example IIn.
Each cDNA sequence is derived from a cDNA library constructed from a human tissue. Each human tissue is classified into one of the following organltissue categories: cardiovascular system; connective tissue; digestive system;
embryonic structures; endocrine system; exocrine glands; genitalia, female;
genitalia, male; germ cells;
heroic and immune system; liver; musculoskeletal system; nervous system;
pancreas; respiratory system; sense organs; skin; stomatognathic system; unclassified/mixed; or urinary tract. The number of libraries in each category is counted and divided by the total number of libraries across all categories.
Similarly, each human tissue is classified into one of the following diseaselcondition categories: cancer, cell line, developmental, inflammation, neurological, trauma, cardiovascular, pooled, and other, and the number of libraries in each category is counted and divided by the total number of libraries across all categories. The resulting percentages reflect the tissue- and disease-specific expression of cDNA
encoding TRICH. cDNA sequences and cDNA library/tissue information are found in the LIFESEQ
GOLD database (Incyte Genomics, Palo Alto CA).
VIII. Extension of TRICH Encoding Polynucleotides Full length polynucleotide sequences were also 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 was synthesized 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 Mg2+, (NH4)2S04, and 2-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 SI~+ 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 ~l PICOGREEN
quantitationreagent (0.25% (v/v) PICOGREEN; Molecular Probes, Eugene OR) dissolved in 1X TE
and 0.5 ~1 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 ~d to 10 /C1 aliquot of the reaction mixture was analyzed by electrophoresis on a 1 % agarose 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 WI), 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, and 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 (PE Biosystems).
In like manner, full length polynucleotide sequences are verified using the above procedure or are used to obtain 5' regulatory sequences using the above procedure along with oligonucleotides designed for such extension, and an appropriate genomic library.

IX. Labeling and Use of Individual Hybridization Probes Hybridization probes derived from SEQ ID N0:28-54 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-32P] adenosine triphosphate (Amersham Pharmacia Biotech), and T4 polynucleotide kinase (DuPont NEN, Boston MA). The labeled oligonucleotides are substantially purified using a SEPHADEX
G-25 superfine 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.
X. Microarrays The linkage or synthesis of array elements upon a microarray can be achieved utilizing photolithography, piezoelectric printing (ink jet printing, See, e.g., Baldeschweiler, supra.), mechanical microspotting technologies, and derivatives thereof. The substrate in each of the aforementioned technologies should be uniform and solid with a non-porous surface (Schena (1999), supra). Suggested substrates include silicon, silica, glass slides, glass chips, and silicon wafers. Alternatively, a procedure 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 using available methods and machines well known to those of ordinary skill in the art and may contain any appropriate number of elements. (See, e.g., Schena, M. et al.
(1995) Science 270:467-470;
Shalom D. et al. (1996) Genome Res. 6:639-645; Marshall, A. and J. Hodgson (1998) Nat. Biotechnol.
16:27-31.) Full length cDNAs, Expressed Sequence Tags (ESTs), or fragments or oligomers thereof may comprise the elements of the microarray. Fragments or oligomers suitable for hybridization can be selected using software well known in the art such as LASERGENE software (DNASTAR). The array elements are hybridized with polynucleotides in a biological sample. The polynucleotides in the biological sample are conjugated to a fluorescent label or other molecular tag for ease of detection.
After hybridization, nonhybridized nucleotides from the biological sample are removed, and a fluorescence scanner is used to detect hybridization at each array element.
Alternatively, laser desorbtion and mass spectrometry may be used for detection of hybridization.
The degree of complementarity and the relative abundance of each polynucleotide which hybridizes to an element on the microarray may be assessed. In one embodiment, microarray preparation and usage is described in detail below.
Tissue or Cell Samule Preparation Total RNA is isolated from tissue samples using the guanidinium thiocyanate method and poly(A)+ RNA is purified using the oligo-(dT) cellulose method. Each poly(A)+
RNA sample is reverse transcribed using MMLV reverse-transcriptase, 0.05 pg/~d oligo-(dT) primer (2lmer), 1X first strand buffer, 0.03 units/f.~l RNase inhibitor, 500 pM dATP, 500 pM dGTP, 500 ~M dTTP, 40 E.~M
dCTP, 40 ~M dCTP-Cy3 (BDS) or dCTP-Cy5 (Amersham Pharmacia Biotech). The reverse transcription reaction is performed in a 25 ml volume containing 200 ng poly(A)+ RNA with GEMBRIGHT kits (Incyte). Specific control poly(A)+ RNAs are synthesized by in vitro transcription from non-coding yeast genomic DNA. After incubation at 37° C for 2 hr, each reaction sample (one with Cy3 and another with Cy5 labeling) is treated with 2.5 ml of O.SM sodium hydroxide and incubated for 20 minutes at 85° C to the stop the reaction and degrade the RNA. Samples are purified using two successive CHROMA SPIN 30 gel filtration spin columns (CLONTECH
Laboratories, Inc.
(CLONTECH), Palo Alto CA) and after combining, both reaction samples are ethanol precipitated using 1 ml of glycogen (1 mg/ml), 60 ml sodium acetate, and 300 ml of 100%
ethanol. The sample is then dried to completion using a SpeedVAC (Savant Instruments Inc., Holbrook NY) and resuspended in 14 Ed SX SSC/0.2% SDS.
Microarrap Preparation Sequences of the present invention are used to generate array elements. Each array element is amplified from bacterial cells containing vectors with cloned cDNA inserts.
PCR amplification uses primers complementary to the vector sequences flanking the cDNA insert. Array elements are amplified in thirty cycles of PCR from an initial quantity of 1-2 ng to a final quantity greater than 5 pg. Amplified array elements are then purified using SEPHACRYL-400 (Amersham Pharmacia Biotech).
Purified array elements are immobilized on polymer-coated glass slides. Glass microscope slides (Corning) are cleaned by ultrasound in 0.1 % SDS and acetone, with extensive distilled water washes between and after treatments. Glass slides are etched in 4%
hydrofluoric acid (VWR
Scientific Products Corporation (VWR), West Chester PA), washed extensively in distilled water, and coated with 0.05% aminopropyl silane (Sigma) in 95% ethanol. Coated slides are cured in a 110°C
oven.
Array elements are applied to the coated glass substrate using a procedure described in US
Patent No. 5,807,522 , incorporated herein by reference. 1 i.d of the array element DNA, at an average concentration of 100 ngli.il, is loaded into the open capillary printing element by a high-speed robotic apparatus. The apparatus then deposits about 5 n1 of array element sample per slide.
Microarrays are UV-crosslinked using a STRATALINKER UV-crosslinker (Stratagene).
Microarrays are washed at room temperature once in 0.2% SDS and three times in distilled water.
Non-specific binding sites are blocked by incubation of microarrays in 0.2%
casein in phosphate buffered saline (PBS) (Tropix, Inc., Bedford MA) for 30 minutes at 60°
C followed by washes in 0.2% SDS and distilled water as before.
Hybridization Hybridization reactions contain 9 iil of sample mixture consisting of 0.2 ~g each of Cy3 and Cy5 labeled cDNA synthesis products in SX SSC, 0.2% SDS hybridization buffer.
The sample mixture is heated to 65 ° C for 5 minutes and is aliquoted onto the microarray surface and covered with an 1.8 cm2 coverslip. The arrays are transferred to a waterproof chamber having a cavity just slightly larger than a microscope slide. The chamber is kept at 100% humidity internally by the addition of 140 fil of SX SSC in a corner of the chamber. The chamber containing the arrays is incubated for about 6.5 hours at 60° C. The arrays are washed for 10 min at 45° C in a first wash buffer (1X SSC, 0.1 % SDS), three times for 10 minutes each at 45 ° C in a second wash buffer (0.1X SSC), and dried.
Detection Reporter-labeled hybridization complexes are detected with a microscope equipped with an Innova 70 mixed gas 10 W laser (Coherent, Inc., Santa Clara CA) capable of generating spectral lines at 488 nm for excitation of Cy3 and at 632 nm for excitation of CyS. The excitation laser light is focused on the array using a 20X microscope objective (Nikon, Inc., Melville NY). The slide containing the array is placed on a computer-controlled X-Y stage on the microscope and raster-scanned past the objective. The 1.8 cm x 1.8 cm array used in the present example is scanned with a resolution of 20 micrometers.
In two separate scans, a mixed gas multiline laser excites the two fluorophores sequentially.
Emitted light is split, based on wavelength, into two photomultiplier tube detectors (PMT 81477, Hamamatsu Photonics Systems, Bridgewater NJ~ corresponding to the two fluorophores. Appropriate filters positioned between the array and the photomultiplier tubes are used to filter the signals. The emission maxima of the fluorophores used are 565 nm for Cy3 and 650 nm for CyS. Each array is typically scanned twice, one scan per fluorophore using the appropriate filters at the laser source, although the apparatus is capable of recording the spectra from both fluorophores simultaneously.

The sensitivity of the scans is typically calibrated using the signal intensity generated by a cDNA control species added to the sample mixture at a known concentration. A
specific location on the array contains a complementary DNA sequence, allowing the intensity of the signal at that location to be correlated with a weight ratio of hybridizing species of 1:100,000. When two samples from different sources (e.g., representing test and control cells), each labeled with a different fluorophore, are hybridized to a single array for the purpose of identifying genes that are differentially expressed, the calibration is done by labeling samples of the calibrating cDNA
with the two fluorophores and adding identical amounts of each to the hybridization mixture.
The output of the photomultiplier tube is digitized using a 12-bit RTI-835H
analog-to-digital (A/D) conversion board (Analog Devices, Inc., Norwood MA) installed in an IBM-compatible PC
computer. The digitized data are displayed as an image where the signal intensity is mapped using a linear 20-color transformation to a pseudocolor scale ranging from blue (low signal) to red (high signal). The data is also analyzed quantitatively. Where two different fluorophores are excited and measured simultaneously, the data are first corrected for optical crosstalk (due to overlapping emission spectra) between the fluorophores using each fluorophore's emission spectrum.
A grid is superimposed over the fluorescence signal image such that the signal from each spot is centered in each element of the grid. The fluorescence signal within each element is then integrated to obtain a numerical value corresponding to the average intensity of the signal. The software used for signal analysis is the GEMTOOLS gene expression analysis program (Incyte).
XI. Complementary Polynucleotides Sequences complementary to the TRICH-encoding sequences, or any parts thereof, are used to detect, decrease, or inhibit expression of naturally occurring TRICH. 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 TRICH. 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 TRICH-encoding transcript.
XII. Expression of TRICH
Expression and purification of TRICH is achieved using bacterial or virus-based expression systems. For expression of TRICH 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 TRICH upon induction with isopropyl beta-D-thiogalactopyranoside (1PTG). Expression of TRICH in eukaryotic cells is achieved by infecting insect or mammalian cell lines with recombinant Autogrraphica californica nuclear polyhedrosis virus (AcMNPV), commonly known as baculovirus. The nonessential polyhedrin gene of baculovirus is replaced with cDNA encoding TRICH 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, TRICH 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 yponicum, 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 TRICH 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, su ra, ch. 10 and 16). Purified TRICH obtained by these methods can be used directly in the assays shown in Examples XVI, XVII, and XVIII, where applicable.
XIII. ~nctional Assays TRICH function is assessed by expressing the sequences encoding TRICH 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 (Life Technologies) and PCR3.1 (Invitrogen, Carlsbad CA), 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 ~cg 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 C ometry, Oxford, New York NY.
The influence of TRICH on gene expression can be assessed using highly purified populations of cells transfected with sequences encoding TRICH 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 e~ciently 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 TRICH and other genes of interest can be analyzed by northern analysis or microarray techniques.
XIV. Production of TRICH Specific Antibodies TRICH 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 TRICH 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 (PE Biosystems) 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-KL,H
complex in complete Freund's adjuvant. Resulting antisera are tested for antipeptide and anti-TRICH

activity by, for example, binding the peptide or TRICH to a substrate, blocking with 1 % BSA, reacting with rabbit antisera, washing, and reacting with radio-iodinated goat anti-rabbit IgG.
XV. Purification of Naturally Occurring TRICH Using Specific Antibodies Naturally occurring or recombinant TRICH is substantially purified by immunoaffinity chromatography using antibodies specific for TRICH. An immunoaffinity column is constructed by covalently coupling anti-TRICH 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 TRICH are passed over the immunoaffinity column, and the column is washed under conditions that allow the preferential absorbance of TRICH (e.g., high ionic strength buffers in the presence of detergent). The column is eluted under conditions that disrupt antibody/TRICH 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 TRICH is collected.
XVI. Identification of Molecules Which Interact with TRICH
Molecules which interact with TRICH may include transporter substrates, agonists or antagonists, modulatory proteins such as G(37 proteins (Reimann, supra) or proteins involved in TRICH
localization or clustering such as MAGUKs (Craven, supra). TRICH, or biologically active fragments thereof, are labeled with lzsl 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 multi-well plate are incubated with the labeled TRICH, washed, and any wells with labeled TRICH
complex are assayed. Data obtained using different concentrations of TRICH axe used to calculate values for the number, affinity, and association of TRICH with the candidate molecules.
Alternatively, proteins that interact with TRICH are isolated using the yeast 2-hybrid system (Fields, S. and O. Song (1989) Nature 340:245-246). TRICH, or fragments thereof, are expressed as fusion proteins with the DNA binding domain of Gal4 or lexA and potential interacting proteins are expressed as fusion proteins with an activation domain. Interactions between the TRICH fusion protein and the reconstitutes a transactivation function that is observed by expression of a reporter gene. Yeast 2-hybrid systems are commercially available, and methods for use of the yeast 2-hybrid system with ion channel proteins are discussed in Niethammer, M. and M. Sheng (1998, Meth.
Enzymol. 293:104-122).
TRICH may also be used in the PATHCALLING process (CuraGen Corp., New Haven CT) which employs the yeast two-hybrid system in a high-throughput manner to determine all interactions between the proteins encoded by two large libraries of genes (Nandabalan, K.
et al. (2000) U.S. Patent No. 6,057,101 ).
Potential TRICH agonists or antagonists may be tested for activation or inhibition of TRICH

ion channel activity using the assays described in section XVIII.
XVII. Demonstration of TRICH Activity Ion channel activity of TRICH is demonstrated using an electrophysiological assay for ion conductance. TRICH can be expressed by transforming a mammalian cell line such as COS7, HeLa or CHO with a eukaryotic expression vector encoding TRICH. Eukaryotic expression vectors are commercially available, and the techniques to introduce them into cells are well known to those skilled in the art. A second plasmid which expresses any one of a number of marker genes, such as 13-galactosidase, is co-transformed into the cells to allow rapid identification of those cells which have taken up and expressed the foreign DNA. The cells are incubated for 48-72 hours after transformation under conditions appropriate for the cell line to allow expression and accumulation of TRICH and 13-galactosidase.
Transformed cells expressing 13-galactosidase are stained blue when a suitable colorimetric substrate is added to the culture media under conditions that are well known in the art. Stained cells are tested for differences in membrane conductance by electrophysiological techniques that are well known in the art. Untransformed cells, and/or cells transformed with either vector sequences alone or 13-galactosidase sequences alone, are used as controls and tested in parallel.
Cells expressing TRICH
will have higher anion or canon conductance relative to control cells. The contribution of TRICH to conductance can be confirmed by incubating the cells using antibodies specific for TRICH. The antibodies will bind to the extracellular side of TRICH, thereby blocking the pore in the ion channel, and the associated conductance.
Alternatively, ion channel activity of TRICH is measured as current flow across a TRICH-containing Xenopus laevis oocyte membrane using the two-electrode voltage-clamp technique (Ishi et al., supra; Jegla, T. and L. Salkoff (1997) J. Neurosci. 17:32-44). TRICH is subcloned into an appropriate Xeno us oocyte expression vector, such as pBF, and 0.5-5 ng of mRNA is injected into mature stage IV oocytes. Injected oocytes are incubated at 18°C for 1-5 days. Inside-out macropatches are excised into an intracellular solution containing 116 mM K-gluconate, 4 mM KCl, and 10 mM Hepes (pH 7.2). The intracellular solution is supplemented with varying concentrations of the TRICH mediator, such as cAMP, cGMP, or Ca+2 (in the form of CaCl2), where appropriate.
Electrode resistance is set at 2-5 MSS and electrodes are filled with the intracellular solution lacking mediator. Experiments are performed at room temperature from a holding potential of 0 mV. Voltage ramps (2.5 s) from -100 to 100 mV are acquired at a sampling frequency of 500 Hz. Current measured is proportional to the activity of TRICH in the assay.
Transport activity of TRICH is assayed by measuring uptake of labeled substrates into Xenopus laevis oocytes. Oocytes at stages V and VI are injected with TRICH
mRNA (10 ng per oocyte) and incubated for 3 days at 18°C in OR2 medium (82.SmM NaCl, 2.5 mM KCl, 1mM CaCl2, 1mM MgCl2, 1mM Na2HP04, 5 mM Hepes, 3.8 mM NaOH , SOpg/ml gentamycin, pH 7.8) to allow expression of TRICH. Oocytes are then transferred to standard uptake medium (100mM NaCl, 2 mM
KCl, 1mM CaCl2, 1mM MgCl2, 10 mM Hepes/Tris pH 7.5). Uptake of various substrates (e.g., amino acids, sugars, drugs, ions, and neurotransmitters) is initiated by adding labeled substrate (e.g.
radiolabeled with 3H, fluorescently labeled with rhodamine, etc.) to the oocytes. After incubating for 30 minutes, uptake is terminated by washing the oocytes three times in Na+-free medium, measuring the incorporated label, and comparing with controls. TRICH activity is proportional to the level of internalized labeled substrate.
ATPase activity associated with TRICH can be measured by hydrolysis of radiolabeled ATP-[Y szP], separation of the hydrolysis products by chromatographic methods, and quantitation of the recovered 32P using a scintillation counter. The reaction mixture contains ATP-[y-32P] and varying amounts of TRICH in a suitable buffer incubated at 37 ° C for a suitable period of time. The reaction is terminated by acid precipitation with trichloroacetic acid and then neutralized with base, and an aliquot of the reaction mixture is subjected to membrane or filter paper-based chromatography to separate the reaction products. The amount of 32P liberated is counted in a scintillation counter. The amount of radioactivity recovered is proportional to the ATPase activity of TRICH in the assay.
XVIII. Identification of TRICH Agonists and Antagonists TRICH is expressed in a eukaryotic cell line such as CHO (Chinese Hamster Ovary) or HEK
(Human Embryonic Kidney) 293. Ion channel activity of the transformed cells is measured in the presence and absence of candidate agonists or antagonists. Ion channel activity is assayed using patch clamp methods well known in the art or as described in Example XVII.
Alternatively, ion channel activity is assayed using fluorescent techniques that measure ion flux across the cell membrane (Velicelebi, G. et al. (1999) Meth. Enzymol. 294:20-47; West, M.R. and C.R.
Molloy (1996) Anal.
Biochem. 241:51-58). These assays may be adapted for high-throughput screening using microplates.
Changes in internal ion concentration are measured using fluorescent dyes such as the Ca2+ indicator Fluo-4 AM, sodium-sensitive dyes such as SBFI and sodium green, or the Cl-indicator MQAE (all available from Molecular Probes) in combination with the FLIPR fluorimetric plate reading system (Molecular Devices). In a more generic version of this assay, changes in membrane potential caused by ionic flux across the plasma membrane are measured using oxonyl dyes such as DiBAC4 (Molecular Probes). DiBAC4 equilibrates between the extracellular solution and cellular sites according to the cellular membrane potential. The dye's fluorescence intensity is 20-fold greater when bound to hydrophobic intracellular sites, allowing detection of DiBAC4 entry into the cell (Gonzalez, J.E. and P.A. Negulescu (1998) Curr. Opin. Biotechnol. 9:624-631). Candidate agonists or antagonists may be selected from known ion channel agonists or antagonists, peptide libraries, or combinatorial chemical libraries.
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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<110> INCYTE GENOMICS, INC.
BAUGHN, Mariah R.
BURFORD, Neil AU-YOUNG, Janice LU, Dyung Aina M.
YANG, Junming REDDY, Roopa LAL, Preeti HILLMAN, Jennifer L.
AZIMZAI, Yalda YUE, Henry NGUYEN, Danniel B.
YAO, Monique G.
GANDHI, Ameeni R.
TANG, Y. Tom KHAN, Farrah A.
<120> TRFNSPORTERS AND ION CHANNELS
<130> PI-0005 PCT
<140> To Be Assigned <141> Herewith <150> 60/172,000; 60/176,083; 60/177,332; 60/178,572; 60/179,758;
60/181,625 <151> 1999-12-23; 2000-01-14; 2000-01-21; 2000-01-28; 2000-02-02;

<160> 54 <170> PERL Program <210> 1 <211> 477 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1416107CD1 <400> 1 Met Thr Pro Glu Asp Pro Glu Glu Thr Gln Pro Leu Leu Gly Pro Pro Gly Gly Ser Ala Pro Arg Gly Arg Arg Val Phe Leu Ala Ala Phe Ala Ala Ala Leu Gly Pro Leu Ser Phe Gly Phe Ala Leu Gly Tyr Ser Ser Pro Ala Ile Pro Ser Leu Gln Arg Ala Ala Pro Pro Ala Pro Arg Leu Asp Asp Ala Ala Ala Ser Trp Phe Gly Ala Val Val Thr Leu Gly Ala Ala Ala Gly Gly Val Leu Gly Gly Trp Leu Val Asp Arg Ala Gly Arg Lys Leu Ser Leu Leu Leu Cys Ser Val Pro Phe Val Ala Gly Phe Ala Val Ile Thr Ala Ala Gln Asp Val Trp Met Leu Leu Gly Gly Arg Leu Leu Thr Gly Leu Ala Cys Gly Val Ala Ser Leu Val Ala Pro Val Tyr Ile Ser G1u Ile Ala Tyr Pro Ala Val Arg Gly Leu Leu Gly Ser Cys Val Gln Leu Met Val Val Val G1y Ile Leu Leu Ala Tyr Leu Ala Gly Trp Val Leu Glu Trp Arg Trp Leu Ala Val Leu Gly Cys Val Pro Pro Ser Leu Met Leu Leu Leu Met Cys Phe Met Pro Glu Thr Pro Arg Phe Leu Leu Thr Gln His Arg Arg Gln Glu Ala Met A1a Ala Leu Arg Phe Leu Trp Gly Ser Glu Gln Gly Trp Glu Asp Pro Pro Ile G1y Ala Glu Gln Ser Phe His Leu Ala Leu Leu Arg Gln Pro Gly Ile Tyr Lys Pro Phe Ile Ile Gly Val Ser Leu Met Ala Phe Gln Gln Leu Ser Gly Val Asn Ala Val Met Phe Tyr Ala Glu Thr Ile Phe Glu Glu Ala Lys Phe Lys Asp Ser Ser Leu Ala Ser Val Val Val Gly Val Ile Gln Val Leu Phe Thr Ala Val Ala Ala Leu Ile Met Asp Arg Ala Gly Arg Arg Leu Leu Leu Val Leu Ser Gly Val Val Met Val Phe Ser Thr Ser Ala Phe Gly Ala Tyr Phe Lys Leu Thr Gln Gly Gly Pro Gly Asn Ser Ser His Val Ala Ile Ser A1a Pro Val Ser Ala Gln Pro Val Asp Ala Ser Val Gly Leu Ala Trp Leu Ala Val Gly Ser Met Cys Leu Phe Ile Ala Gly Phe Ala Val Gly Trp Gly Pro Ile Pro Trp Leu Leu Met Ser Glu Ile Phe Pro Leu His Val Lys Gly Val Ala Thr Gly Ile Cys Va1 Leu Thr Asn Trp Leu Met Ala Phe Leu Val Thr Lys Glu Phe Ser Ser Leu Met Glu Val Leu Arg Pro Tyr Gly Ala Phe Trp Leu Ala Ser Ala Phe Cys Ile Phe Ser Val Leu Phe Thr Leu Phe Cys Val Pro Glu Thr Lys Gly Lys Thr Leu Glu Gln Ile Thr Ala His Phe Glu Gly Arg <210> 2 <211> 498 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 1682513CD1 <400> 2 Met Arg Arg Gln Asp Ser Arg Gly Asn Thr Val Leu His Ala Leu Val Ala Ile Ala Asp Asn Thr Arg Glu Asn Thr Lys Phe Val Thr Lys Met Tyr Asp Leu Leu Leu Leu Lys Cys Ala Arg Leu Phe Pro Asp Ser Asn Leu Glu Ala Val Leu Asn Asn Asp G1y Leu Ser Pro Leu Met Met Ala Ala Lys Thr Gly Lys Ile Gly Asn Arg His Glu Met Leu Ala Val Glu Pro Ile Asn Glu Leu Leu Arg Asp Lys Trp Arg Lys Phe Gly Ala Val Ser Phe Tyr Ile Asn Val Val Ser Tyr Leu Cys Ala Met Val Ile Phe Thr Leu Thr Ala Tyr Tyr Gln Pro Leu Glu Gly Thr Pro Pro Tyr Pro Tyr Arg Thr Thr Val Asp Tyr Leu Arg Leu Ala Gly Glu Val Ile Thr Leu Phe Thr Gly Val Leu Phe Phe Phe Thr Asn Ile Lys Asp Leu Phe Met Lys Lys Cys Pro Gly Val Asn Ser Leu Phe Ile Asp Gly Ser Phe Gln Leu Leu Tyr Phe Ile Tyr Ser Val Leu Val Ile Val Ser Ala Ala Leu Tyr Leu Ala Gly Ile G1u Ala Tyr Leu Ala Va1 Met Val Phe Ala Leu Val Leu Gly Trp Met Asn Ala Leu Tyr Phe Thr Arg Gly Leu Lys Leu Thr Gly Thr Tyr Ser Ile Met Ile Gln Lys Ile Leu Phe Lys Asp Leu Phe Arg Phe Leu Leu Val Tyr Leu Leu Phe Met Ile Gly Tyr Ala Ser Ala Leu Va1 Ser Leu Leu Asn Pro Cys Ala Asn Met Lys Val Cys Asn Gly Asp Gln Thr Asn Cys Thr Val Pro Thr Tyr Pro Ser Cys Arg Asp Ser Glu Thr Phe Ser Thr Phe Leu Leu Asp Leu Phe Lys Leu Thr Ile Gly Met Gly Asp Leu Glu Met Leu Ser Ser Thr Lys Tyr Pro Val Val Phe Ile Ile Leu Leu Val Thr Tyr Ile Ile Leu Thr Phe Val Leu Leu Leu Asn Met Leu Ile Ala Leu Met Gly Glu Thr Val Gly Gln Val Ser Lys Glu Ser Lys His Ile Trp Lys Leu Gln Trp Ala Thr Thr Ile Leu Asp Ile Glu Arg Ser Phe Pro Val Phe Leu Arg Lys Ser Phe Arg Ser Gly Glu Met Val Thr Val Gly Lys Ser Ser Asp Gly Thr Pro Asp Arg Arg Trp Cys Phe Arg Val Asp Glu Val Asn Trp Ser His Trp Asn Gln Asn Leu Gly Ile Ile Asn Glu Asp Pro Gly Lys Asn Glu Thr Tyr Gln Tyr Tyr Gly Phe Ser His Thr Val Gly Arg Leu Arg Arg Asp Arg Trp Ser Ser Val Val Pro Arg Val Val Glu Leu Asn Lys Asn Ser Asn Pro Asp Glu Val Val Val Pro Leu Asp Ser Thr Gly Asn Pro Arg Cys Asp Gly His Gln Gln Gly Tyr Pro Arg Lys Trp Arg Thr Asp Asp Ala Pro Leu <210> 3 <211> 764 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 2446438CD1 <400> 3 Met Thr Ser Pro Ser Ser Ser Pro Val Phe Arg Leu Glu Thr Leu Asp Ala Gly G1n Glu Asp Gly Ser Glu Ala Asp Arg Gly Lys Leu Asp Phe Gly Ser Gly Leu Pro Pro Met Glu Ser Gln Phe Gln Gly Glu Asp Arg Lys Phe Ala Pro Gln Ile Arg Val Asn Leu Asn Tyr Arg Lys GIy Thr Gly Ala Ser Gln Pro Asp Pro Asn Arg Phe Asp Arg Asp Arg Leu Phe Asn Ala Val Ser Arg Gly Val Pro Glu Asp Leu Ala Gly Leu Pro Glu Tyr Leu Ser Lys Thr Ser Lys Tyr Leu Thr Asp Ser Glu Tyr Thr Glu Gly Ser Thr Gly Lys Thr Cys Leu Met Lys Ala Val Leu Asn Leu Lys Asp Gly Val Asn Ala Cys Ile Leu Pro Leu Leu Gln Ile Asp Arg Asp Ser Gly Asn Pro Gln Pro Leu Val Asn Ala Gln Cys Thr Asp Asp Tyr Tyr Arg Gly His Ser A1a Leu His Ile Ala Ile Glu Lys Arg Ser Leu Gln Cys Val Lys Leu Leu Val Glu Asn Gly Ala Asn Val His Ala Arg Ala Cys Gly Arg Phe Phe Gln Lys Gly Gln Gly Thr Cys Phe Tyr Phe Gly Glu Leu Pro Leu Ser Leu Ala Ala Cys Thr Lys Gln Trp Asp Val Val Ser Tyr Leu Leu Glu Asn Pro His Gln Pro Ala Ser Leu Gln Ala Thr Asp Ser Gln Gly Asn Thr Val Leu His Ala Leu Val Met Ile Ser Asp Asn Ser Ala Glu Asn Ile Ala Leu Val Thr Ser Met Tyr Asp Gly Leu Leu Gln Ala Gly Ala Arg Leu Cys Pro Thr Val Gln Leu Glu Asp Ile Arg Asn Leu Gln Asp Leu Thr Pro Leu Lys Leu Ala Ala Lys GIu Gly Lys Ile Glu Ile Phe Arg His Ile Leu Gln Arg Glu Phe Ser Gly Leu Ser His Leu Ser Arg Lys Phe Thr Glu Trp Cys Tyr Gly Pro Val Arg Val Ser Leu Tyr Asp Leu Ala Ser Val Asp Ser Cys Glu Glu Asn Ser Val Leu Glu Ile Ile A1a Phe His Cys Lys Ser Pro His Arg His Arg Met Val Val Leu Glu Pro Leu Asn Lys Leu Leu Gln Ala Lys Trp Asp Leu Leu Ile Pro Lys Phe Phe Leu Asn Phe Leu Cys Asn Leu Ile Tyr Met Phe Ile Phe Thr Ala Val Ala Tyr His Gln Pro Thr Leu Lys Lys Gln Ala Ala Pro His Leu Lys Ala Glu Val Gly Asn Ser Met Leu Leu Thr Gly His Ile Leu Ile Leu Leu Gly Gly Ile Tyr Leu Leu Val Gly Gln Leu Trp Tyr Phe Trp Arg Arg His Val Phe Ile Trp Ile Ser Phe Ile Asp Ser Tyr Phe Glu Ile Leu Phe Leu Phe G1n Ala Leu Leu Thr Val Val Ser Gln Val Leu Cys Phe Leu Ala Ile Glu Trp Tyr Leu Pro Leu Leu Val Ser Ala Leu Val Leu Gly Trp Leu Asn Leu Leu Tyr Tyr Thr Arg Gly Phe Gln His Thr Gly Ile Tyr Ser Val Met Ile Gln Lys Val Ile Leu Arg Asp Leu Leu Arg Phe Leu Leu Ile Tyr Leu Val Phe Leu Phe Gly Phe Ala Val Ala Leu Val Ser Leu Ser Gln Glu Ala Trp Arg Pro Glu Ala Pro Thr Gly Pro Asn Ala Thr G1u Ser Val Gln Pro Met Glu Gly G1n Glu Asp Glu Gly Asn Gly Ala Gln Tyr Arg Gly Ile Leu Glu Ala Ser Leu Glu Leu Phe Lys Phe Thr Ile Gly Met Gly Glu Leu Ala Phe Gln Glu Gln Leu His Phe Arg Gly Met Val Leu Leu Leu Leu Leu A1a Tyr Val Leu Leu Thr Tyr Ile Leu Leu Leu Asn Met Leu Ile Ala Leu Met Ser Glu Thr Val Asn Ser Val Ala Thr Asp Ser Trp Ser Ile Trp Lys Leu Gln Lys Ala Ile Ser Val Leu Glu Met Glu Asn Gly Tyr Trp Trp Cys Arg Lys Lys G1n Arg Ala Gly Val Met Leu Thr Val Gly Thr Lys Pro Asp Gly Ser Pro Asp Glu Arg Trp Cys Phe Arg Val Glu Glu Val Asn Trp Ala Ser Trp Glu Gln Thr Leu Pro Thr Leu Cys Glu Asp Pro Ser Gly Ala Gly Val Pro Arg Thr Leu Glu Asn Pro Val Leu Ala Ser Pro Pro Lys Glu Asp Glu Asp Gly Ala Ser Glu Glu Asn Tyr Val Pro Val Gln Leu Leu Gln Ser Asn <210> 4 <211> 255 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 2817822CD1 <400> 4 Met Trp Gln Gly Cys Ala Val Glu Arg Pro Va1 Gly Arg Met Thr Ser Gln Thr Pro Leu Pro Gln Ser Pro Arg Pro Arg Arg Pro Thr Met Ser Thr Val Val Glu Leu Asn Val Gly Gly Glu Phe His Thr Thr Thr Leu Gly Thr Leu Arg Lys Phe Pro Gly Ser Lys Leu Ala Glu Met Phe Ser Ser Leu Ala Lys Ala Ser Thr Asp Ala Glu Gly Arg Phe Phe Ile Asp Arg Pro Ser Thr Tyr Phe Arg Pro Ile Leu Asp Tyr Leu Arg Thr Gly Gln Val Pro Thr Gln His Ile Pro Glu Val Tyr Arg Glu Ala Gln Phe Tyr Glu Ile Lys Pro Leu Val Lys Leu Leu Glu Asp Met Pro Gln Ile Phe Gly Glu Gln Val Ser Arg Lys Gln Phe Leu Leu Gln Val Pro Gly Tyr Ser Glu Asn Leu Glu Leu Met Val Arg Leu Ala Arg Ala G1u Ala Ile Thr Ala Arg Lys Ser Ser Val Leu Val Cys Leu Val G1u Thr Glu Glu Gln Asp Ala Tyr Tyr Ser Glu Val Leu Cys Phe Leu Gln Asp Lys Lys Met Phe Lys Ser Val Val Lys Phe Gly Pro Trp Lys Ala Val Leu Asp Asn Ser Asp Leu Met His Cys Leu Glu Met Asp Ile Lys Ala Gln Gly Tyr Lys Val Phe Ser Lys Phe Tyr Leu Thr Tyr Pro Thr Lys Arg Asn Glu Phe His Phe Asn Ile Tyr Ser Phe Thr Phe Thr Trp Trp <210> 5 <211> 584 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 4009329CD1 <400> 5 Met Ala Gly Arg Arg Leu Asn Leu Arg Trp Ala Leu Ser Val Leu Cys Val Leu Leu Met Ala Glu Thr Val Ser Gly Thr Arg Gly Ser Ser Thr Gly Ala His Ile Ser Pro Gln Phe Pro Ala Ser Gly Val Asn Gln Thr Pro Val Val Asp Cys Arg Lys Val Cys Gly Leu Asn Val Ser Asp Arg Cys Asp Phe Ile Arg Thr Asn Pro Asp Cys His Ser Asp Gly Gly Tyr Leu Asp Tyr Leu Glu Gly Ile Phe Cys His Phe Pro Pro Ser Leu Leu Pro Leu Ala Val Thr Leu Tyr Val Ser Trp Leu Leu Tyr Leu Phe Leu Ile Leu Gly Val Thr Ala Ala Lys Phe Phe Cys Pro Asn Leu Ser A1a Ile Ser Thr Thr Leu Lys Leu Ser His Asn Val Ala Gly Val Thr Phe Leu Ala Phe Gly Asn Gly Ala Pro Asp Ile Phe Ser Ala Leu Val Ala Phe Ser Asp Pro His Thr Ala Gly Leu Ala Leu G1y Ala Leu Phe Gly Ala Gly Val Leu Val Thr Thr Val Val Ala Gly Gly Ile Thr Ile Leu His Pro Phe Met Ala Ala Ser Arg Pro Phe Phe Arg Asp Ile Val Phe Tyr Met Val Ala Val Phe Leu Thr Phe Leu Met Leu Phe Arg Gly Arg Val Thr Leu A1a Trp Ala Leu Gly Tyr Leu Gly Leu Tyr Val Phe Tyr Val Val Thr Val Ile Leu Cys Thr Trp Ile Tyr Gln Arg Gln Arg Arg Gly Ser Leu Phe Cys Pro Met Pro Val Thr Pro Glu Ile Leu Ser Asp Ser Glu Glu Asp Arg Val Ser Ser Asn Thr Asn Ser Tyr Asp Tyr Gly Asp Glu Tyr Arg Pro Leu Phe Phe Tyr Gln Glu Thr Thr Ala Gln Ile Leu Val Arg Ala Leu Asn Pro Leu Asp Tyr Met Lys Trp Arg Arg Lys Ser A1a Tyr Trp Lys Ala Leu Lys Val Phe Lys Leu Pro Val Glu Phe Leu Leu Leu Leu Thr Val Pro Val Val Asp Pro Asp Lys Asp Asp Gln Asn Trp Lys Arg Pro Leu Asn Cys Leu His Leu Va1 I1e Ser Pro Leu Val Val Val Leu Thr Leu Gln Ser Gly Thr Tyr Gly Val Tyr Glu Ile Gly Gly Leu Val Pro Val Trp Val Val Val Val Tle Ala Gly Thr Ala Leu Ala Ser Va1 Thr Phe Phe A1a Thr Ser Asp Ser Gln Pro Pro Arg Leu His Trp Leu Phe Ala Phe Leu Gly Phe Leu Thr Ser Ala Leu Trp Ile Asn Ala Ala Ala Thr Glu Val Val Asn Ile Leu Arg Ser Leu Gly Val Val Phe Arg Leu Ser Asn Thr Val Leu Gly Leu Thr Leu Leu A1a Trp G1y Asn Ser Ile Gly Asp Ala Phe Ser Asp Phe Thr Leu Ala Arg Gln Gly Tyr Pro Arg Met Ala Phe Ser Ala Cys Phe Gly Gly Ile Ile Phe Asn Ile Leu Val Gly Val Gly Leu Gly Cys Leu Leu Gln Ile Ser Arg Ser His Thr Glu Val Lys Leu Glu Pro Asp Gly Leu Leu Val Trp Val Leu Ala Gly Ala Leu Gly Leu Ser Leu Val Phe Ser Leu Val Ser Val Pro Leu Gln Cys Phe Gln Leu Ser Arg Val Tyr Gly Phe Cys Leu Leu Leu Phe Tyr Leu Asn Phe Leu Val Val Ala Leu Leu Ile Glu Phe Gly Val Ile His Leu Lys Ser Met <210> 6 <211> 416 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 6618083CD1 <400> 6 Met Lys Leu Ser Lys Lys Asp Arg Gly Glu Asp G1u Glu Ser Asp Ser Ala Lys Lys Lys Leu Asp Trp Ser Cys Ser Leu Leu Val Ala Ser Leu Ala Gly Ala Phe Gly Ser Ser Phe Leu Tyr Gly Tyr Asn Leu Ser Val Val Asn Ala Pro Thr Pro Tyr Ile Lys Ala Phe Tyr Asn Glu Ser Trp Glu Arg Arg His Gly Arg Pro Ile Asp Pro Asp Thr Leu Thr Leu Leu Trp Ser Val Thr Val Ser Ile Phe Ala Ile Gly Gly Leu Val Gly Thr Leu Ile Val Lys Met Ile Gly Lys Val Leu Gly Arg Lys His Thr Leu Leu Ala Asn Asn Gly Phe Ala Tle Ser Ala Ala Leu Leu Met A1a Cys Ser Leu Gln Ala Gly Ala Phe Glu Met Leu Ile Val Gly Arg Phe Ile Met Gly Ile Asp G1y Gly Val Ala Leu Ser Val Leu Pro Met Tyr Leu Ser Glu Ile Ser Pro Lys Glu Ile Arg G1y Ser Leu Gly Gln Val Thr Ala Ile Phe Ile Cys Ile Gly Va1 Phe Thr Gly Gln Leu Leu Gly Leu Pro Glu Leu Leu Gly Lys Glu Ser Thr Trp Pro Tyr Leu Phe Gly Val Ile Val Val Pro Ala Val Val Gln Leu Leu Ser Leu Pro Phe Leu Pro Asp Ser Pro Arg Tyr Leu Leu Leu Glu Lys His Asn Glu Ala Arg Ala Val Lys Ala Phe Gln Thr Phe Leu Gly Lys Ala Asp Val Ser Gln Glu Val Glu Glu Val Leu Ala Glu Ser His Val Gln Arg Ser Ile Arg Leu Val Ser Val Leu Glu Leu Leu Arg Ala Pro Tyr Val Arg Trp Gln Val Val Thr Val Ile Val Thr Met Ala Cys Tyr Gln Leu Cys Gly Leu Asn Ala Ile Trp Phe Tyr Thr Asn Ser Ile Phe Gly Lys Ala Gly Ile Pro Leu Ala Lys Ile Pro Tyr Val Thr Leu Ser Thr Gly Gly Ile Glu Thr Leu Ala Ala Val Phe Ser Gly Leu Val Ile Glu His Leu Gly Arg Arg Pro Leu Leu Ile Gly Gly Phe Gly Leu Met Gly Leu Phe Phe Gly Thr Leu Thr Ile Thr Leu Thr Leu Gln Asp His Ala Pro Trp Val Pro Tyr Leu Ser Ile Val Gly Ile Leu Ala Ile Ile Ala Ser Phe Cys Ser Gly Pro Ala Val Phe Pro Glu Glu Thr Val Asn Val Ser Ile Val Ser Glu <210> 7 <211> 664 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 7472002CD1 <400> 7 Met Thr Glu Lys Thr Asn Gly Val Lys Ser Ser Pro Ala Asn Asn His Asn His His Ala Pro Pro Ala Ile Lys Ala Asn Gly Lys Asp Asp His Arg Thr Ser Ser Arg Pro His Ser Ala Ala Asp Asp Asp Thr Ser Ser Glu Leu Gln Arg Leu Ala Asp Val Asp Ala Pro Gln Gln Gly Arg Ser Gly Phe Arg Arg Ile Val Arg Leu Val Gly Ile Ile Arg Glu Trp Ala Asn Lys Asn Phe Arg Glu Glu Glu Pro Arg Pro Asp Ser Phe Leu Glu Arg Phe Arg Gly Pro Glu Leu Gln Thr Val Thr Thr Gln Glu Gly Asp Gly Lys Gly Asp Lys Asp Gly Glu Asp Lys Gly Thr Lys Lys Lys Phe Glu Leu Phe Val Leu Asp Pro Ala Gly Asp Trp Tyr Tyr Cys Trp Leu Phe Val Ile Ala Met Pro Val Leu Tyr Asn Trp Cys Leu Leu Val Ala Arg Ala Cys Phe Ser Asp Leu Gln Lys Gly Tyr Tyr Leu Val Trp Leu Val Leu Asp Tyr Val Ser Asp Va1 Va1 Tyr Ile Ala Asp Leu Phe Ile Arg Leu Arg Thr Gly Phe Leu Glu G1n Gly Leu Leu Val Lys Asp Thr Lys Lys Leu Arg Asp Asn Tyr Ile His Thr Leu Gln Phe Lys Leu Asp Val Ala Ser Ile Ile Pro Thr Asp Leu Ile Tyr Phe Ala Val Asp Ile His Ser Pro Glu Val Arg Phe Asn Arg Leu Leu His Phe Ala Arg Met Phe Glu Phe Phe Asp Arg Thr Glu Thr Arg Thr Asn Tyr Pro Asn Ile Phe Arg Ile Ser Asn Leu Val Leu Tyr Ile Leu Val Ile Ile His Trp Asn Ala Cys Ile Tyr Tyr Ala Ile Ser Lys Ser Ile Gly Phe Gly Val Asp Thr Trp Val Tyr Pro Asn Ile Thr Asp Pro Glu Tyr Gly Tyr Leu Ala Arg Glu Tyr Ile Tyr Cys Leu Tyr Trp Ser Thr Leu Thr Leu Thr Thr Ile Gly Glu Thr Pro Pro Pro Val Lys Asp Glu Glu Tyr Leu Phe Val Ile Phe Asp Phe Leu Ile Gly Val Leu Ile Phe Ala Thr Ile Val Gly Asn Val Gly Ser Met Ile Ser Asn Met Asn Ala Thr Arg Ala Glu Phe Gln Ala Lys Ile Asp Ala Val Lys His Tyr Met Gln Phe Arg Lys Val Ser Lys Gly Met Glu Ala Lys Val Ile Arg Trp Phe Asp Tyr Leu Trp Thr Asn Lys Lys Thr Val Asp Glu Arg Glu Ile Leu Lys Asn Leu Pro Ala Lys Leu Arg Ala Glu Ile Ala Tle Asn Val His Leu Ser Thr Leu Lys Lys Val Arg Ile Phe His Asp Cys Glu Ala Gly Leu Leu Val G1u Leu Val Leu Lys Leu Arg Pro Gln Val Phe Ser Pro Gly Asp Tyr Ile Cys Arg Lys Gly Asp Ile Gly Lys Glu Met Tyr Ile Ile Lys Glu G1y Lys Leu Ala Val Val Ala Asp Asp Gly Val Thr Gln Tyr Ala Leu Leu Ser Ala Gly Ser Cys Phe G1y Glu Ile Ser Ile Leu Asn Ile Lys Gly Ser Lys Met Gly Asn Arg Arg Thr Ala Asn Ile Arg Ser Leu Gly Tyr Ser Asp Leu Phe Cys Leu Ser Lys Asp Asp Leu Met Glu Ala Val Thr Glu Tyr Pro Asp Ala Lys Lys Val Leu Glu Glu Arg Gly Arg Glu Tle Leu Met Lys Glu Gly Leu Leu Asp Glu Asn Glu Val Ala Thr Ser Met Glu Val Asp Val Gln Glu Lys Leu Gly Gln Leu Glu Thr Asn Met Glu Thr Leu Tyr Thr Arg Phe Gly Arg Leu Leu Ala Glu Tyr Thr Gly Ala Gln Gln Lys Leu Lys Gln Arg Ile Thr Val Leu Glu Thr Lys Met Lys Gln Asn Asn Glu Asp Asp Tyr Leu Ser Asp Gly Met Asn Ser Pro Glu Leu Ala A1a Ala Asp Glu Pro <210> 8 <211> 242 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1812692CD1 <400> 8 Met Ser Phe Arg Ala Ala Arg Leu Ser Met Arg Asn Arg Arg Asn Asp Thr Leu Asp Ser Thr Arg Thr Leu Tyr Ser Ser Ala Ser Arg Ser Thr Asp Leu Ser Tyr Ser Glu Ser Asp Leu Val Asn Phe Ile Gln Ala Asn Phe Lys Lys Arg Glu Cys Val Phe Phe Thr Lys Asp Ser Lys Ala Thr Glu Asn Val Cys Lys Cys Gly Tyr Ala Gln Ser Gln His Met Glu Gly Thr Gln Ile Asn Gln Ser Glu Lys Trp Asn Tyr Lys Lys His Thr Lys Glu Phe Pro Thr Asp Ala Phe Gly Asp Ile Gln Phe Glu Thr Leu Gly Lys Lys Gly Lys Tyr Ile Arg Leu Ser Cys Asp Thr Asp Ala Glu Ile Leu Tyr Glu Leu Leu Thr Gln His Trp His Leu Lys Thr Pro Asn Leu Val Ile Ser Val Thr Gly Gly Ala Lys Asn Phe Ala Leu Lys Pro Arg Met Arg Lys Ile Phe Ser Arg Leu Ile Tyr Ile Ala Gln Ser Lys Gly Ala Trp Ile Leu Thr Gly G1y Thr His Tyr Gly Leu Met Lys Tyr Ile Gly Glu Val Val Arg Asp Asn Thr Ile Ser Arg Ser Ser Glu Glu Asn Ile Val Ala Ile Gly Ile Ala Ala Trp Gly Met Val Ser Asn Arg Asp Thr Leu Ile Arg Asn Cys Asp Ala Glu Val Pro Val Gly Gln Glu Glu Val Cys <210> 9 <211> 398 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 3232992CD1 <400> 9 Met Val Ala Ala Pro Ile Phe Gly Tyr Leu Gly Asp Arg Phe Asn Arg Lys Val Ile Leu Ser Cys Gly Ile Phe Phe Trp Ser Ala Val Thr Phe Ser Ser Ser Phe Ile Pro Gln Gln Tyr Phe Trp Leu Leu Val Leu Ser Arg Gly Leu Val Gly Ile Gly Glu Ala Ser Tyr Ser Thr Ile Ala Pro Thr Tle Ile Gly Asp Leu Phe Thr Lys Asn Thr Arg Thr Leu Met Leu Ser Val Phe Tyr Phe Ala Ile Pro Leu Gly Ser Gly Leu Gly Tyr Ile Thr Gly Ser Ser Val Lys Gln Ala Ala Gly Asp Trp His Trp Ala Leu Arg Val Ser Pro Val Leu Gly Met Ile Thr Gly Thr Leu Ile Leu Ile Leu Val Pro Ala Thr Lys Arg Gly His Ala Asp Gln Leu Gly Asp Gln Leu Lys Ala Arg Thr Ser Trp Leu Arg. Asp Met Lys Ala Leu Ile Arg Asn Arg Ser Tyr Val Phe Ser Ser Leu A1a Thr Ser Ala Val Ser Phe Ala Thr Gly Ala Leu Gly Met Trp Ile Pro Leu Tyr Leu His Arg Ala Gln Val Val Gln Lys Thr Ala Glu Thr Cys Asn Ser Pro Pro Cys Gly Ala Lys Asp Ser Leu 21e Phe Gly Ala Ile Thr Cys Phe Thr G1y Phe Leu Gly Val Val Thr Gly Ala Gly Ala Thr Arg Trp Cys Arg Leu Lys Thr Gln Arg Ala Asp Pro Leu Val Cys Ala Val Gly Met Leu Gly Ser Ala Ile Phe Ile Cys Leu Ile Phe Val Ala Ala Lys Ser Ser Ile Val Gly Ala Tyr Ile Cys Ile Phe Val Gly Glu Thr Leu Leu Phe Ser Asn Trp Ala Ile Thr Ala Asp Ile Leu Met Tyr Val Val Ile Pro Thr Arg Arg Ala Thr Ala Val Ala Leu Gln Ser Phe Thr Ser His Leu Leu Gly Asp Ala Gly Ser Pro Tyr Leu Ile Gly Phe Ile Ser Asp Leu Ile Arg Gln Ser Thr Lys Asp Ser Pro Leu Trp Glu Phe Leu Ser Leu Gly Tyr Ala Leu Met Leu Cys Pro Phe Val Val Val Leu Gly Gly Met Phe Phe Leu Ala Thr Ala Leu Phe Phe Val Ser Asp Arg Ala Arg Ala Glu Gln Gln Val Asn Gln Leu Ala Met Pro Pro Ala Ser Val Lys Val <210> 10 <211> 553 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 3358383CD1 <400> 10 Met Ala Phe Gln Asp Leu Leu Gly His Ala Gly Asp Leu Trp Arg Phe Gln Ile Leu Gln Thr Val Phe Leu Ser Ile Phe Ala Val Ala Thr Tyr Leu His Phe Met Leu Glu Asn Phe Thr Ala Phe Ile Pro Gly His Arg Cys Trp Val His Ile Leu Asp Asn Asp Thr Val Ser Asp Asn Asp Thr Gly Ala Leu Ser Gln Asp Ala Leu Leu Arg Ile Ser Ile Pro Leu Asp Ser Asn Met Arg Pro Glu Lys Cys Arg Arg Phe Val His Pro Gln Trp Gln Leu Leu His Leu Asn Gly Thr Phe Pro Asn Thr Ser Asp Ala Asp Met Glu Pro Cys Val Asp Gly Trp Val Tyr Asp Arg Ile Ser Phe Ser Ser Thr Ile Val Thr Glu Trp Asp Leu Val Cys Asp Ser Gln Ser Leu Thr Ser Val Ala Lys Phe Val Phe Met Ala Gly Met Met Val Gly Gly Ile Leu Gly Gly His Leu Ser Asp Arg Phe Gly Arg Arg Phe Val Leu Arg Trp Cys Tyr Leu Gln Val Ala Ile Val Gly Thr Cys Ala Ala Leu A1a Pro Thr Phe Leu Ile Tyr Cys Ser Leu Arg Phe Leu Ser Gly Ile Ala Ala Met Ser Leu I1e Thr Asn Thr Ile Met Leu Ile Ala Glu Trp Ala Thr His Arg Phe Gln Ala Met Gly Ile Thr Leu Gly Met Cys Pro Ser Gly Ile Ala Phe Met Thr Leu Ala Gly Leu Ala Phe Ala Ile Arg Asp Trp His Ile Leu Gln Leu Val Val Ser Val Pro Tyr Phe Val Ile Phe Leu Thr Ser Ser Trp Leu Leu Glu Ser Ala Arg Trp Leu Ile Ile Asn Asn Lys Pro Glu Glu Gly Leu Lys Glu Leu Arg Lys Ala Ala His Arg Ser Gly Met Lys Asn Ala Arg Asp Thr Leu Thr Leu Glu Ile Leu Lys Ser Thr Met Lys Lys Glu Leu Glu Ala Ala Gln Lys Lys Lys Pro Ser Leu Cys Glu Met Leu His Met Pro Asn Ile Cys Lys Arg Ile Ser Leu Leu Ser Phe Thr Arg Phe Ala Asn Phe Met Ala Tyr Phe Gly Leu Asn Leu His Val Gln His Leu Gly Asn Asn Val Phe Leu Leu Gln Thr Leu Phe G1y Ala Va1 Ile Leu Leu A1a Asn Cys Val Ala Pro Trp Ala Leu Lys Tyr Met Thr Arg Arg Ala Ser Gln Met Arg Leu Met Tyr Leu Leu Ala Ile Cys Phe Met Ala Ile Ile Phe Val Pro Gln Glu Met Gln Thr Leu Arg Glu Val Leu Ala Thr Leu Gly Leu Gly Ala Ser Ala Leu Thr Asn Thr Leu Ala Phe Ala His Gly Asn Glu Val Ile Pro Thr Ile Ile Arg Ala Arg Ala Met Gly Ile Asn Ala Thr Phe Ala Asn Ile Ala Gly Ala Leu Ala Pro Leu Met Met Ile Leu Ser Val Tyr Ser Pro Pro Leu Pro Trp Ile Ile Tyr Gly Val Phe Pro Phe Ile Ser Gly Phe Ala Phe Leu Leu Leu Pro Glu Thr Arg Asn Lys Pro Leu Phe Asp Thr Ile Gln Asp Glu Lys Asn Glu Arg Lys Asp Pro Arg Glu Pro Lys Gln Glu Asp Pro Arg Val Glu Val Thr Gln Phe <210> 11 <211> 213 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 4250091CD1 <400> 11 Met Ser Ser Gln Glu Leu Val Thr Leu Asn Val Gly Gly Lys Ile Phe Thr Thr Arg Phe Ser Thr Ile Lys Gln Phe Pro Ala Ser Arg Leu Ala Arg Met Leu Asp Gly Arg Asp Gln Glu Phe Lys Met Val Gly Gly Gln Ile Phe Val Asp Arg Asp Gly Asp Leu Phe Ser Phe Ile Leu Asp Phe Leu Arg Thr His Gln Leu Leu Leu Pro Thr Glu Phe Ser Asp Tyr Leu Arg Leu Gln Arg Glu Ala Leu Phe Tyr Glu Leu Arg Ser Leu Val Asp Leu Leu Asn Pro Tyr Leu Leu Gln Pro Arg Pro Ala Leu Val Glu Val His Phe Leu Ser Arg Asn Thr Gln Ala Phe Phe Arg Val Phe Gly Ser Cys Ser Lys Thr Ile Glu Met Leu Thr Gly Arg Ile Thr Val Phe Thr Glu Gln Pro Ser A1a Pro Thr Trp Asn Gly Asn Phe Phe Pro Pro Gln Met Thr Leu Leu Pro Leu Pro Pro Gln Arg Pro Ser Tyr His Asp Leu Val Phe G1n Cys Gly Ser Asp Ser Thr Thr Asp Asn Gln Thr Gly Val Arg Tyr Phe Val Leu Cys Ser Ile Ser Leu Val Tyr Gln Phe Val Met Phe Ser Leu Lys Thr <210> 12 <211> 476 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 70064803CD1 <400> 12 Met Ala Gly Ser Asp Thr Ala Pro Phe Leu Ser Gln Ala Asp Asp Pro Asp Asp Gly Pro Val Pro Gly Thr Pro Gly Leu Pro Gly Ser Thr Gly Asn Pro Lys Ser Glu Glu Pro G1u Val Pro Asp Gln Glu Gly Leu Gln Arg Ile Thr Gly Leu Ser Pro G1y Arg Ser Ala Leu Ile Val Ala Val Leu Cys Tyr Ile Asn Leu Leu Asn Tyr Met Asp Arg Phe Thr Val Ala Gly Val Leu Pro Asp Ile Glu Gln Phe Phe Asn Ile Gly Asp Ser Ser Ser Gly Leu Ile Gln Thr Val Phe Ile Ser Ser Tyr Met Val Leu Ala Pro Val Phe Gly Tyr Leu Gly Asp Arg Tyr Asn Arg Lys Tyr Leu Met Cys Gly Gly I1e Ala Phe Trp Ser Leu Val Thr Leu Gly Ser Ser Phe Ile Pro Gly Glu His Phe Trp Leu Leu Leu Leu Thr Arg Gly Leu Val Gly Val Gly Glu Ala Ser Tyr Ser Thr Ile A1a Pro Thr Leu Ile Ala Asp Leu Phe Val Ala Asp Gln Arg Ser Arg Met Leu Ser Ile Phe Tyr Phe Ala Ile Pro Val Gly Ser Gly Leu Gly Tyr Ile Ala Gly Ser Lys Val Lys Asp Met Ala Gly Asp Trp His Trp Ala Leu Arg Val Thr Pro Gly Leu Gly Val Val A1a Val Leu Leu Leu Phe Leu Va1 Val Arg G1u Pro Pro Arg Gly Ala Val Glu Arg His Ser Asp Leu Pro Pro Leu Asn Pro Thr Ser Trp Trp Ala Asp Leu Arg Ala Leu Ala Arg Asn Leu Ile Phe Gly Leu Ile Thr Cys Leu Thr Gly Val Leu Gly Val Gly Leu Gly Val Glu Ile Ser Arg Arg Leu Arg His Ser Asn Pro Arg Ala Asp Pro Leu Val Cys Ala Thr Gly Leu Leu Gly Ser Ala Pro Phe Leu Phe Leu Ser Leu A1a Cys Ala Arg Gly Ser Ile Val Ala Thr Tyr Ile Phe Ile Phe Ile Gly Glu Thr Leu Leu Ser Met Asn Trp Ala Ile Val Ala Asp Ile Leu Leu Tyr Val Val Ile Pro Thr Arg Arg Ser Thr Ala Glu Ala Phe Gln Ile Val Leu Ser His Leu Leu Gly Asp Ala Gly Ser Pro Tyr Leu Ile Gly Leu Ile Ser Asp Arg Leu Arg Arg Asn Trp Pro Pro Ser Phe Leu Ser Glu Phe Arg Ala Leu Gln Phe Ser Leu Met Leu Cys Ala Phe Val Gly Ala Leu Gly Gly Ala Ala Phe Leu Gly Thr Ala Ile Phe Ile Glu Ala Asp Arg Arg Arg A1a Gln Leu His Val Gln Gly Leu Leu His Glu A1a Gly Ser Thr Asp Asp Arg Ile Val Val Pro Gln Arg Gly Arg Ser Thr Arg Val Pro Val Ala Ser Val Leu Ile <210> 13 <211> 246 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 70356768CD1 <400> 13 Met Leu His Ala Leu Leu Arg Ser Arg Met Ile Gln Gly Arg Ile Leu Leu Leu Thr Ile Cys Ala Ala Gly Ile Gly Gly Thr Phe Gln Phe Gly Tyr Asn Leu Ser Ile Ile Asn Ala Pro Thr Leu His Ile Gln Glu Phe Thr Asn Glu Thr Trp G1n Ala Arg Thr Gly Glu Pro Leu Pro Asp His Leu Val Leu Leu Met Trp Ser Leu I1e Val Ser Leu Tyr Pro Leu Gly Gly Leu Phe Gly Ala Leu Leu Ala Gly Pro Leu Ala Ile Thr Leu Gly Arg Lys Lys Ser Leu Leu Val Asn Asn Ile Phe Val Val Ser Ala Ala Ile Leu Phe Gly Phe Ser Arg Lys Ala Gly Ser Phe Glu Met Ile Met Leu Gly Arg Leu Leu Val Gly Val Asn Ala Gly Val Ser Met Asn Ile Gln Pro Met Tyr Leu Gly Glu Ser Ala Pro Lys Glu Leu Arg Gly Ala Val Ala Met Ser Ser Ala Ile Phe Thr Ala Leu Gly Ile Val Met Gly Gln Val Val Gly Leu Arg Glu Leu Leu Gly Gly Pro Gln Ala Trp Pro Leu Leu Leu Ala Ser Cys Leu Va1 Pro Gly Ala Leu Gln Leu Ala Ser Leu Pro Leu Leu Pro Glu Ser Pro Arg Tyr Leu Leu Ile Asp Cys Gly Asp Thr Glu Ala Cys Leu Ala Glu Thr Gly Ser Arg Leu Ser Arg Leu Glu Cys Cys Gly Cys Ser <210> 14 <211> 436 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 5674114CD1 <400> 14 Met Gly Leu Ala Arg Ala Leu Arg Arg Leu Ser Gly Ala Leu Asp Ser Gly Asp Ser Arg A1a Gly Asp Glu Glu Glu Ala Gly Pro Gly Leu Cys Arg Asn Gly Trp Ala Pro Ala Pro Val Gln Ser Pro Val 35 40 ~ 45 Gly Arg Arg Arg Gly Arg Phe Val Lys Lys Asp Gly His Cys Asn Val Arg Phe Val Asn Leu Gly Gly Gln Gly Ala Arg Tyr Leu Ser Asp Leu Phe Thr Thr Cys Val Asp Val Arg Trp Arg Trp Met Cys Leu Leu Phe Ser Cys Ser Phe Leu Ala Ser Trp Leu Leu Phe Gly Leu Ala Phe Trp Leu Ile Ala Ser Leu His Gly Asp Leu Ala Ala Pro Pro Pro Pro Ala Pro Cys Phe Ser His Va1 Ala Ser Phe Leu Ala Ala Phe Leu Phe Ala Leu Glu Thr Gln Thr Ser Ile Gly Tyr Gly Val Arg Ser Val Thr Glu Glu Cys Pro Ala Ala Val Ala Ala Val Val Leu Gln Cys Ile Ala Gly Cys Val Leu Asp Ala Phe Val Val Gly Ala Val Met Ala Lys Met Ala Lys Pro Lys Lys Arg Asn Glu Thr Leu Val Phe Ser Glu Asn Ala Val Val Ala Leu Arg Asp His Arg Leu Cys Leu Met Trp Arg Val Gly Asn Leu Arg Arg Ser His Leu Val Glu Ala His Val Arg Ala Gln Leu Leu Gln Pro Arg Val Thr Pro Glu Gly Glu Tyr Ile Pro Leu Asp His Gln Asp Val Asp Val Gly Phe Asp Gly Gly Thr Asp Arg Ile Phe Leu Val Ser Pro Ile Thr Ile Val His Glu Ile Asp Ser Ala Ser Pro Leu Tyr Glu Leu Gly Arg Ala Glu Leu Ala Arg Ala Asp Phe Glu Leu Val Val Ile Leu G1u Gly Met Val Glu Ala Thr Ala Met Thr Thr Gln Cys Arg Ser Ser Tyr Leu Pro Gly Glu Leu Leu Trp Gly His Arg Phe Glu Pro Va1 Leu Phe Gln Arg Gly Ser Gln Tyr Glu Val Asp Tyr Arg His Phe His Arg Thr Tyr Glu Val Pro Gly Thr Pro Val Cys Ser Ala Lys Glu Leu Asp Glu Arg Ala Glu Gln Ala Ser His Ser Leu Lys Ser Ser Phe Pro Gly Ser Leu Thr Ala Phe Cys Tyr Glu Asn Glu Leu Ala Leu Ser Cys Cys Gln Glu Glu Asp Glu Asp Asp Glu Thr Glu Glu Gly Asn Gly Val Glu Thr Glu Asp Gly Ala Ala Ser Pro Arg Val Leu Thr Pro Thr Leu Ala Leu Thr Leu Pro Pro <210> 15 <211> 453 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 1254635CD1 <400> 15 Met Leu Lys Met Val Leu Thr Glu Asn Pro Asn Gln Glu Ile Ala Thr Ser Leu Glu Phe Leu Leu Leu Gln Asn Ser Pro Gly Ser Leu Arg Ala Gln Gln Arg Met Ser Tyr Tyr Gly Ser Ser Tyr His Ile Ile Asn Ala Asp Ala Lys Tyr Pro Gly Tyr Pro Pro Glu His Ile Ile Ala Glu Lys Arg Arg Ala Arg Arg Arg Leu Leu His Lys Asp Gly Ser Cys Asn Val Tyr Phe Lys His Ile Phe Gly Glu Trp Gly Ser Tyr Val Val Asp Ile Phe Thr~Thr Leu Val Asp Thr Lys Trp Arg His Met Phe Val Ile Phe Ser Leu Ser Tyr Ile Leu Ser Trp Leu Ile Phe Gly Ser Val Phe Trp Leu Ile Ala Phe His His Gly Asp Leu Leu Asn Asp Pro Asp Ile Thr Pro Cys Val Asp Asn Val His Ser Phe Thr Gly Ala Phe Leu Phe Ser Leu Glu Thr Gln Thr Thr Ile Gly Tyr Gly Tyr Arg Cys Val Thr Glu Glu Cys Ser Val Ala Val Leu Met Val Ile Leu Gln Ser Ile Leu Ser Cys Ile Ile Asn Thr Phe Ile Ile Gly A1a Ala Leu Ala Lys Met Ala Thr Ala Arg Lys Arg Ala Gln Thr Ile Arg Phe Ser Tyr Phe Ala Leu Ile Gly Met Arg Asp Gly Lys Leu Cys Leu Met Trp Arg Ile Gly Asp Phe Arg Pro Asn His Val Val Glu Gly Thr Val Arg Ala Gln Leu Leu Arg Tyr Thr Glu Asp Ser Glu Gly Arg Met Thr Met Ala Phe Lys Asp Leu Lys Leu Val Asn Asp Gln Ile Ile Leu Val Thr Pro Val Thr Ile Val His Glu Ile Asp His Glu Ser Pro Leu Tyr Ala Leu Asp Arg Lys Ala Val Ala Lys Asp Asn Phe Glu Ile Leu Val Thr Phe Ile Tyr Thr Gly Asp Ser Thr Gly Thr Ser His Gln Ser Arg Ser Ser Tyr Val Pro Arg Glu Ile Leu Trp Gly His Arg Phe Asn Asp Val Leu Glu Val Lys Arg Lys Tyr Tyr Lys Val Asn Cys Leu Gln Phe Glu Gly Ser Val Glu Val Tyr Ala Pro Phe Cys Ser Ala Lys G1n Leu Asp Trp Lys Asp Gln G1n Leu His Ile Glu Lys Ala Pro Pro Val Arg Glu Ser Cys Thr Ser Asp Thr Lys Ala Arg Arg Arg Ser Phe Ser Ala Val Ala Ile Val Ser Ser Cys Glu Asn Pro Glu Glu Thr Thr Thr Ser Ala Thr His Glu Tyr Arg Glu Thr Pro Tyr Gln Lys Ala Leu Leu Thr Leu Asn Arg Ile Ser Val Glu Ser Gln Met <210> 16 <211> 299 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1670595CD1 <400> 16 Met Ala Ser Glu Ser Ser Pro Leu Leu Ala Tyr Arg Leu Leu Gly Glu Glu Gly Val Ala Leu Pro Ala Asn Gly Ala Gly Gly Pro Gly Gly Ala Ser Ala Arg Lys Leu Ser Thr Phe Leu Gly Val Val Val Pro Thr Val Leu Ser Met Phe Ser Ile Val Val Phe Leu Arg Ile Gly Phe Val Val Gly His Ala Gly Leu Leu Gln Ala Leu Ala Met 65 70 . 75 Leu Leu Val Ala Tyr Phe Ile Leu Ala Leu Thr Val Leu Ser Val Cys Ala Ile Ala Thr Asn Gly Ala Val Gln Gly Gly Gly Ala Tyr Cys Ile Leu Gln His Arg Trp Thr G1y Met Pro Gln Gly Pro Val Gly Ser Gly Ser Cys Pro Arg Ala Thr Ala Trp Asn Leu Leu Tyr Gly Ser Leu Leu Leu Gly Leu Val Gly Gly Val Cys Thr Leu G1y Ala Gly Leu Tyr Ala Arg Ala Ser Phe Leu Thr Phe Leu Leu Val Ser Gly Ser Leu Ala Ser Val Leu Ile Ser Phe Val Ala Val Gly Pro Arg Asp Ile Arg Leu Thr Pro Arg Pro Gly Pro Asn Gly Ser Ser Leu Pro Pro Arg Phe Gly His Phe Thr Gly Phe Asn Ser Ser Thr Leu Lys Asp Asn Leu Gly Ala Gly Tyr Ala Glu Asp Tyr Thr Thr Gly A1a Val Met Asn Phe Ala Ser Val Phe Ala Val Leu Phe Asn Gly Arg His His Gly Trp Gly Gln His Val Arg Gly Ala Glu Gly Pro Gln Pro Gly Asp Pro Ser Gly His Asp Arg Arg Arg Arg Leu His Leu Leu Arg Leu Cys Pro Ala Phe Leu Ser Leu Gln Pro Pro Phe Thr Gly A1a Leu Met Leu G1y Ala Arg Pro Pro Leu <210> 17 <211> 606 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1859560CD1 <400> 17 Met Pro Ser Ser Val Thr Ala Leu Gly Gln Ala Arg Ser Ser Gly Pro Gly Met Ala Pro Ser Ala Cys Cys Cys Ser Pro Ala Ala Leu Gln Arg Arg Leu Pro Ile Leu Ala Trp Leu Pro Ser Tyr Ser Leu Gln Trp Leu Lys Met Asp Phe Val Ala Gly Leu Ser Val Gly Leu Thr Ala Ile Pro Gln Ala Leu Ala Tyr Ala Glu Val Ala Gly Leu Pro Pro Gln Tyr Gly Leu Tyr Ser Ala Phe Met Gly Cys Phe Val Tyr Phe Phe Leu G1y Thr Ser Arg Asp Val Thr Leu Gly Pro Thr Ala Ile Met Ser Leu Leu Val Ser Phe Tyr Thr Phe His Glu Pro Ala Tyr Ala Val Leu Leu Ala Phe Leu Ser Gly Cys Ile Gln Leu Ala Met Gly Val Leu Arg Leu Gly Phe Leu Leu Asp Phe Ile Ser Tyr Pro Val Ile Lys Gly Phe Thr Ser Ala Ala Ala Val Thr Ile Gly Phe Gly Gln Ile Lys Asn Leu Leu Gly Leu Gln Asn Ile Pro Arg Pro Phe Phe Leu G1n Val Tyr His Thr Phe Leu Arg Ile Ala Glu Thr Arg Val Gly Asp Ala Val Leu G1y Leu Val Cys Met Leu Leu Leu Leu Val Leu Lys Leu Met Arg Asp His Val Pro Pro Val His Pro Glu Met Pro Pro Gly Va1 Arg Leu Ser Arg Gly Leu Val Trp Ala Ala Thr Thr Ala Arg Asn Ala Leu Val Val Ser Phe Ala Ala Leu Val Ala Tyr Ser Phe Glu Val Thr Gly Tyr Gln Pro Phe Ile Leu Thr Gly Glu Thr Ala Glu Gly Leu Pro Pro Val Arg Ile Pro Pro Phe Ser Val Thr Thr Ala Asn Gly Thr Ile Ser Phe Thr Glu Met Val Gln Asp Met Gly Ala Gly Leu Ala Val Val Pro Leu Met Gly Leu Leu Glu Ser Ile Ala Val Ala Lys Ala Phe Ala Ser Gln Asn Asn Tyr Arg Ile Asp Ala Asn Gln Glu Leu Leu Ala Ile Gly Leu Thr Asn Met Leu Gly Ser Leu Val Ser Ser Tyr Pro Val Thr G1y Ser Phe G1y Arg Thr Ala Va1 Asn Ala Gln Ser Gly Val Cys Thr Pro Ala Gly Gly Leu Val Thr Gly Val Leu Val Leu Leu Ser Leu Asp Tyr Leu Thr Ser Leu Phe Tyr Tyr Ile Pro Lys Ser Ala Leu Ala Ala Val Ile Ile Met Ala Val Ala Pro Leu Phe Asp 410 4l5 420 Thr Lys Ile Phe Arg Thr Leu Trp Arg Val Lys Arg Leu Asp Leu Leu Pro Leu Cys Val Thr Phe Leu Leu Cys Phe Trp Glu Val Gln Tyr Gly Ile Leu A1a Gly Ala Leu Val Ser Leu Leu Met Leu Leu His Ser Ala Ala Arg Pro Glu Thr Lys Val Ser Glu Gly Pro Val Leu Val Leu Gln Pro Ala Ser Gly Leu Ser Phe Pro Ala Met Glu Ala Leu Arg Glu Glu Ile Leu Ser Arg Ala Leu Glu Val Ser Pro 500 505 5l0 Pro Arg Cys Leu Val Leu Glu Cys Thr His Val Cys Ser Ile Asp Tyr Thr Val Val Leu Gly Leu Gly Glu Leu Leu Gln Asp Phe Gln Lys Gln Gly Val Ala Leu Ala Phe Val Gly Leu Gln Val Pro Val Leu Arg Val Leu Leu Ser Ala Asp Leu Lys Gly Phe Gln Tyr Phe Ser Thr Leu Glu Glu Ala Glu Lys His Leu Arg G1n G1u Pro Gly Thr Gln Pro Tyr Asn Ile Arg Glu Asp Ser Ile Leu Asp Gln Lys Val Ala Leu Leu Lys Ala <2l0> 18 <211> 324 <2l2> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 5530164CD1 <400> 18 Met Ser Val Glu Asp Gly Gly Met Pro Gly Leu Gly Arg Pro Arg Gln Ala Arg Trp Thr Leu Met Leu Leu Leu Ser Thr Ala Met Tyr Gly Ala His Ala Pro Leu Leu Ala Leu Cys His Val Asp Gly Arg Val Pro Phe Arg Pro Ser Ser A1a Val Leu Leu Thr Glu Leu Thr Lys Leu Leu Leu Cys Ala Phe Ser Leu Leu Val Gly Trp Gln Ala Trp Pro Gln Gly Pro Pro Pro Trp Arg Gln Ala Ala Pro Phe Ala Leu Ser Ala Leu Leu Tyr Gly Ala Asn Asn Asn Leu Val Ile Tyr Leu Gln Arg Tyr Met Asp Pro Ser Thr Tyr Gln Val Leu Ser Asn 1l0 1l5 120 Leu Lys Ile Gly Ser Thr Ala Val Leu Tyr Cys Leu Cys Leu Arg His Arg Leu Ser Val Arg Gln Gly Leu Ala Leu Leu Leu Leu Met Ala Ala Gly Ala Cys Tyr Ala Ala Gly Gly Leu Gln Val Pro Gly Asn Thr Leu Pro Ser Pro Pro Pro Ala Ala Ala Ala Ser Pro Met Pro Leu His Ile Thr Pro Leu Gly Leu Leu Leu Leu Ile Leu Tyr Cys Leu Ile Ser Gly Leu Ser Ser Val Tyr Thr Glu Leu Leu Met Lys Arg Gln Arg Leu Pro Leu Ala Leu Gln Asn Leu Phe Leu Tyr Thr Phe Gly Val Leu Leu Asn Leu Gly Leu His Ala Gly Gly Gly Ser Gly Pro Gly Leu Leu Glu Gly Phe Ser Gly Trp Ala Ala Leu Val Val Leu Ser Gln Ala Leu Asn Gly Leu Leu Met Ser Ala Val Met Lys His Gly Ser Ser Ile Thr Arg Leu Phe Val Val Ser Cys Ser Leu Val Val Asn Ala Val Leu Ser Ala Val Leu Leu Arg Leu Gln Leu Thr Ala Ala Phe Phe Leu Ala Thr Leu Leu Ile G1y Leu A1a Met Arg Leu Tyr Tyr Gly Ser Arg <210> 19 <211> 445 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 139115CD1 <400> 19 Met Thr Leu Thr Gly Pro Leu Thr Thr Gln Tyr Val Tyr Arg Arg Ile Trp Glu Glu Thr G1y Asn Tyr Thr Phe Ser Ser Asp Ser Asn Ile Ser G1u Cys Glu Lys Asn Lys Ser Ser Pro Ile Phe Ala Phe Gln Glu Glu Val Gln Lys Lys Val Ser Arg Phe Asn Leu Gln Met Asp Ile Ser Gly Leu Ile Pro Gly Leu Val Ser Thr Phe Ile Leu Leu Ser Ile Ser Asp His Tyr Gly Arg Lys Phe Pro Met Ile Leu Ser Ser Val Gly A1a Leu Ala Thr Ser Val Trp Leu Cys Leu Leu Cys Tyr Phe Ala Phe Pro Phe Gln Leu Leu Ile Ala Ser Thr Phe Ile Gly Ala Phe Cys Gly Asn Tyr Thr Thr Phe Trp Gly A1a Cys Phe Ala Tyr Ile Val Asp Gln Cys Lys Glu His Lys Gln Lys Thr Ile Arg Ile Ala Ile Ile Asp Phe Leu Leu Gly Leu Val Thr Gly Leu Thr Gly Leu Ser Ser Gly Tyr Phe Ile Arg Glu Leu Gly Phe Glu Trp Ser Phe Leu Ile Ile Ala Val Ser Leu Ala Val Asn Leu Ile Tyr Tle Leu Phe Phe Leu Gly Asp Pro Val Lys Glu Cys Ser Ser Gln Asn Val Thr Met Ser Cys Ser Glu Gly Phe Lys Asn Leu Phe Tyr Arg Thr Tyr Met Leu Phe Lys Asn Ala Ser Gly Lys Arg 230 ~ 235 240 Arg Phe Leu Leu Cys Leu Leu Leu Phe Thr Val Ile Thr Tyr Phe Phe Val Val Ile Gly Ile Ala Pro Ile Phe Ile Leu Tyr Glu Leu Asp Ser Pro Leu Cys Trp Asn Glu Val Phe Tle Gly Tyr Gly Ser Ala Leu Gly Ser Ala Ser Phe Leu Thr Ser Phe Leu Gly Ile Trp Leu Phe Ser Tyr Cys Met Glu Asp Ile His Met Ala Phe Ile Gly Ile Phe Thr Thr Met Thr Gly Met Ala Met Thr Ala Phe Ala Ser Thr Thr Leu Met Met Phe Leu Ala Arg Val Pro Phe Leu Phe Thr Ile Val Pro Phe Ser Val Leu Arg Ser Met Leu Ser Lys Val Val Arg Ser Thr Glu Gln Gly Thr Leu Phe Ala Cys Ile Ala Phe Leu Glu Thr Leu Gly Gly Val Thr Ala Val Ser Thr Phe Asn Gly Ile Tyr Ser Ala Thr Val Ala Trp Tyr Pro Gly Phe Thr Phe Leu Leu Ser Ala Gly Leu Leu Leu Leu Pro Ala Ile Ser Leu Cys Val Val Lys Cys Thr Ser Trp Asn Glu Gly Ser Tyr Glu Leu Leu I1e Gln G1u Glu Ser Ser Glu Asp Ala Ser Asp Arg <210> 20 <211> 337 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1702940CD1 <400> 20 Met Asn Pro Glu Ser Ser Ile Phe Ile Glu Asp Tyr Leu Lys Tyr Phe Gln Asp Gln Val Ser Arg Glu Asn Leu Leu Gln Leu Leu Thr Asp Asp Glu Ala Trp Asn Gly Phe Val Ala Ala Ala Glu Leu Pro Arg Asp Glu Ala Asp G1u Leu Arg Lys Ala Leu Asn Lys Leu Ala Ser His Met Val Met Lys Asp Lys Asn Arg His Asp Lys Asp Gln G1n His Arg Gln Trp Phe Leu Lys Glu Phe Pro Arg Leu Lys Arg Glu Leu Glu Asp His Ile Arg Lys Leu Arg Ala Leu Ala Glu Glu Va1 Glu Gln Val His Arg Gly Thr Thr Ile Ala Asn Val Val Ser Asn Ser Val Gly Thr Thr Ser Gly Ile Leu Thr Leu Leu Gly Leu Gly Leu A1a Pro Phe Thr Glu Gly Ile Ser Phe Val Leu Leu Asp Thr Gly Met Gly Leu Gly Ala A1a Ala Ala Val Ala Gly Ile Thr Cys Ser Val Val Glu Leu Val Asn Lys Leu Arg Ala Arg Ala Gln Ala Arg Asn Leu Asp Gln Ser Gly Thr Asn Val A1a Lys Val Met Lys Glu Phe Val Gly Gly Asn Thr Pro Asn Val Leu Thr Leu Val Asp Asn Trp Tyr Gln Val Thr Gln Gly Ile Gly Arg Asn Ile Arg Ala I1e Arg Arg Ala Arg Ala Asn Pro Gln Leu Gly Ala Tyr Ala Pro Pro Pro His Val Ile Gly Arg Ile Ser Ala Glu Gly G1y Glu Gln Va1 Glu Arg Val Val Glu Gly Pro Ala Gln Ala Met Ser Arg Gly Thr Met Ile Val Gly Ala Ala Thr Gly Gly Ile Leu Leu Leu Leu Asp Val Va1 Ser Leu Ala Tyr Glu Ser Lys His Leu Leu Glu Gly Ala Lys Ser Glu Ser Ala Glu Glu Leu Lys Lys Arg Ala Gln Glu Leu Glu Gly Lys Leu Asn Phe Leu Thr Lys Ile His Glu Met Leu Gln Pro Gly Gln Asp Gln <210> 21 <211> 273 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1703342CD1 <400> 21 Met Ala Thr Trp Asp Glu Lys Ala Val Thr Arg Arg Ala Lys Val Ala Pro Ala Glu Arg Met Ser Lys Phe Leu Arg His Phe Thr Val Val Gly Asp Asp Tyr His Ala Trp Asn Ile Asn Tyr Lys Lys Trp Glu Asn Glu Glu Glu Glu G1u Glu Glu G1u Gln Pro Pro Pro Thr Pro Val Ser Gly Glu Glu Gly Arg Ala Ala Ala Pro Asp Val Ala Pro Ala Pro Gly Pro Ala Pro Arg Ala Pro Leu Asp Phe Arg Gly Met Leu Arg Lys Leu Phe Ser Ser His Arg Phe Gln Val Ile Ile Ile Cys Leu Val Val Leu Asp Ala Leu Leu Val Leu Ala Glu Leu Ile Leu Asp Leu Lys Ile Ile Gln Pro Asp Lys Asn Asn Tyr Ala Ala Met Val Phe His Tyr Met Ser Ile Thr Ile Leu Val Phe Phe Met Met Glu Ile Ile Phe Lys Leu Phe Val Phe Arg Leu Glu Phe Phe His His Lys Phe Glu Ile Leu Asp Ala Val Val Val Val Val Ser Phe Ile Leu Asp Ile Val Leu Leu Phe Gln Glu His Gln Phe Glu Ala Leu Gly Leu Leu Ile Leu Leu Arg Leu Trp Arg Val Ala Arg Ile Ile Asn Gly Tle Ile Ile Ser Va1 Lys Thr Arg Ser Glu Arg Gln Leu Leu Arg Leu Lys Gln Met Asn Val Gln Leu Ala Ala Lys Ile Gln His Leu Glu Phe Ser Cys Ser Glu Lys Glu Gln Glu Ile Glu Arg Leu Asn Lys Leu Leu Arg Gln His Gly Leu Leu Gly Glu Val Asn <210> 22 <211> 710 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1727529CD1 <400> 22 Met Gly Gly Lys Gln Arg Asp Glu Asp Asp Glu Ala Tyr Gly Lys Pro Val Lys Tyr Asp Pro Ser Phe Arg Gly Pro Ile Lys Asn Arg Ser Cys Thr Asp Val Ile Cys Cys Val Leu Phe Leu Leu Phe Ile Leu Gly Tyr Ile Val Val Gly Ile Val Ala Trp Leu Tyr Gly Asp Pro Arg Gln Val Leu Tyr Pro Arg Asn Ser Thr Gly Ala Tyr Cys Gly Met Gly Glu Asn Lys Asp Lys Pro Tyr Leu Leu Tyr Phe Asn Ile Phe Ser Cys Ile Leu Ser Ser Asn Ile Ile Ser Val Ala Glu Asn Gly Leu Gln Cys Pro Thr Pro Gln Val Cys Val Ser Ser Cys Pro Glu Asp Pro Trp Thr Val Gly Lys Asn Glu Phe Ser Gln Thr Val Gly Glu Val Phe Tyr Thr Lys Asn Arg Asn Phe Cys Leu Pro Gly Val Pro Trp Asn Met Thr Val Ile Thr Ser Leu Gln Gln Glu Leu Cys Pro Ser Phe Leu Leu Pro Ser Ala Pro Ala Leu Gly Arg Cys Phe Pro Trp Thr Asn Ile Thr Pro Pro Ala Leu Pro Gly Ile Thr Asn Asp Thr Thr I1e Gln Gln Gly I1e Ser Gly Leu Ile Asp Ser Leu Asn Ala Arg Asp Ile Ser Val Lys Ile Phe Glu Asp Phe Ala Gln Ser Trp Tyr Trp Ile Leu Val Ala Leu Gly Val Ala Leu Val Leu Ser Leu Leu Phe Ile Leu Leu Leu Arg Leu Val A1a Gly Pro Leu Val Leu Val Leu Ile Leu Gly Val Leu Gly Val Leu Ala Tyr Gly Ile Tyr Tyr Cys Trp Glu Glu Tyr Arg Val Leu Arg Asp Lys Gly Ala Ser Ile Ser Gln Leu Gly Phe Thr Thr Asn Leu Ser Ala Tyr Gln Ser Val Gln Glu Thr Trp Leu Ala Ala Leu Ile Val Leu Ala Val Leu Glu Ala Ile Leu Leu Leu Val Leu I1e Phe Leu Arg Gln Arg Ile Arg Ile Ala Ile Ala Leu Leu Lys Glu Ala Ser Lys Ala Val Gly Gln Met Met Ser Thr Met Phe Tyr Pro Leu Val Thr Phe Val Leu Leu Leu Ile Cys I1e Ala Tyr Trp Ala Met Thr Ala Leu Tyr Leu Ala Thr Ser Gly Gln Pro Gln Tyr Val Leu Trp Ala Ser Asn Ile Ser Ser Pro Gly Cys Glu Lys Val Pro Ile Asn Thr Ser Cys Asn Pro Thr Ala His Leu Val Asn Ser Ser Cys Pro Gly Leu Met Cys Val Phe Gln G1y Tyr Ser Ser Lys Gly Leu Ile Gln Arg Ser Val Phe Asn Leu Gln I1e Tyr Gly Val Leu Gly Leu Phe Trp Thr Leu Asn Trp Val Leu Ala Leu Gly Gln Cys Val Leu Ala Gly Ala Phe Ala Ser Phe Tyr Trp Ala Phe His Lys Pro Gln Asp Ile Pro Thr Phe Pro Leu Ile Ser Ala Phe Ile Arg Thr Leu Arg Tyr His Thr Gly Ser Leu Ala Phe Gly Ala Leu Ile Leu Thr Leu Val Gln Ile Ala Arg Val Ile Leu Glu Tyr Ile Asp His Lys Leu Arg Gly Val Gln Asn Pro Val Ala Arg Cys I1e Met Cys Cys Phe Lys Cys Cys Leu Trp Cys Leu Glu Lys Phe Ile Lys Phe Leu Asn Arg Asn Ala Tyr Ile Met Ile Ala Ile Tyr Gly Lys Asn Phe Cys Val Ser Ala Lys Asn Ala Phe Met Leu Leu Met Arg Asn Ile Val Arg Val Val Val Leu Asp Lys Val Thr Asp Leu Leu Leu Phe Phe Gly Lys Leu Leu Val Val Gly Gly Val Gly Val Leu Ser Phe Phe Phe Phe Ser Gly Arg Ile Pro Gly Leu Gly Lys Asp Phe Lys Ser Pro His Leu Asn Tyr Tyr Trp Leu Pro Ile Met Thr Ser Ile Leu Gly Ala Tyr Val Ile Ala Ser Gly Phe Phe Ser Val Phe Gly Met Cys Val Asp Thr Leu Phe Leu Cys Phe Leu Glu Asp Leu Glu Arg Asn Asn Gly Ser Leu Asp Arg Pro Tyr Tyr Met Ser Lys Ser Leu Leu Lys Ile Leu Gly Lys Lys Asn Glu Ala Pro Pro Asp Asn Lys Lys Arg Lys Lys <210> 23 <211> 476 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 2289333CD1 <400> 23 Glu Gln Asn Phe Asp Gly Thr Ser Asp Glu Glu His Glu Gln Glu Leu Leu Pro Val Gln Lys His Tyr Gln Leu Asp Asp Gln Glu Gly Ile Ser Phe Va1 Gln Thr Leu Met His Leu Leu Lys Gly Asn Ile Gly Thr Gly Leu Leu Gly Leu Pro Leu Ala Ile Lys Asn Ala G1y Ile Val Leu Gly Pro Ile Ser Leu Val Phe Ile Gly Ile Ile Ser Val His Cys Met His Ile Leu Val Arg Cys Ser His Phe Leu Cys Leu Arg Phe Lys Lys Ser Thr Leu Gly Tyr Ser Asp Thr Val Ser Phe A1a Met Glu Val Ser Pro Trp Ser Cys Leu Gln Lys Gln Ala Ala Trp Gly Arg Ser Val Val Asp Phe Phe Leu Val Ile Thr Gln Leu Gly Phe Cys Ser Val Tyr Ile Val Phe Leu A1a Glu Asn Val Lys Gln Va1 His Glu Gly Phe Leu Glu Ser Lys Val Phe Ile Ser Asn Ser Thr Asn Ser Ser Asn Pro Cys Glu Arg Arg Ser Val Asp Leu Arg I1e Tyr Met Leu Cys Phe Leu Pro Phe Ile Ile Leu Leu Val Phe Ile Arg Glu Leu Lys Asn Leu Phe Val Leu Ser Phe Leu Ala Asn Val Ser Met Ala Val Ser Leu Val Ile Ile Tyr Gln Tyr Val Val Arg Asn Met Pro Asp Pro His Asn Leu Pro Ile Val Ala Gly Trp Lys Lys Tyr Pro Leu Phe Phe Gly Thr Ala Val Phe Ala Phe Glu Gly Ile Gly Val Val Leu Pro Leu Glu Asn Gln Met Lys Glu Ser Lys Arg Phe Pro Gln Ala Leu Asn Ile Gly Met Gly Ile Val Thr Thr Leu Tyr Val Thr Leu Ala Thr Leu Gly Tyr Met Cys Phe His Asp Glu Ile Lys Gly Ser Ile Thr Leu Asn Leu Pro Gln Asp Val Trp Leu Tyr Gln Ser Val Lys Ile Leu Tyr Ser Phe Gly Ile Phe Val Thr Tyr Ser Ile Gln Phe Tyr Val Pro Ala Glu Ile Ile Ile Pro Gly Ile Thr Ser Lys Phe His Thr Lys Trp Lys Gln Ile Cys Glu Phe Gly Ile Arg Ser Phe Leu Val Ser I1e Thr Cys Ala Gly Ala Ile Leu Ile Pro Arg Leu Asp Ile Val Ile Ser Phe Val Gly Ala Val Ser Ser Ser Thr Leu Ala Leu Ile Leu Pro Pro Leu Val Glu Ile Leu Thr Phe Ser Lys Glu His Tyr Asn Ile Trp Met Val Leu Lys Asn I1e Ser Ile Ala Phe Thr Gly Val Val Gly Phe Leu Leu Gly Thr Tyr Ile Thr Va1 Glu Glu Ile Ile Tyr Pro Thr Pro Lys Val Val Ala Gly Thr Pro Gln Ser Pro Phe Leu Asn Leu Asn Ser Thr Cys Leu Thr Ser Gly Leu Lys <210> 24 <211> 237 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 2720354CD1 <400> 24 Met Gly Leu Thr Phe Ile Asn Ala Leu Val Phe Gly Val Gln G1y Asn Thr Leu Arg Ala Leu Gly His Asp Ser Pro Leu Asn Gln Phe Leu Ala Gly Ala Ala Ala Gly Ala Ile Gln Cys Val Ile Cys Cys Pro Met Glu Leu Ala Lys Thr Arg Leu Gln Leu Gln Asp Ala Gly Pro Ala Arg Thr Tyr Lys Gly Ser Leu Asp Cys Leu Ala Gln Ile Tyr Gly His Glu Gly Leu Arg Gly Val Asn Arg Gly Met Val Ser Thr Leu Leu Arg Glu Thr Pro Ser Phe Gly Val Tyr Phe Leu Thr Tyr Asp Ala Leu Thr Arg Ala Leu Gly Cys Glu Pro Gly Asp Arg Leu Leu Val Pro Lys Leu Leu Leu Ala Gly Gly Thr Ser Gly Ile Val Ser Trp Leu Ser Thr Tyr Pro Val Asp Val Val Lys Ser Arg Leu Gln Ala Asp Gly Leu Arg Gly Ala Pro Arg Tyr Arg Gly Ile Leu Asp Cys Val His Gln Ser Tyr Arg Ala Glu Gly Trp Arg Val Phe Thr Arg Gly Leu Ala Ser Thr Leu Leu Arg Ala Phe Pro Val Asn Ala Ala Thr Phe Ala Thr Val Thr Val Val Leu Thr Tyr Ala Arg Gly Glu Glu Ala Gly Pro Glu Gly Glu Ala Val Pro Ala Ala Pro Ala Gly Pro Ala Leu Ala Gln Pro Ser Ser Leu <210> 25 <211> 345 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 3038193CD1 <400> 25 Met Arg Leu Leu G1u Arg Met Arg Lys Asp Trp Phe Met Val Gly Ile Val Leu Ala Ile Ala Gly Ala Lys Leu Glu Pro Ser Ile Gly Val Asn Gly Gly Pro Leu Lys Pro Glu Ile Thr Val Ser Tyr Ile Ala Val Ala Thr Ile Phe Phe Asn Ser Gly Leu Ser Leu Lys Thr Glu Glu Leu Thr Ser Ala Leu Val His Leu Lys Leu His Leu Phe Ile Gln Ile Phe Thr Leu Ala Phe Phe Pro Ala Thr Ile Trp Leu Phe Leu G1n Leu Leu Ser Ile Thr Pro Ile Asn Glu Trp Leu Leu Lys Gly Leu Gln Thr Val G1y Cys Met Pro Pro Pro Val Ser Ser Ala Val Ile Leu Thr Lys Ala Val G1y Gly Asn Glu Gly Ile Val Ile Thr Pro Leu Leu Leu Leu Leu Phe Leu Gly Ser Ser Ser Ser Val Pro Phe Thr Ser Ile Phe Ser Gln Leu Phe Met Thr Val Val Val Pro Leu Ile Ile Gly Gln Ile Val Arg Arg Tyr Ile Lys Asp Trp Leu Glu Arg Lys Lys Pro Pro Phe Gly Ala Ile Ser Ser Ser Val Leu Leu Met Ile Ile Tyr Thr Thr Phe Cys Asp Thr Phe Ser Asn Pro Asn Ile Asp Leu Asp Lys Phe Ser Leu Val Leu Ile Leu Phe Ile Ile Phe Ser Ile Gln Leu Ser Phe Met Leu Leu Thr Phe Ile Phe Ser Thr Arg Asn Asn Ser Gly Phe Thr Pro Ala Asp Thr Val Ala Ile Ile Phe Cys Ser Thr His Lys Ser Leu Thr Leu Gly Il.e Pro Met Leu Lys Ile Val Phe Ala Gly Tyr Glu His Leu Ser Leu Ile Ser Val Pro Leu Leu Ile Tyr His Pro Ala Gln Ile Leu Leu Gly Ser Val Leu Val Pro Thr Ile Lys Ser Trp Met Val Ser Arg Gln Lys Lys Leu Leu Gln Thr Arg Gly Pro Leu Ala Asn Leu Asn Asn Pro Glu Gly Leu Glu Tyr Leu Ser Ile Lys Phe Gly His <210> 26 <211> 521 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 3460979CD1 <400> 26 Met Ala Ala Leu Ala Pro Val Gly Ser Pro Ala Ser Arg Gly Pro Arg Leu Ala Ala Gly Leu Arg Leu Leu Pro Met Leu Gly Leu Leu Gln Leu Leu Ala Glu Pro Gly Leu Gly Arg Val His His Leu A1a Leu Lys Asp Asp Val Arg His Lys Val His Leu Asn Thr Phe Gly Phe Phe Lys Asp Gly Tyr Met Val Val Asn Val Ser Ser Leu Ser Leu Asn Glu Pro Glu Asp Lys Asp Val Thr Ile Gly Phe Ser Leu Asp Arg Thr Lys Asn Asp Gly Phe Ser Ser Tyr Leu Asp Glu Asp Val Asn Tyr Cys Ile Leu Lys Lys Gln Ser Val Ser Val Thr Leu Leu Ile Leu Asp Ile Ser Arg Ser Glu Val Arg Val Lys Ser Pro Pro Glu Ala Gly Thr Gln Leu Pro Lys Ile Ile Phe Ser Arg Asp Glu Lys Va1 Leu Gly Gln Ser Gln Glu Pro Asn Val Asn Pro Ala Ser Ala G1y Asn Gln Thr Gln Lys Thr Gln Asp Gly Gly Lys Ser Lys Arg Ser Thr Val Asp Ser Lys Ala Met Gly Glu Lys Ser Phe Ser Val His Asn Asn Gly Gly Ala Val Ser Phe Gln Phe Phe Phe Asn Ile Ser Thr Asp Asp Gln Glu Gly Leu Tyr Ser Leu Tyr Phe His Lys Cys Leu Gly Lys Glu Leu Pro Ser Asp Lys Phe Thr Phe Ser Leu Asp Ile Glu Tle Thr Glu Lys Asn Pro Asp Ser Tyr Leu Ser Ala Gly Glu Ile Pro Leu Pro Lys Leu Tyr Ile Ser Met A1a Phe Phe Phe Phe Leu Ser Gly Thr Ile Trp Ile His Ile Leu Arg Lys Arg Arg Asn Asp Val Phe Lys Ile His Trp Leu Met Ala Ala Leu Pro Phe Thr Lys Ser Leu Ser Leu Val Phe His Ala Ile Asp Tyr His Tyr Ile Ser Ser Gln Gly Phe Pro Ile Glu Gly Trp Ala Val Val Tyr Tyr Ile Thr His Leu Leu Lys Gly Ala Leu Leu Phe Ile Thr Ile Ala Leu Ile Gly Thr Gly Trp Ala Phe Ile Lys His Ile Leu Ser Asp Lys Asp Lys Lys Ile Phe Met Ile Val Ile Pro Leu Gln Val Leu Ala Asn Val Ala Tyr Ile Ile Ile Glu Ser Thr Glu Glu Gly Thr Thr Glu Tyr Gly Leu Trp Lys Asp Ser Leu Phe Leu Val Asp Leu Leu Cys Cys Gly Ala Ile Leu Phe Pro Val Val Trp Ser Ile Arg His Leu Gln Glu Ala Ser Ala Thr Asp Gly Lys Ala Ala Ile Asn Leu Ala Lys Leu Lys Leu Phe Arg His Tyr Tyr Val Leu Ile Val Cys Tyr Ile Tyr Phe Thr Arg Ile Ile Ala Phe Leu Leu Lys Leu Ala Val Pro Phe Gln Trp Lys Trp Leu Tyr Gln Leu Leu Asp Glu Thr Ala Thr Leu Val Phe Phe Val Leu Thr Gly Tyr Lys Phe Arg Pro Ala Ser Asp Asn Pro Tyr Leu Gln Leu Ser Gln Glu Glu Glu Asp Leu Glu Met G1u Ser Val <210> 27 <211> 555 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 7472200CD1 <400> 27 Met Thr Leu Val Tyr Phe Pro Pro Ser Lys Leu Gln Gln Gln Gln Gln Pro Ser Arg Ser Ser Arg Leu Ala Gln Gln Leu Ala Gln Ser Ser Trp Gln Leu Ala Leu Arg Phe Gly Lys Arg Thr Thr I1e His Gly Leu Asp Arg Leu Leu Ser Ala Lys A1a Ser Arg Trp Glu Arg Phe Val Trp Leu Cys Thr Phe Val Ser Ala Phe Leu Gly Ala Val Tyr Val Cys Leu Ile Leu Ser Ala Arg Tyr Asn Ala Ala His Phe Gln Thr Val Val Asp Ser Thr Arg Phe Pro Val Tyr Arg Ile Pro Phe Pro Val Ile Thr Ile Cys Asn Arg Asn Arg Leu Asn Trp Gln Arg Leu Ala Glu Ala Lys Ser Arg Phe Leu Ala Asn Gly Ser Asn Ser Ala Gln Gln Glu Leu Phe Glu Leu Ile Val Gly Thr Tyr Asp Asp Ala Tyr Phe Gly His Phe Gln Ser Phe Glu Arg Leu Arg Asn Gln Pro Thr Glu Leu Leu Asn Tyr Val Asn Phe Ser Gln Val Val Asp Phe Met Thr Trp Arg Cys Asn Glu Leu Leu Ala Glu Cys Leu Trp Arg His His Ala Tyr Asp Cys Cys Glu Ile Arg Ser Lys Arg Arg Ser Lys Asn Gly Leu Cys Trp Ala Phe Asn Ser Leu Glu Thr Glu Glu Gly Arg Arg Met Gln Leu Leu Asp Pro Met Trp Pro Trp Arg Thr Gly Ser Ala Gly Pro Met Ser Ala Leu Ser Val Arg Val Leu Ile Gln Pro Ala Lys His Trp Pro Gly His Arg Glu Thr Asn Ala Met Lys Gly Ile Asp Val Met Val Thr Glu Pro Phe Val Trp His Asn Asn Pro Phe Phe Val A1a Ala Asn Thr Glu Thr Thr Met Glu Ile Glu Pro Val Ile Tyr Phe Tyr Asp Asn Asp Thr Arg Gly Val Arg Ser Asp Gln Arg Gln Cys Val Phe Asp Asp Glu His Asn Ser Lys Asp Phe Lys Ser Leu Gln Gly Tyr Val Tyr Met Ile Glu Asn Cys Gln Ser Glu Cys His Gln Glu Tyr Leu Val Arg Tyr Cys Asn Cys Thr Met Asp Leu Leu Phe Pro Pro Asp Leu Leu Ile Tyr Ser His Asn Pro Gly Glu Lys Glu Phe Val Arg Asn Gln Phe Gln Gly Met Ser Cys Lys Cys Phe Arg Asn Cys Tyr Ser Leu Asn Tyr Ile Ser Asp Val Arg Pro Ala Phe Leu Pro Pro Asp Val Tyr Ala Asn Asn Ser Tyr Val Asp Leu Asp Val His Phe Arg Phe Glu Thr Ile Met Val Tyr Arg Thr Ser Leu Val Phe Gly Trp Val Asp Leu Met Val Ser Phe Gly Gly Ile Ala Gly Leu Phe Leu Gly Cys Ser Leu Ile Ser Gly Met Glu Leu Ala Tyr Phe Leu Cys Ile Glu Val Pro Ala Phe Gly Leu Asp Gly Leu Arg Arg Arg Trp Lys Ala Arg Arg Gln Met Asp Leu Gly Val Thr Val Pro Thr Pro Thr Leu Asn Phe Gln Gln Thr Thr Pro Ser Gln Leu Met Glu Asn Tyr Ile Met Gln Leu Lys Ala Glu Lys Ala Gln Gln Gln Lys Ala Asn Phe Gln Asn Trp His Arg Ile Thr Phe Ala Gln Lys His Val Ile Gly Lys <210> 28 <211> 2080 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte TD No: 1416107CB1 <400> 28 ggcggttcag gcgccagagc tggccgatcg gcgttggccg ccgacatgac gcccgaggac 60 ccagaggaaa cccagccgct tctggggcct cctggcggca gcgcgccccg cggccgccgc 120 gtcttcctcg ccgccttcgc cgctgccctg ggcccactca gcttcggctt cgcgctcggc 180 tacagctccc cggccatccc tagcctgcag cgcgccgcgc ccccggcccc gcgcctggac 240 gacgccgccg cctcctggtt cggggctgtc gtgaccctgg gtgccgcggc ggggggagtg 300 ctgggcggct ggctggtgga ccgcgccggg cgcaagctga gcctcttgct gtgctccgtg 360 cccttcgtgg ccggctttgc cgtcatcacc gcggcccagg acgtgtggat gctgctgggg 420 ggccgcctcc tcaccggcct ggcctgcggt gttgcctccc tagtggcccc ggtctacatc 480 tccgaaatcg cctacccagc agtccggggg ttgctcggct cctgtgtgca gctaatggtc 540 gtcgtcggca tcctcctggc ctacctggca ggctgggtgc tggagtggcg ctggctggct 600 gtgctgggct gcgtgccccc ctccctcatg ctgcttctca tgtgcttcat gcccgagacc 660 ccgcgcttcc tgctgactca gcacaggcgc caggaggcca tggccgccct gcggttcctg 720 tggggctccg agcagggctg ggaagacccc cccatcgggg ctgagcagag ctttcacctg 780 gccctgctgc ggcagcccgg catctacaag cccttcatca tcggcgtctc cctgatggcc 840 ttccagcagc tgtcgggggt caacgccgtc atgttctatg cagagaccat ctttgaagag 900 gccaagttca aggacagcag cctggcctcg gtcgtcgtgg gtgtcatcca ggtgctgttc 960 acagctgtgg cggctctcat catggacaga gcagggcgga ggctgctcct ggtcttgtca 1020 ggtgtggtca tggtgttcag cacgagtgcc ttcggcgcct acttcaagct gacccagggt 1080 ggccctggca actcctcgca cgtggccatc tcggcgcctg tctctgcaca gcctgttgat 1140 gccagcgtgg ggctggcctg gctggccgtg ggcagcatgt gcctcttcat cgccggcttt 1200 gcggtgggct gggggcccat cccctggctc ctcatgtcag agatcttccc tctgcatgtc 1260 aagggcgtgg cgacaggcat ctgcgtcctc accaactggc tcatggcctt tctcgtgacc 1320 aaggagttca gcagcctcat ggaggtcctc aggccctatg gagccttctg gcttgcctcc 1380 gctttctgca tcttcagtgt ccttttcact ttgttctgtg tccctgaaac taaaggaaag 1440 actctggaac aaatcacagc ccattttgag gggcgatgac agccactcac taggggatgg 1500 agcaagcctg tgactccaag ctgggcccaa gcccagagcc cctgcctgcc ccaggggagc 1560 cagaatccag ccccttggag ccttggtctg cagggtccct ccttcctgtc atgctccctc 1620 cagcccatga cccggggcta ggaggctcac tgcctcctgt tccagctcct gctgctgctc 1680 tgaggactca ggaacacctt cgagctttgc agacctgcgg tcagccctcc atgcgcaaga 1740 ctaaagcagc ggaagaggag gtgggcctct aggatctttg tcttctggct ggaggtgctt 1800 ttggaggttg ggtgctgggc attcagtcgc toctctcacg cggctgcctt atcgggaagg 1860 aaatttgttt gccaaataaa gactgacaca gaaaatcagg tcagtgtctc tgggctttgt 1920 gcaagctcag tttgaaaagg gtttattccc atcactgccc aggacaccct gtggctttac 1980 ttgctcatgg tcagccaagc ttacccttca cactgagaag tcatttctgg ctacttcctt 2040 gggctcagtt ccctgggtca tcagccatca aatcttgttg 2080 <210> 29 <211> 2128 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 1682513CB1 <400> 29 ctggccctag ggagctgccc ctgtcgctgg ctgcctgcac caaccagccc cacattgtca 60 actacctgac ggagaacccc cacaagaagg cggacatgcg gcgccaggac tcgcgaggca 120 acacagtgct gcatgcgctg gtggccattg ctgacaacac ccgtgagaac accaagtttg 180 ttaccaagat gtacgacctg ctgctgctca agtgtgcccg cctcttcccc gacagcaacc 240 tggaggccgt gctcaacaac gacggcctct cgcccctcat gatggctgcc aagacgggca 300 agattgggaa ccgccacgag atgctggctg tggagcccat caatgaactg ctgcgggaca 360 agtggcgcaa gttcggggcc gtctccttct acatcaacgt ggtctcctac ctgtgtgcca 420 tggtcatctt cactctcacc gcctactacc agccgctgga gggcacaccg ccgtaccctt 480 accgcaccac ggtggactac ctgcggctgg ctggcgaggt cattacgctc ttcactgggg 540 tcctgttctt cttcaccaac atcaaagact tgttcatgaa gaaatgccct ggagtgaatt 600 ctctcttcat tgatggctcc ttccagctgc tctacttcat ctactctgtc ctggtgatcg 660 tctcagcagc cctctacctg gcagggatcg aggcctacct ggccgtgatg gtctttgccc 720 tggtcctggg ctggatgaat gccctttact tcacccgtgg gctgaagctg acggggacct 780 atagcatcat gatccagaag attctcttca aggacctttt ccgattcctg ctcgtctact 840 tgctcttcat gatcggctac gcttcagccc tggtctccct cctgaacccg tgtgccaaca 900 tgaaggtgtg caatggggac cagaccaact gcacagtgcc cacttacccc tcgtgccgtg 960 acagcgagac cttcagcacc ttcctcctgg acctgtttaa gctgaccatc ggcatgggcg 1020 acctggagat gctgagcagc accaagtacc ccgtggtctt catcatcctg ctggtgacct 1080 acatcatcct cacctttgtg ctgctcctca acatgctcat tgccctcatg ggcgagacag 1140 tgggccaggt ctccaaggag agcaagcaca tctggaagct gcagtgggcc accaccatcc 1200 tggacattga gcgctccttc cccgtattcc tgaggaagtc cttccgctct ggggagatgg 1260 tcaccgtggg caagagctcg gacggcactc ctgaccgcag gtggtgcttc agggtggatg 1320 aggtgaactg gtctcactgg aaccagaact tgggcatcat caacgaggac ccgggcaaga 1380 atgagaccta ccagtattat ggcttctcgc ataccgtggg ccgcctccgc agggatcgct 1440 ggtcctcggt ggtaccccgc gtggtggaac tgaacaagaa ctcgaacccg gacgaggtgg 1500 tggtgcctct ggacagcacg gggaaccccc gctgcgatgg ccaccagcag ggttaccccc 1560 gcaagtggag gactgatgac gccccgctct agggactgca gcccagcccc agcttctctg 1620 cccactcatt tctagtccag ccgcatttca gcagtgcctt ctggggtgtc cccccacacc 1680 ctgctttggc cccagaggcg agggaccagt ggaggtgcca gggaggcccc aggaccctgt 1740 ggtcccctgg ctctgcctcc ccaccctggg gtgggggctc ccggccacct gtcttgctcc 1800 tatggagtca cataagccaa cgccagagcc cctccacctc aggccccagc ccctgcctct 1860 ccattattta tttgctctgc tctcaggaag cgacgtgacc cctgccccag ctggaacctg 1920 gcagaggcct taggaccccg ttccaagtgc actgcccggc caagccccag cctcagcctg 1980 cgcctgagct gcatgcgcca ccatttttgg cagcgtggca gctttgcaag gggctggggc 2040 cctcggcgtg gggccatgcc ttctgtgtgt tctgtagtgt ctgggatttg ccggtgctca 2100 ataaatgttt attcattgaa aaaaaaaa 2128 <210> 30 <211> 2825 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 2446438CB1 <400> 30 cgttgtgcac gtaattcggc tcgacgtgtg tccagatggt cagtctctgg tggctagcct 60 gtcctgacag gggagagtta agctcccgtt ctccaccgtg ccggctggcc aggtgggctg 120 agggtgaccg agagaccaga acctgcttgc tggagcttag tgctcagagc tggggaggga 180 ggttccgccg ctcctctgct gtcagcgccg gcagcccctc ccggcttcac ttcctcccgc 240 agcccctgct actgagaagc tccgggatcc cagcagccgc cacgccctgg cctcagcctg 300 cggggctcca gtcaggccaa caccgacgcg cagctgggag gaagacagga cccttgacat 360 ctccatctgc acagaggtcc tggctggacc gagcagcctc ctcctcctag gatgacctca 420 ccctccagct ctccagtttt caggttggag acattagatg caggccaaga agatggctct 480 gaggcggaca gaggaaagct ggattttggg agcgggctgc ctcccatgga gtcacagttc 540 cagggcgagg accggaaatt cgcccctcag ataagagtca acctcaacta ccgaaaggga 600 acaggtgcca gtcagccgga tccaaaccga tttgaccgag atcggctctt caatgcggtc 660 tcccggggtg tccccgagga tctggctgga cttccagagt acctgagcaa gaccagcaag 720 tacctcaccg actcggaata cacagagggc tccacaggta agacgtgcct gatgaaggct 780 gtgctgaacc ttaaggacgg ggtcaatgcc tgcattctgc cactgctgca gatcgaccgg 840 gactctggca atcctcagcc cctggtaaat gcccagtgca cagatgacta ttaccgaggc 900 cacagcgctc tgcacatcgc cattgagaag aggagtctgc agtgtgtgaa gctcctggtg 960 gagaatgggg ccaatgtgca tgcccgggcc tgcggccgct tcttccagaa gggccaaggg 1020 acttgctttt atttcggtga gctacccctc tctttggccg cttgcaccaa gcagtgggat 1080 gtggtaagct acctcctgga gaacccacac cagcccgcca gcctgcaggc cactgactcc 1140 cagggcaaca cagtcctgca tgccctagtg atgatctcgg acaactcagc tgagaacatt 1200 gcactggtga ccagcatgta tgatgggctc ctccaagctg gggcccgcct ctgccctacc 1260 gtgcagcttg aggacatccg caacctgcag gatctcacgc ctctgaagct ggccgccaag 1320 gagggcaaga tcgagatttt caggcacatc ctgcagcggg agttttcagg actgagccac 1380 ctttcccgaa agttcaccga gtggtgctat gggcctgtcc gggtgtcgct gtatgacctg 1440 gcttctgtgg acagctgtga ggagaactca gtgctggaga tcattgcctt tcattgcaag 1500 agcccgcacc gacaccgaat ggtcgttttg gagcccctga acaaactgct gcaggcgaaa 1560 tgggatctgc tcatccccaa gttcttctta aacttcctgt gtaatctgat ctacatgttc 1620 atcttcaccg ctgttgccta ccatcagcct accctgaaga agcaggccgc ccctcacctg 1680 aaagcggagg ttggaaactc catgctgctg acgggccaca tccttatcct gctagggggg 1740 atctacctcc tcgtgggcca gctgtggtac ttctggcggc gccacgtgtt catctggatc 1800 tcgttcatag acagctactt tgaaatcctc ttcctgttcc aggccctgct cacagtggtg 1860 tcccaggtgc tgtgtttcct ggccatcgag tggtacctgc ccctgcttgt gtctgcgctg 1920 gtgctgggct ggctgaacct gctttactat acacgtggct tccagcacac aggcatctac 1980 agtgtcatga tccagaaggt catcctgcgg gacctgctgc gcttccttct gatctactta 2040 gtcttccttt tcggcttcgc tgtagccctg gtgagcctga gccaggaggc ttggcgcccc 2100 gaagctccta caggccccaa tgccacagag tcagtgcagc ccatggaggg acaggaggac 2160 gagggcaacg gggcccagta caggggtatc ctggaagcct ccttggagct cttcaaattc 2220 accatcggca tgggcgagct ggccttccag gagcagctgc acttccgcgg catggtgctg 2280 ctgctgctgc tggcctacgt gctgctcacc tacatcctgc tgctcaacat gctcatcgcc 2340 ctcatgagcg agaccgtcaa cagtgtcgcc actgacagct ggagcatctg gaagctgcag 2400 aaagccatct ctgtcctgga gatggagaat ggctattggt ggtgcaggaa gaagcagcgg 2460 gcaggtgtga tgctgaccgt tggcactaag ccagatggca gccccgatga gcgctggtgc 2520 ttcagggtgg aggaggtgaa ctgggcttca tgggagcaga cgctgcctac gctgtgtgag 2580 gacccgtcag gggcaggtgt ccctcgaact ctcgagaacc ctgtcctggc ttcccctccc 2640 aaggaggatg aggatggtgc ctctgaggaa aactatgtgc ccgtccagct cctccagtcc 2700 aactgatggc ccagatgcag caggaggcca gaggacagag cagaggatct ttccaaccac 2760 atctgctggc tctggggtcc cagtgaattc tggtggcaaa tatatatttt cactaaaaaa 2820 aaaaa 2825 <210> 31 <211> 1718 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 2817822CB1 <400> 31 gcctcggtgt tcccacctag gggcgggcag ccaggggcac ttccgctggc ccaagtgatc 60 tgcatgtggc agggctgcgc agtggagcgg ccagtgggca ggatgacgag ccagacccct 120 ctgccccagt ccccccggcc caggcggcca acgatgtcta ctgttgtgga gctgaacgtc 180 gggggtgagt tccacaccac caccctgggt accctgagga agtttccggg ctcaaagctg 240 gcagagatgt tctctagctt agccaaggcc tccacggacg cggagggccg cttcttcatc 300 gaccgcccca gcacctattt cagacccatc ctggactacc tgcgcactgg gcaagtgccc 360 acacagcaca tccctgaagt gtaccgtgag gctcagttct acgaaatcaa gcctttggtc 420 aagctgctgg aggacatgcc acagatcttt ggtgagcagg tgtctcggaa gcagtttttg 480 ctgcaagtgc cgggctacag cgagaacctg gagctcatgg tgcgcctggc acgtgcagaa 540 gccataacag cacggaagtc cagcgtgctt gtgtgcctgg tggaaactga ggagcaggat 600 gcatattatt cagaggtcct gtgttttctg caggataaga agatgttcaa gtctgttgtc 660 aagtttgggc cctggaaggc ggtcctagac aacagcgacc tcatgcactg cctggagatg 720 gacattaagg cccaggggta caaggtattc tccaagttct acctgacgta ccccaccaaa 780 agaaacgaat tccattttaa catttattca ttcaccttca cctggtggtg atcctcagga 840 gcagagactg ttatgaattc tggcgtggct tatgaaatta aaagttgcca tcaaagccat 900 tttcttttaa tttcacaaac atcaggcaat ttccagggtt ggtctagagt cttgccacta 960 aatattgatc actcgtttaa ggactttcca ctccattgca actgatgcca ctatatttgc 1020 ctagcaactt gcagctactt ccttttcaaa gcctcatgta tctcccagac ccttctcttg 1080 aagtccaata acaagaccaa gtaagaatgt ttcaacaatg cgttggcaag agatgtgaga 1140 tgacaacagg aacatacaag atactgtgaa tctagatgtt ctgacctaaa gatgtagtct 1200 acatagcccc agcttggggt ccaatccatc tgtccctggc atgtgccttc atgtagtagg 1260 tgctttcctg atcccctttg cgagatgctg tgggtgctaa cacctcagag ctgtcctctt 1320 ctctagagtg gaggttttca aagtgcatca tcagcattac ctgtgaactt gctggaaata 1380 caaatcctca ggccccacct cagacctact gaatcagaat ctctgggggt tggcacagca 1440 ttctgattta ccaaaccctc caagtgattt tgatgtattc taattttgag accatctcta 1500 gaaaagaatt gctacctctt gtatggaggt acaaaagact gacctcttac atcaaggaac 1560 ttcctttccc agagctcctc atggaatcaa gctgaagtca gtcttcttct gagagcacat 1620 tcttactcag tttttttcct ctgtcctacg ctgcttccct cactcccctt ctcctaagag 1680 cactccatca ataaaccact tgcacgagaa aaaaaaaa 1718 <210> 32 <211> 2000 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 4009329CB1 <400> 32 gacgaatttg aaaccagggg gtgtcctgtt tgaacttggt gccagataga gtaactcgga 60 ctccagttgg aggggttcgg gagaaccata gaagaggaag ggccgtgtct tccgtggaca 120 ggccaccgga gccgccagct gtttggaact gagctactgc agaaagggaa gtggagagta 180 agggccaggc cccgtggggg cagatggccg gcagaaggct gaatctgcgc tgggcactga 240 gtgtgctttg tgtgctgcta atggcggaga cagtgtctgg gactaggggc tcgtctacag 300 gagctcacat tagcccccag tttccagctt caggtgtgaa ccagaccccc gtggtagact 360 gccgcaaggt gtgtggcctg aatgtctctg accgctgtga cttcatccgg accaaccctg 420 actgccacag tgatgggggg tacctggact acctggaagg catcttctgc cacttccctc 480 ccagcctcct ccctctggct gtcactctct acgtttcctg gctgctctac ctgtttctga 540 ttctgggagt caccgcagcc aagtttttct gccccaactt gtcggccatt tctaccacac 600 tgaagctctc ccacaacgtg gcaggcgtca ccttcctggc atttgggaat ggtgcacctg 660 acatcttcag tgccctggtg gccttctctg acccgcacac agccggcctg gcccttgggg 720 cactgtttgg cgctggcgtg ctggttacca cagtggtggc cggaggcatt accatcctac 780 accccttcat ggctgcctcc aggcccttct tcagggacat cgttttctac atggtggctg 840 aactgatggc ccagatgcag caggaggcca gaggacagag cagaggatct ttccaacc tgttcctgac cttcctcatg ctcttccgtg gcagggtcac cctggcatgg gctctgggtt 900 acctgggctt gtatgtgttc tatgtggtca ctgtgattct ctgcacctgg atctaccaac 960 ggcaacggag aggatctctg ttctgcccca tgccagttac tccagagatc ctctcagact 1020 ccgaggagga ccgggtatct tctaatacca acagctatga ctacggtgat gagtaccggc 1080 cgctgttctt ctaccaggag accacggctc agatcctggt ccgggccctc aatcccctgg 1140 attacatgaa gtggagaagg aaatcagcat actggaaagc cctcaaggtg ttcaagctgc 1200 ctgtggagtt cctgctgctc ctcacagtcc ccgtcgtgga cccggacaag gatgaccaga 1260 actggaaacg gcccctcaac tgtctgcatc tggttatcag ccccctggtt gtggtcctga 1320 ccctgcagtc ggggacctat ggtgtctatg agataggcgg cctcgttccc gtctgggtcg 1380 tggtggtgat cgcaggcaca gccttggctt cagtgacctt ttttgccaca tctgacagcc 1440 agccccccag gcttcactgg ctctttgctt tcctgggctt tctgaccagc gccctgtgga 1500 tcaacgcggc cgccacagag gtggtgaaca tcttgcggtc cctgggtgtg gtcttccggc 1560 tgagcaacac tgtgctgggg ctcacgctgc tggcctgggg gaacagcatt ggagatgcct 1620 tctcggattt cacactggct cgccagggct acccacggat ggcgttctcc gcctgctttg 1680 gcggcatcat cttcaacatc ctcgtgggtg tggggctggg ctgcctgctc cagatctccc 1740 gaagccacac agaagtgaag ctggagccag acggactgct ggtgtgggtc ctggcaggcg 1800 ccctggggct cagcctcgtc ttctccctgg tctcagtccc attgcagtgc ttccagctca 1860 gcagagtcta tggcttctgc ctgctcctct tctacctgaa cttccttgtc gtggccctcc 1920 tcattgaatt tggagtgatt cacctgaaaa gcatgtgact gaagccgctt agtgctgtgg 1980 cctcactgca ggcaggagcc 2000 <210> 33 <211> 2216 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 6618083CB1 <400> 33 gaaaactctt cctgaaggag atgcagagga agattcgaac tggaggaaaa ccctaaaata 60 aacaataaca acaaaagttc aaaacctgaa aagtgaacca tgaagctcag taaaaaggac 120 cgaggagaag atgaagaaag tgattcagcg aaaaagaaat tggactggtc ctgctcgctc 180 ctcgtggcct ccctcgcggg cgccttcggc tcctccttcc tctacggcta caacctgtcg 240 gtggtgaatg cccccacccc gtacatcaag gccttttaca atgagtcatg ggaaagaagg 300 catggacgtc caatagaccc agacactctg actttgctct ggtctgtgac tgtgtccata 360 ttcgccatcg gtggacttgt ggggacgtta attgtgaaga tgattggaaa ggttcttggg 420 aggaagcaca ctttgctggc caataatggg tttgcaattt ctgctgcatt gctgatggcc 480 tgctcgctcc aggcaggagc ctttgaaatg ctcatcgtgg gacgcttcat catgggcata 540 gatggaggcg tcgccctcag tgtgctcccc atgtacctca gtgagatctc acccaaggag 600 atccgtggct ctctggggca ggtgactgcc atctttatct gcattggcgt gttcactggg 660 cagcttctgg gcctgcccga gctgctggga aaggagagta cctggccata cctgtttgga 720 gtgattgtgg tccctgccgt tgtccagctg ctgagccttc cctttctccc ggacagccca 780 cgctacctgc tcttggagaa gcacaacgag gcaagagctg tgaaagcctt ccaaacgttc 840 ttgggtaaag cagacgtttc ccaagaggta gaggaggtcc tggctgagag ccacgtgcag 900 aggagcatcc gcctggtgtc cgtgctggag ctgctgagag ctccctacgt ccgctggcag 960 gtggtcaccg tgattgtcac catggcctgc taccagctct gtggcctcaa tgcaatttgg 1020 ttctatacca acagcatctt tggaaaagct gggatccctc tggcaaagat cccatacgtc 1080 accttgagta cagggggcat cgagactttg gctgccgtct tctctggttt ggtcattgag 1140 cacctgggac ggagacccct cctcattggt ggctttgggc tcatgggcct cttctttggg 1200 accctcacca tcacgctgac cctgcaggac cacgccccct gggtccccta cctgagtatc 1260 gtgggcattc tggccatcat cgcctctttc tgcagtgggc cagctgtttt cccagaagaa 1320 acggtaaatg tcagcattgt atctgagtga aaagttgacc ttcttcccca cccatgcaca 1380 caaacaagcc agattggact catctgcata tctgcctgaa gttctttgct aaccaaaaat 1440 cactaagctt agccttctct gttttttttt tcctaagccc tcccaagact ttttgcaatg 1500 atcctgattc tgttccaagt gtttgcaact gtggctttct tttgactgta gaacatgctg 1560 catttccagg gctttaaatg ctgggctccc catcagtgtc tatgggactc cctggaggga 1620 aggccacctg cacctcccaa tcccagatca cctgtcagcc cctgccctcc gcttcctcaa 1680 tccatcttca accccctgtg ttgacccagc acctgggcct tgctggctag caatgacttt 1740 agccacaaga tggaccaggg tttagaagct tcatttaaac tcacattgac agtgtacagt 1800 ttaaagcctc agggaactta cctgtctaag aaaagctgcc acttagacca tgagaccatc 1860 ttgcatcttc ctaagtggac agggaagagc aagtccccag gggagccacc cgggaaagtg 1920 tggcaggaag atgctcagag ctgaatggca gagagactca tgggcctgct ctccatgatt 1980 aaagaagagg gatggatctc ccaggagagg gccaggaggc cgcctgaggc agcttctgtg 2040 aggaacaggt cgatgtaaga agacttgaca aggagttgaa attaggtgaa agcaaagaaa 2100 gaaaacaaga gaggcagttt cctgctgcat attttatttg tgtgcataac cccaaggcag 2160 tggcagggaa gtctaataaa tgaggcaaaa taaaagagct tcacctttta aaaaaa 2216 <210> 34 <211> 1995 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 7472002CB1 <400> 34 atgaccgaaa aaaccaatgg tgtgaagagc tccccagcca ataatcacaa ccatcatgca 60 cctcctgcca tcaaggccaa tggcaaagat gaccacagga caagcagcag gccacactct 120 gcagctgacg atgacacctc ctcagaactg cagaggctgg cagacgtgga tgccccacag 180 cagggaagga gtggcttccg caggatagtt cgcctggtgg ggatcatcag agaatgggcc 240 aacaagaatt tccgagagga ggaacctagg cctgactcat tcctcgagcg ttttcgtggg 300 cctgaactcc agactgtgac cacacaggag ggggatggca aaggcgacaa ggatggcgag 360 gacaaaggca ccaagaagaa atttgaacta tttgtcttgg acccagctgg ggattggtac 420 tactgctggc tatttgtcat tgccatgccc gtcctttaca actggtgcct gctggtggcc 480 agagcctgct tcagtgacct acagaaaggc tactacctgg tgtggctggt gctggattat 540 gtctcagatg tggtctacat tgcggacctc ttcatccgat tgcgcacagg tttcctggag 600 caggggctgc tggtcaaaga taccaagaaa ctgcgagaca actacatcca caccctgcag 660 ttcaagctgg atgtggcttc catcatcccc actgacctga tctattttgc tgtggacatc 720 cacagccctg aggtgcgctt caaccgcctg ctgcactttg cccgcatgtt tgagttcttt 780 gaccggacag agacacgcac caactaccct aacatcttcc gcatcagcaa ccttgtcctc 840 tacatcttgg tcatcatcca ctggaatgcc tgcatctatt atgccatctc caaatccata 900 ggctttgggg tcgacacctg ggtttaccca aacatcactg accctgagta tggctacctg 960 gctagggaat acatctattg cctttactgg tccacactga ctctcactac cattggggag 1020 acaccacccc ctgtaaagga tgaggagtac ctatttgtca tctttgactt cctgattggc 1080 gtcctcatct ttgccaccat cgtgggaaat gtgggctcca tgatctccaa catgaatgcc 1140 acccgggcag agttccaggc taagatcgat gccgtgaaac actacatgca gttccgaaag 1200 gtcagcaagg ggatggaagc caaggtcatt aggtggtttg actacttgtg gaccaataag 1260 aagacagtgg atgagcgaga aattctcaag aatctgccag ccaagctcag ggctgagata 1320 gccatcaatg tccacttgtc cacactcaag aaagtgcgca tcttccatga ttgtgaggct 1380 ggcctgctgg tagagctggt actgaaactc cgtcctcagg tcttcagtcc tggggattac 1440 atttgccgca aaggggacat cggcaaggag atgtacatca ttaaggaggg caaactggca 1500 gtggtggctg atgatggtgt gactcagtat gctctgctgt cggctggaag ctgctttggc 1560 gagatcagta tccttaacat taagggcagt aaaatgggca atcgacgcac agctaatatc 1620 cgcagcctgg gctactcaga tctcttctgc ttgtccaagg atgatcttat ggaagctgtg 1680 actgagtacc ctgatgccaa gaaagtccta gaagagaggg gtcgggagat cctcatgaag 1740 gagggactgc tggatgagaa cgaagtggca accagcatgg aggtcgacgt gcaggagaag 1800 ctagggcagc tggagaccaa catggaaacc ttgtacactc gctttggccg cctgctggct 1860 gagtacacgg gggcccagca gaagctcaag cagcgcatca cagttctgga aaccaagatg 1920 aaacagaaca atgaagatga ctacctgtct gatgggatga acagccctga gctggctgct 1980 gctgacgagc cataa 1995 <210> 35 <211> 988 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1812692CB1 <400> 35 cttgggtgaa agaaaatcct gcttgacaaa aaccgtcact taggaaaaga tgtcctttcg 60 ggcagccagg ctcagcatga ggaacagaag gaatgacact ctggacagca cccggaccct 120 gtactccagc gcgtctcgga gcacagactt gtcttacagt gaaagcgact tggtgaattt 180 tattcaagca aattttaaga aacgagaatg tgtcttcttt accaaagatt ccaaggccac 240 ggagaatgtg tgcaagtgtg gctatgccca gagccagcac atggaaggca cccagatcaa 300 ccaaagtgag aaatggaact acaagaaaca caccaaggaa tttcctaccg acgcctttgg 360 ggatattcag tttgagacac tggggaagaa agggaagtat atacgtctgt cctgcgacac 420 ggacgcggaa atcctttacg agctgctgac ccagcactgg cacctgaaaa cacccaacct 480 ggtcatttct gtgaccgggg gcgccaagaa cttcgccctg aagccgcgca tgcgcaagat 540 cttcagccgg ctcatctaca tcgcgcagtc caaaggtgct tggattctca cgggaggcac 600 ccattatggc ctgatgaagt acatcgggga ggtggtgaga gataacacca tcagcaggag 660 ttcagaggag aatattgtgg ccattggcat agcagcttgg ggcatggtct ccaaccggga 720 caccctcatc aggaattgcg atgctgaggt accggtggga caggaggagg tctgctaggt 780 cacatggaag aaagaccatg gcatgggcct gtggcctgaa ccctggggct ctgtgatgga 840 gccagccaga tcatggggaa gtctgccttt caaggagtgc ctttgggacc ttaaaggaat 900 tgaaaacaag gatgacgtac ctaattaact gctgggaaag agttaacaat gaatgttttg 960 ttcattaaaa tgtgttctca gcaatctc 988 <210> 36 <211> 3179 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 3232992CB1 <400> 36 gcggagcggc ggcgccggcg ccggggggcg cagcgagggg ctggcggtag cggttgctgc 60 ggggcgcggg gcgcgggcgg cgctggagtc tcggccgcgg gcgatgaggt gcagacgctg 120 tcgggcagcg taaggcgggc cccgaccgga ccccccggca cccccggcac ccccggctgc 180 gcagctactg caaaggggcc ccggcgctca gcagcccaaa ccggccagct tgggccgcgg 240 gcggggggca agccgccgcc atcctcagct tgggcaacgt gctcaactac ctggacaggt 300 acaccgtggc aggcgtcctt ctggacatcc agcagcactt tggggtcaag gaccgaggcg 360 ccggcctgct gcagtcagtg ttcatctgta gcttcatggt ggctgccccc atcttcggct 420 acctgggcga ccgcttcaac aggaaggtga ttctcagctg cggcattttc ttctggtcgg 480 ccgtcacctt ctccagctcc ttcattcccc agcagtactt ctggctgctg gtcctgtccc 540 gggggctggt gggcatcggg gaggccagct actccaccat cgcccccact atcattggcg 600 acctcttcac caagaacacg cgtacgctca tgctgtccgt cttctacttc gccatcccac 660 tgggcagtgg cctgggctac attactggct ccagcgtgaa gcaggcagcc ggagactggc 720 actgggcatt gcgggtgtcc cctgtcctgg gcatgatcac aggaacactc atcctcattc 780 tggtcccagc cactaaaagg ggtcatgccg accagctcgg ggaccagctc aaggcccgga 840 cctcatggct ccgagatatg aaggccctga ttcgaaaccg cagctacgtc ttctcctccc 900 tggccacgtc ggctgtctcc ttcgccacgg gggccctggg catgtggatc ccgctctacc 960 tgcaccgcgc ccaagttgtg cagaagacag cagagacgtg caacagcccg ccctgtgggg 1020 ccaaggacag cctcatcttt ggggccatca cctgctttac gggatttctg ggcgtggtca 1080 cgggggcagg agccacgcgc tggtgccgcc tgaagaccca gcgggccgac ccactggtgt 1140 gtgccgtggg catgctgggc tctgccatct tcatctgcct gatcttcgtg gctgccaaga 1200 gcagcatcgt aggagcctat atctgtatct tcgtcgggga gacgctgctg ~ttttctaact 1260 gggccatcac tgcagacatc ctcatgtacg tggtcatccc cacgcggcgc gccactgccg 1320 tggccttgca gagcttcacc tcccacctgc tgggggacgc cgggagcccc tacctcattg 1380 gctttatctc agacctgatc cgccagagca ctaaggactc cccgctctgg gagttcctga 1440 gcctgggcta cgcgctcatg ctctgccctt tcgtcgtggt cctgggcggc atgttcttcc 1500 tcgccactgc gctcttcttc gtcagcgacc gcgccagggc tgagcagcag gtgaaccagc 1560 tggcgatgcc gcccgcatct gtgaaagtct gaggtggtgc cattgggaca atgaagaacc 1620 cacactccca cctcgtctgg gaggtgtcct acagcgtccg ggaccggctg ggctgcccca 1680 aagctttctg tgtgatccac ggctaggcac ccaccctctc tggcccaggc ctgctgagtg 1740 gccctggcat caagaggagg ctgtgtcctc agttaccctg gaaggatgtg tgtgttggag 1800 ccacacggtt ggacaggttc ccagccctag gtttgggccg cagggcccct ggggccaagg 1860 aagaagacag ccccaagtgg gtgtccgggg agagcctggc ctgccaccag cttatgtgat 1920 cttgggcaag tccctgccct ccctggaacg aagggccagg gggctggact ttcccacaca 1980 acttgctggg caaagcacga tctgcagctt tgaagactca acagaccctg gaccatacgg 2040 agagcaggtg gcccaggcct cagggcggca gtcccggctt tgaggctcac gcgagggcct 2100 ggtatgcagg gaccactgct cagctgggcc tcggaccttg gggatattgg acgcaacctg 2160 gcaaatgaag ctgggcgccc aagtctctgg gtactccctg gaggacactg tctcactgtc 2220 tcgggttggc tcccagcctg gaggtcccag atggggactg ttctgacaag ctggcatcac 2280 caggggtgaa ggccctggct gcagctgtac accacctgtg cccccaggct caaggtctct 2340 ggcaggtgca caccagccca actctgcagg gcttctctcc ctgccaccac cccccaagcc 2400 aggaccccac tccttccccg aggctgagct gagccttttc caggggcagg gcccaggaga 2460 ccattcccag aatccatggg gcagtagcca gggctccggc tgctggagga agcagctatc 2520 cacaaagctt cctgccccag agctgaggct gaggccccgg gagaggcggc ccctacccaa 2580 acactggctg ctggcattcc accaagtgac cccaggggcc aggccttcga tcacccacct 2640 cccatccatg cacacaccag gatgcagctg ccaacttcac accagcccca acccgctttg 2700 ggggagctta gccccctgcg tcacccactg cctgcacttc tgctgcaatc aaggtggttc 2760 tggtgcgggg gtggggtggg gggtgaggcc ttgtggccaa tgggggaccc cccaagagcc 2820 agcttggaca atgctcttct tgccccttag ttactggctg gctgtggctt cagtggtgtg 2880 taagcaggtg gaatactcac ccaccaagct ctggggtacc ccgagggcct gacaagagga 2940 tggggtgggg gtggcatcct ccaaagacca gcctccaccc ccactccagc ctcagcgggg 3000 ccccagcgat gttttcttgt tgtacaagaa ccaggtccga gtgttgcctc ctcttccttc 3060 cggaagccaa actgctcctt tattttttag agctgctgat tgtgaatctc agagtcttaa 3120 gagagaagcc aaatatattc ctcttgtaaa tgaagaaata aacctattta aatcacaaa 3179 <210> 37 <211> 1986 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 3358383CB1 <400> 37 ggagtatctg agcaaattat ttcttacgtg actttagaga aaacggctac ctatctgacc 60 ccaaaacgac ttgaggaaac tgtttccacg gtcctgctgc aggggggaag cacagtcgtc 120 aagaagagag tggggtcagg atcaaaacac atttagtgtg acttagggaa agaaaacatt 180 ttccctcttt gaacctctct ggatacagtc attttgcctc tacttgagga tcaactgttc 240 aacctcaatg gcctttcagg acctcctggg tcacgctggt gacctgtgga gattccagat 300 ccttcagact gtttttctct caatctttgc tgttgctaca taccttcatt ttatgctgga 360 gaacttcact gcattcatac ctggccatcg ctgctgggtc cacatcctgg acaatgacac 420 tgtctctgac aatgacactg gggccctcag ccaagatgca ctcttgagaa tctccatccc 480 actggactca aacatgaggc cagagaagtg tcgtcgcttt gttcatcctc agtggcagct 540 ccttcacctg aatgggacct tccccaacac aagtgacgca gacatggagc cctgtgtgga 600 tggctgggtg tatgacagaa tctccttctc atccaccatc gtgactgagt gggatctggt 660 atgtgactct caatcactga cttcagtggc taaatttgta ttcatggctg gaatgatggt 720 gggaggcatc ctaggcggtc atttatcaga caggtttggg agaaggttcg tgctcagatg 780 gtgttacctc caggttgcca ttgttggcac ctgtgcagcc ttggctccca ccttcctcat 840 ttactgctca ctacgcttct tgtctgggat tgctgcaatg agcctcataa caaatactat 900 tatgttaata gccgagtggg caacacacag attccaggcc atgggaatta cattgggaat 960 gtgcccttct ggtattgcat ttatgaccct ggcaggcctg gcttttgcca ttcgagactg 1020 gcatatcctc cagctggtgg tgtctgtacc atactttgtg atctttctga cctcaagttg 1080 gctgctagag tctgctcggt ggctcattat caacaataaa ccagaggaag gcttaaagga 1140 acttagaaaa gctgcacaca ggagtggaat gaagaatgcc agagacaccc taaccctgga 1200 gattttgaaa tccaccatga aaaaagaact ggaggcagca caaaaaaaaa aaccttctct 1260 gtgtgaaatg ctccacatgc ccaacatatg taaaaggatc tccctcctgt cctttacgag 1320 atttgcaaac tttatggcct attttggcct taatctccat gtccagcatc tggggaacaa 1380 tgttttcctg ttgcagactc tctttggtgc agtcatcctc ctggccaact gtgttgcacc 1440 ttgggcactg aaatacatga cccgtcgagc aagccagatg cgtctcatgt acctactggc 1500 aatctgcttt atggccatca tatttgtgcc acaagaaatg cagacgctgc gtgaggtttt 1560 ggcaacactg ggcttaggag cgtcggctct gaccaatacc cttgcttttg cccatggaaa 1620 tgaagtaatt cccaccataa tcagggcaag agctatgggg atcaatgcaa cctttgctaa 1680 tatagcagga gccctggctc ccctcatgat gatcctaagt gtgtattctc cacccctgcc 1740 ctggatcatc tatggagtct tccccttcat ctctggcttt gctttcctcc tccttcctga 1800 aaccaggaac aagcctctgt ttgacaccat ccaggatgag aaaaatgaga gaaaagaccc 1860 cagagaacca aagcaagagg atccgagagt ggaagtgacg cagttttaag gaattccagg 1920 agctgactgc cgatcaatga gccagatgaa gggaacaatc aggactattc ctagacacta 1980 gcaaat 1986 <210> 38 <211> 3294 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 4250091CB1 <400> 38 tgtaagacag gaaagggatc tatttgatgt ctatcttcag atatattggc agttttcctt 60 aagctattta gttcctcatc tgttgctttt tcattttgta tactgcaagt tcccaggcaa 120 ctcgaatttg caaacacagc catggataca ctatttacct tacagtagtt tcctgggaat 180 ctaagtctgg tttttgttat tcttccctcc cctccactgc ataatcatgt ataactagca 240 acatttatgg ttataggttg atttcctaag tgtggctgat ggtagcctct agtttgaagt 300 gagggaagaa tgagtagtca ggaactggtc actttgaatg tgggagggaa gatattcacg 360 acaaggtttt ctacgataaa gcagtttcct gcttctcgtt tggcacgcat gttagatggc 420 agagaccaag aattcaagat ggttggtggc cagatttttg tagacagaga tggtgatttg 480 tttagtttca tcttagattt tttgagaact caccagcttt tattacccac tgaattttca 540 gactatctta ggcttcagag agaggctctt ttctatgaac ttcgttctct agttgatctc 600 ttaaacccat acctgctaca gccaagacct gctcttgtgg aggtacattt cctaagccgg 660 aacactcaag cttttttcag ggtgtttggc tcttgcagca aaacaattga gatgctaaca 720 gggaggatta cagtgtttac agaacaacct tcggcgccga cctggaatgg taactttttc 780 cctcctcaga tgaccttact tccactgcct ccacaaagac cttcttacca tgacctggtt 840 ttccagtgtg gttctgacag cactactgat aaccaaactg gagtcaggta ttttgtactt 900 tgcagtattt ctcttgtata ccagtttgtg atgttttctc taaaaacttg aagttcctca 960 ggcctgtaac ttctggaaaa gatgattatt caaaataatg ttttggggta accagtggag 1020 ttgggtagaa tgaccaaata attattttcc aaactgggat actttttaga gtgaaagggg 1080 ctattattag gtgggacaaa aggaataaat gaagactgcc cagaaaaaac tgagactatg 1140 gacattcaaa tcatgggaga aaataatttt gtagattatg ttccattgct aatgaatttg 1200 acttagaaaa gaattgcctt atttttaaga gattgtttca gtggttcaca taaaggctcg 1260 ctcactggtt tctcttgagt tccttacaca ctatataagt tgttctttca gttttatgat 1320 tcaactactg tttttccttc agctgacttt atttttaaac acccttaaag acagatatat 1380 ctcatggcaa atttggtatc ctgttacagc cttggctctt aaacaactca aaatattggg 1440 ataggctgtc agtatgttaa ggatagttgc tcctgagtca attcttcact tactccctct 1500 gttgttcttg gctggatcct aacgctgatt tccactctgc tgtcacaaac atttttcccc 1560 ccgtaaaatg tcttaatgct gtcctaccat tattttacca actgtgaaag ctggctttaa 1620 tttttaggag gaaaagaaaa gcctgcatgt gttctttatt ggtatcattt aaaatatact 1680 tttttttttt ttttggtaaa ggtaggcgta ttttaagata ttttcttaac ttgagcagta 1740 gccaacagga aggataccag tgtctctctc tcttagcgac acactccttg gtcttgctta 1800 ccaactggag gacactaggt agaataaccg agtatgacaa ttcttaattg tttacatttt 1860 ataacttcct gtccttcaaa agagtttgaa atgtcatttt gggaaaagag agccagtcaa 1920 gctagtaggc tgattgtgaa gaaaatctaa taccttatct ttatctcaaa cctctgtaca 1980 actttatttt cattgatggg atactttaac aaaaatgaaa ttttttttgg tttttaaaat 2040 atgagtgatt atgacctctt tggggatcat gcttcaaaaa gtcagaaacc tagagacaaa 2100 actgtcattg atttttaaga agaaacacac taggtcaaaa gaagatgtcc tggaaatacg 2160 aagtactctt taaaaaccat gcatttggag aaagtaattg tttccttgaa aaacatgatt 2220 aaaaactaaa actgggatgt tcctgtgtgt acacagtgcc aaatggtttt ccctttttat 2280 gttgtgtttt agaaacagca cgaaagtttt ttccatttta aagtgagaaa acattatatt 2340 tagacttcca taattccaaa atcagaagct atttttaaaa ttagcatttt cttgcatcac 2400 caaatggtat tcaattgttt gaagctcaaa atttttacca ttccataaat gtttgtgaat 2460 ttttagacag tgccaattta aaagtagaga tagccaatct gaatacggtg aaattatggg 2520 gatctctggt gattgggatg aaaactctgg ccttaaaagg tccactttta gtatataatt 2580 gcctaattag caatcatttt tattttttgc tcactccctg gtctgaatct atctgtctat 2640 tcagatattt tttggtaggt ttggaaaatg gagaagtgag cctaattggt gcctaattgt 2700 ctggtgtatc attcacttta ttcagtttgt tctatcaata tgatttaccc ctcaaggtta 2760 acctagcagg ttgctcagtt attatctctc aaggtcacag tactagaaat acttggcttg 2820 catctttcag atgccattca tgttatcaag ctcaaattat agttggtcac aggattctaa 2880 agtctttatt tgacttctcc tttttgaact ggctcaaatg gaaaagtgta gttgctttta 2940 aatgttaaaa ataagtttaa actttatatt tcccattggt ttcccctatt ttgtcctttc 3000 tttgtgtgct tgaaatattt tatttttcag tttgtcctca tagggaatca agtattttag 3060 ctaggtgatg tcttgcaagt acgttccact ttgttacaat ctactatctg tatatactat 3120 ttgtatctta attcttttat gagatgttct gtaacatttt tctcactttg acaaatgttt 3180 ttagactgta cagtcaagat ctggcgcttg ggggtaagtg gaatgatttg ctaatattga 3240 gaatctgttg tatcaaacat aataaacttt ttttgagatg tgaaaaaaaa aaaa 3294 <210> 39 <211> 2043 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 70064803CB1 <400> 39 gcaacatggc ggctgccgtg gtgcagcgcc cgggctgagc gacagcaagt gcagcgggct 60 cctaccccgg gtgaggggtg gcctccgcgt gggatcgtgc cctcttcagc ccgctcctgt 120 ccccgacatc acgtgtattc cgcacgtccc ctccgcgctg tgtgtctact gagacgggga 180 ggcgtgacag ggcccgggtc ccttctcagt ggtgctctgt gcttcagggc aagctccccg 240 tctccgggcg cacttccctc gcctgtgttc ggtccatcct cctttctcca gcctcctccc 300 ctcgcaggtg ggatcgtcgg tgggaccgga gcgcgggcgg gcgcggcccc ccgggaccat 360 ggccgggtcc gacaccgcgc ccttcctcag ccaggcggat gacccggacg acgggccagt 420 gcctggcacc ccggggttgc cagggtccac ggggaacccg aagtccgagg agcccgaggt 480 cccggaccag gaggggctgc agcgcatcac cggcctgtct cccggccgtt cggctctcat 540 agtggcggtg ctgtgctaca tcaatctcct gaactacatg gaccgcttca ccgtggctgg 600 cgtccttccc gacatcgagc agttcttcaa catcggggac agtagctctg ggctcatcca 660 gaccgtgttc atctccagtt acatggtgtt ggcacctgtg tttggctacc tgggtgacag 720 gtacaatcgg aagtatctca tgtgcggggg cattgccttc tggtccctgg tgacactggg 780 gtcatccttc atccccggag agcatttctg gctgctcctc ctgacccggg gcctggtggg 840 ggtcggggag gccagttatt ccaccatcgc gcccactctc attgccgacc tctttgtggc 900 cgaccagcgg agccggatgc tcagcatctt ctactttgcc attccggtgg gcagtggtct 960 gggctacatt gcaggctcca aagtgaagga tatggctgga gactggcact gggctctgag 1020 ggtgacaccg ggtctaggag tggtggccgt tctgctgctg ttcctggtag tgcgggagcc 1080 gccaagggga gccgtggagc gccactcaga tttgccaccc ctgaacccca cctcgtggtg 1140 ggcagatctg agggctctgg caagaaatct catctttgga ctcatcacct gcctgaccgg 1200 agtcctgggt gtgggcctgg gtgtggagat cagccgccgg ctccgccact ccaacccccg 1260 ggctgatccc ctggtctgtg ccactggcct cctgggctct gcacccttcc tcttcctgtc 1320 ccttgcctgc gcccgtggta gcatcgtggc cacttatatt ttcatcttca ttggagagac 1380 cctcctgtcc atgaactggg ccatcgtggc cgacattctg ctgtacgtgg tgatccctac 1440 ccgacgctcc accgccgagg ccttccagat cgtgctgtcc cacctgctgg gtgatgctgg 1500 gagcccctac ctcattggcc tgatctctga ccgcctgcgc cggaactggc ccccctcctt 1560 cttgtccgag ttccgggctc tgcagttctc gctcatgctc tgcgcgtttg ttggggcact 1620 gggcggcgca gccttcctgg gcaccgccat cttcattgag gccgaccgcc ggcgggcaca 1680 gctgcacgtg cagggcctgc tgcacgaagc agggtccaca gacgaccgga ttgtggtgcc 1740 ccagcggggc cgctccaccc gcgtgcccgt ggccagtgtg ctcatctgag aggctgccgc 1800 tcacctacct gcacatctgc cacagctggc cctgggccca ccccacgaag ggcctgggcc 1860 taaccccttg gcctggccca gcttccagag ggaccctggg ccgtgtgcca gctcccagac 1920 actacatggg tagctcaggg gaggaggtgg gggtccagga gggggatccc tctccacagg 1980 ggcagcccca agggctcggt gctatttgta acggaataaa atttgtagcc agacaaaaaa 2040 aaa 2043 <210> 40 <211> 1915 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 70356768CB1 <400> 40 caccactggg cgctgcgcgc tgcccttccc tccgcgcaca ggctgccggc tcaccgcttg 60 ctaatggcag ccggggtctc cctgggacag caagacctcc gctcaggccc ctctttcgaa 120 tgctccacgc cctcctgcga tctagaatga ttcagggcag gatcctgctc ctgaccatct 180 gcgctgccgg cattggtggg acttttcagt ttggctataa cctctctatc atcaatgccc 240 cgaccttgca cattcaggaa ttcaccaatg agacatggca ggcgcgtact ggagagccac 300 tgcccgatca cctagtcctg cttatgtggt ccctcatcgt gtctctgtat cccctgggag 360 gcctctttgg agcactgctt gcaggtccct tggccatcac gctgggaagg aagaagtccc 420 tcctggtgaa taacatcttt gtggtgtcag cagcaatcct gtttggattc agccgcaaag 480 caggctcctt tgagatgatc atgctgggaa gactgctcgt gggagtcaat gcaggtgtga 540 gcatgaacat ccagcccatg tacctggggg agagcgcccc taaggagctc cgaggagctg 600 tggccatgag ctcagccatc tttacggctc tggggatcgt gatgggacag gtggtcggac 660 tcagggagct cctaggtggc cctcaggcct ggcccctgct gctggccagc tgcctggtgc 720 ccggggcgct ccagctcgcc tccctgcctc tgctccctga aagcccgcgc tacctcctca 780 ttgactgtgg agacaccgag gcctgcctgg cagagacggg ttctcgcttg tccaggctgg 840 agtgctgtgg ctgttcatag gcatgacccc attgttgatc agcacggaag ttttcttctt 900 ttttgttttt gtttttttgg ttttgtttgg gacggggtct cactctgtcg cccaggctgg 960 agtggtgtga tctcggctcg ctgcagcctc cacctcccgg gcccaatcgg ttctcccgcc 1020 tcagcctcct gggtggctgg gactgctggc ccgtgccacc acgcttggct aatttttttt 1080 tattattgta ttttttgtaa agatggagtt tcacctcttt gcctgggcag gtctcaaact 1140 cctgagatca aatgatcctc cccccttggc ctcccaaagt gcgtggatta taggcatgag 1200 ccattgtatc tggctagcat gggagttttg aactgtccca tttccaacct gggccagtgc 1260 attcctcctt aggcagcctg gtggtccctg ctcctgggat gtcactatat tgatgctgaa 1320 cttagtgcag acacctgatc tgcctagcgt actgcaaccc agagctcctg ggcccaggcg 1380 atcctcctgt ctcagcctcc tgagtagctg ggactctagg cacacaccac tatgcgtggc 1440 tctccatgct tcttgggtct accctctgag atgtttttcc ttttctttca ccttccttga 1500 ttccttctga agagggcgtt gcacaatgtg ctgcttttga tggttgagca aatttctcag 1560 cctccttcct gcctatagag agttggggca ggctgggcgc cagctcacgc ctgtaatccc 1620 agggaggctg aggcgggcag atcacgaggt caggacatca agaccggcct ggccgacatg 1680 gtgggacccc atctctacta acaatacaaa aattggctgg gtatggtggc acgtgcctgt 1740 ggtcccggct gctggggagg ctgaggcggg agagttgctt gggcccggga ggcggaggtt 1800 gcagtggcgg gagaattgct tggggcccgg gaggcggagg ttgcggtgag ccgagattgt 1860 gccagtgcac actgcactcc agcctggtga cagagtgaga ctccgtcttc aaaaa 1915 <210> 41 <211> 1809 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 5674114CB1 <400> 41 atgggcctgg ccagggccct acgccgcctc agcggcgccc tggattcggg agacagccgg 60 gcgggcgatg aagaggaggc cgggcccggg ttgtgccgca acgggtgggc gccggcaccg 120 gtgcagtcac ccgtgggccg gcgccgcggt cgcttcgtca agaaagacgg gcactgcaac 180 gtgcgtttcg taaacctggg tggccagggc gcgcgctacc tgagcgacct gttcaccaca 240 tgcgtggacg tgcgctggcg ctggatgtgc ctgctcttct cctgctcctt cctcgcctcc 300 tggctgctct tcggcctggc cttctggctc attgcctcgc tgcacggcga cctggccgcc 360 ccgccaccgc ccgcgccctg cttctcacac gtggccagct tcctggccgc cttcctcttc 420 gcgctggaga cgcagacgtc catcggctac ggcgtgcgca gcgtcaccga ggagtgcccg 480 gccgctgtgg ccgccgtggt gctgcagtgc attgccggct gcgtgctcga cgccttcgtc 540 gtgggtgctg tcatggccaa gatggccaaa cccaagaagc gcaacgagac gctggtcttc 600 agcgagaacg ccgtcgtggc gctgcgcgac caccgcctct gcctcatgtg gcgcgtcggc 660 aacctgcgcc gcagccacct ggtcgaggcc cacgtgcgtg cccagctgct gcagccccgt 720 gtgaccccag agggtgagta catcccgctg gaccaccagg atgtggatgt gggctttgat 780 ggaggcaccg atcgtatctt cctcgtgtcc cccatcacca tcgtccatga gatcgactct 840 gccagtcctc tgtatgagct aggacgtgcc gagctggcca gggctgactt tgagctggtg 900 gtcattctcg aggggatggt tgaggccaca gccatgacca cacagtgtcg ctcgtcctac 960 ctccctggtg aactgctctg gggccatcgt tttgagccag ttctcttcca gcgtggctcc 1020 cagtatgagg tcgactatcg ccacttccat cgcacttatg aggtcccagg gacaccggtc 1080 tgcagtgcta aggagctgga tgaacgggca gagcaggctt cccacagcct caagtctagt 1140 ttccccggct ctctgactgc attttgttat gagaatgaac ttgctctgag ctgctgccag 1200 gaggaagatg aggacgatga gactgaggaa gggaatgggg tggaaacaga agatggggct 1260 gctagccccc gagttctcac accaaccctg gcgctgaccc tgcctccatg atgcaaactg 2320 atgtcccctt ccccgtgtat gcccccttcc ccaaggtagc aagatggagg gatggggctc 1380 tctcctggga tgggggcagg tgttcctgaa taccgacagg cctgctgggt aaatgactag 1440 gtggtaaggt tctgccatgc ctggtgaccc accatggaca tactggacct taattcctct 1500 gcttctgtgc tccctcctga gaacccttta tgagcctgat tcctcagtct caccagaatt 1560 ctggatcacc caagaggaaa agactggcag ttctagattc ctctatatgg ggagacctgg 1620 attgttgacc agggtgagaa gccaatggta tagactgcct ctggggaagc aagttggcag 1680 ttcttgaaca gcatcagata tcaagagttt gtaggtctgg attcacctaa gattcaaggg 1740 agtgttgctt ctcaactcag ccaactgagt agcaaatcat ttgttctaga ccacctaagg 2800 agggaaggt 1809 <210> 42 <211> 1730 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1254635CB1 <400> 42 ctttggccta ttataccatg gatgctaaaa atggttctaa ctgaaaaccc aaaccaagaa 60 atagcaacaa gtctagaatt cttactacta caaaactcac ctggatccct aagggcacag 120 caaagaatga gctattacgg cagcagctat catattatca atgcggacgc aaaataccca 180 ggctacccgc cagagcacat tatagctgag aagagaagag caagaagacg attacttcac 240 aaagatggca gctgtaatgt ctacttcaag cacatttttg gagaatgggg aagctatgtg 300 gttgacatct tcaccactct tgtggacacc aagtggcgcc atatgtttgt gatattttct 360 ttatcttata ttctctcgtg gttgatattt ggctctgtct tttggctcat agcctttcat 420 catggcgatc tattaaatga tccagacatc acaccttgtg ttgacaacgt ccattctttc 480 acaggggcct ttttgttctc cctagagacc caaaccacca taggatatgg ttatcgctgt 540 gttactgaag aatgttctgt ggccgtgctc atggtgatcc tccagtccat cttaagttgc 600 atcataaata cctttatcat tggagctgcc ttggccaaaa tggcaactgc tcgaaagaga 660 gcccaaacca ttcgtttcag ctactttgca cttataggta tgagagatgg gaagctttgc 720 ctcatgtggc gcattggtga ttttcggcca aaccacgtgg tagaaggaac agttagagcc 780 caacttctcc gctatacaga agacagtgaa gggaggatga cgatggcatt taaagacctc 840 aaattagtca acgaccaaat catcctggtc accccggtaa ctattgtcca tgaaattgac 900 catgagagcc ctctgtatgc ccttgaccgc aaagcagtag ccaaagataa ctttgagatt 960 ttggtgacat ttatctatac tggtgattcc actggaacat ctcaccaatc tagaagctcc 1020 tatgttcccc gagaaattct ctggggccat aggtttaatg atgtcttgga agttaagagg 1080 aagtattaca aagtgaactg cttacagttt gaaggaagtg tggaagtata tgcccccttt 1140 tgcagtgcca agcaattgga ctggaaagac cagcagctcc acatagaaaa agcaccacca 1200 gttcgagaat cctgcacgtc ggacaccaag gcgagacgaa ggtcatttag tgcagttgcc 1260 attgtcagca gctgtgaaaa ccctgaggag accaccactt ccgccacaca tgaatatagg 1320 gaaacacctt atcagaaagc tctcctgact ttaaacagaa tctctgtaga atcccaaatg 1380 tagtcctaaa ttgcaattat gagggctacc actgaatcat tttatctttc agccaatcaa 1440 gtcgttgtaa acgtggcttt tttgaaagtg ttatggctat gttttatgat gatgctgggt 1500 aagtagagta agttaaactt ggtaaaagat aatctaaaaa ttccatagtt ctcagttatt 1560 aaaatttttc ttgttcgcca attttgtatt aagaatgcta ttaagcctaa ttgattaaaa 1620 tttatctttt ttattatctt acatgcttgt atcttcagtt ggaggtgtag tattcaaaaa 1680 cggggaatga aggcaggaag gaggctggaa taaataaaaa taaaatgatt 1730 <210> 43 <211> 1147 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1670595CB1 <400> 43 gcagctgtct tttccggccc ccgtgcactc tccgcccgag gcggagcccc cggctcgcgg 60 ggatcgcccc cgagcgctgc gtcctgcggg tgggtcacct aacccatttg tggcttcctc 220 tacctgtgct cagccatggc cagcgagagc tcacctctgc tggcctaccg gctcctgggg 180 gaggaggggg ttgccctccc tgccaatggg gccgggggtc ctggaggggc gtctgcccgg 240 aagctgtcca ccttcctggg tgtggtggtg cccactgtcc tgtccatgtt cagcatagtt 300 gtttttctga ggattgggtt cgtggtgggt catgctgggc tactgcaggc cctggccatg 360 ctgctggttg cctacttcat cctggcactc accgtcctct ctgtctgtgc catcgccacc 420 aatggagccg tgcagggggg cggagcctac tgtatcctcc aacatcgatg gactgggatg 480 ccacagggcc cagtgggctc cgggtcctgc cccagggcta cggcttggaa cctgctgtat 540.
ggctccctgc tgctgggcct tgtgggtggg gtctgcacct tgggagccgg cctctatgcc 600;
cgggcctcat tcctcacatt cctgctggtc tctggctccc tggcctctgt gctcatcagt 660.
tttgtggctg tggggccgag ggacatccgc ttgactccta ggcctggccc caatggctcc 720 tccctgccgc cccggtttgg ccacttcacc ggcttcaaca gcagtaccct gaaggacaac 780 ttgggcgctg gctatgctga ggactacacc acgggagccg tgatgaattt tgccagcgtc 840 tttgctgtcc tctttaacgg caggcatcat ggctggggcc aacatgtcag gggagctgaa 900 ggaccccagc cgggcgatcc ctctgggcac gatcgtcgcc gtcgcctaca ccttcttcgt 960 ctatgccctg cttttctttc tctccagcct cccttcactg gtgccttgat gctaggggcc 1020 aggcctcctc tgtgactctg ggctacctca gtttccccat tttggccaga ctcaccggcc 1080 caccggggtg gtgatgtttt cgttctgttt tatttttcta actctgcatg accatgaata 1140 aaagacc 1147 <210> 44 <211> 2745 <212 > DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1859560CB1 <400> 44 cggcgacgcc agggacccca cgcatcccga gtgaagcaac tagaactcca gggctgtgaa 60 agccacaggt gggggctgag cgaggcgtgg cctcaggagc ggaggacccc ccactctccc 120 tcgagcgccg cagtccaccg tagcgggtgg agcccgcctt ggtgcgcagt tggaaaacct 180 cggagccccg ctggatctcc tggctgccac ccgcaccccc cgccagccta cgccccaccg 240 tagagatgcc ttcttcggtg acggcgctgg gtcaggccag gtcctctggc cccgggatgg 300 ccccgagcgc ctgctgctgc tcccctgcgg ccctgcagag gaggctgccc atcctggcgt 360 ggctgcccag etactccctg cagtggctga agatggattt cgtcgccggc ctctcagttg 420 gcctcactgc cattccccag gcgctggcct atgctgaagt ggctggactc ccgccccagt 480 atggcctcta ctctgccttc atgggctgct tcgtgtattt cttcctgggc acctcccggg 540 atgtgactct gggccccacc gccattatgt ccctcctggt ctccttctac accttccatg 600 agcccgccta cgctgtgctg ctggccttcc tgtccggctg catccagctg gccatggggg 660 tcctgcgttt ggggttcctg ctggacttca tttcctaccc cgtcattaaa ggcttcacct 720 ctgctgctgc cgtcaccatc ggctttggac agatcaagaa cctgctggga ctacagaaca 780 tccccaggcc gttcttcctg caggtgtacc acaccttcct caggattgca gagaccaggg 840 taggtgacgc cgtcctgggg ctggtctgca tgctgctgct gctggtgctg aagctgatgc 900 gggaccacgt gcctcccgtc caccccgaga tgccccctgg tgtgcggctc agccgtgggc 960 tggtctgggc tgccacgaca gctcgcaacg ccctggtggt ctccttcgca gccctggttg 1020 cgtactcctt cgaggtgact ggataccagc ctttcatcct aacaggggag acagctgagg 1080 ggctccctcc agtccggatc ccgcccttct cagtgaccac agccaacggg acgatctcct 1140 tcaccgagat ggtgcaggac atgggagccg ggctggccgt ggtgcccctg atgggcctcc 1200 tggagagcat tgcggtggcc aaagccttcg catctcagaa taattaccgc atcgatgcca 1260 accaggagct gctggccatc ggtctcacca acatgttggg ctccctcgtc tcctcctacc 1320 cggtcacagg cagctttgga cggacagccg tgaacgctca gtcgggggtg tgcaccccgg 1380 cggggggcct ggtgacggga gtgctggtgc tgctgtctct ggactacctg acctcactgt 1440 tctactacat ccccaagtct gccctggctg ccgtcatcat catggccgtg gccccgctgt 1500 tcgacaccaa gatcttcagg acgctctggc gtgttaagag gctggacctg ctgcccctgt 1560 gcgtgacctt cctgctgtgc ttctgggagg tgcagtacgg catcctggcc ggggccctgg 1620 tgtctctgct catgctcctg cactctgcag ccaggcctga gaccaaggtg tcagaggggc 1680 cggttctggt cctgcagccg gccagcggcc tgtccttccc tgccatggag gctctgcggg 1740 aggagatcct aagccgggcc ctggaagtgt ccccgccacg ctgcctggtc ctggagtgca 1800 cccatgtctg cagcatcgac tacactgtgg tgctgggact cggcgagctc ctccaggact 1860 tccagaagca gggcgtcgcc ctggcctttg tgggcctgca ggtccccgtt ctccgtgtcc 1920 tgctgtccgc tgacctgaag gggttccagt acttctctac cctggaagaa gcagagaagc 1980 acctgaggca ggagccaggg acccagccct acaacatcag agaagactcc attctggacc 2040 aaaaggttgc cctgctcaag gcataatggg gccacccgtg ggcatccaca gtttgcaggg 2100 tgttccggaa ggttcttgtc actgtgattg gatgctggat gccgcctgat agacatgctg 2160 gcctggctga gaaacccctg agcaggtaac ccagggaaga gaaggaagcc aggcctggag 2220 gtccacggca gtgggagtgg ggctcactgg cttcctgtgg gatgactgga aaatgacctc 2280 gctgctgttc cctggcatga ccctctttgg aagagtggtt tggagagagc cttctagaat 2340 gacagactgt gcgaggaagc aggggcaggg gtttccagcc cgggctgtgc gaggcatcct 2400 ggggctggca gcaccttccc ggctcaccag tgccacctgc gggggaggga cggggcaggc 2460 aggagtctgg gaggcgggtc cgctcctctt gtctgcggca tctgtgctct ccgagagaaa 2520 accaaggtgt gtcaaatgac gtcaagtctc tatttaaaaa taattttgtg ttttctaaat 2580 ggaaaaagtg atagctttgg tgattttgta aaagtcataa atgcttattg taaaaaatac 2640 aggaaaccac ccctcaccct gtccacttgg gtgatcattc cagacccctc cccaaacatg 2700 catatgtacc tgtccgtcag tgtgtggatg tatgtttaca gttct ' 2745 <210> 45 <211> 3204 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 5530164CB1 <400> 45 cgacctctgg agctactgcg cctgcaagcc cagcctctct gcgccgcagg ctgcggggcc 60 agctggcgcc gcacaaatac ggggcgggac acggggcggg acacgggccg gtcccggggg 120 agggcctgag ccgcacagcc cgcccagggg tggtgcgtgt aaacgggcgt ctggatcccc 180 gaatggttgc gtgtttccgt gtgtgggtcc gggggaggcc cacgaacgcc agcgaaaccg 240 ctgacaccac cgcccaacta tgaactcatc aggcgcctga agaccgacac gccgaacatg 300 cgccgcgcgc actcgcgcac gagtgagatc atcgcgcccc ggtcgtgagt gcgctcacac 360 gcagcctgag actcgacggg agggggtcac gtggaagtat ctgagagagg cgtacttggc 420 cactaggaaa gcacctcccc ctttccaaaa atgctccgga agtgccttcg ccctccgtaa 480 agatggccgg ggcagtcggc acgagggagg cggggatgcg cctgcgcaac aagttcggcg 540 gggaagatgg cggatgacaa ggattctctg cctaagctta aggacctggc atttctcaag 600 aaccagctgg aaagcctgca gcggcgtgta gaagacgaag tcaacagtgg agtgggccag 660 gatggctcgc tgttgtcctc cccgttcctc aagggattcc tggctggcta tgtggtggcc 720 aaactgaggg catcagcagt attgggcttt gctgtgggca cctgcactgg catctatgcg 780 gctcaggcat atgctgtgcc caacgtggag aagacattaa gggactattt gcagttgcta 840 cgcaaggggc ccgactagct ctaggtgcca tggaagaggc aggatgagca gctcagcctt 900 caggtggaga cactttatct ggattcccca gctgtcatcc atttgctatc tccaactttc 960 ctgccacctt catccttgcc tcccttcctg cagattgtgg acagtagttc ctcagcctgc 1020 accctggatt ccttcttccc cttcctagct ccatgggact cgccccaaga ctgtggcttc 1080 aaggaccacc agccccttac tcttcaagcc ctgactgtgg agttggtaga tgcctctgat 2140 cctcagtatt ctctctggca atgttccacg gcttctcctt cctgggagct ggctccataa 1200 cttgattttc cccaaacgtg ttgcaatccc tgctgcccct tagccaccca gggtcttgtg 1260 tgggtatgag tgtagaggat gggggtatgc caggcctggg ccgtcccagg caggcccgct 1320 ggaccctgat gctactccta tccactgcca tgtacggtgc ccatgcccca ttgctggcac 1380 tgtgccatgt ggacggccga gtgcccttcc ggccctcctc agccgtgctg ctgactgagc 1440 tgaccaagct actgttatgc gccttctccc ttctggtagg ctggcaagca tggccccagg 1500 ggcccccacc ctggcgccag gctgctccct tcgcactatc agccctgctc tatggcgcta 1560 acaacaacct ggtgatctat cttcagcgtt acatggaccc cagcacctac caggtgctga 1620 gtaatctcaa gattggaagc acagctgtgc tctactgcct ctgcctccgg caccgcctct 1680 ctgtgcgtca ggggttagcg ctgctgctgc tgatggctgc gggagcctgc tatgcagcag 1740 ggggccttca agttcccggg aacacccttc ccagtccccc tccagcagct gctgccagcc 1800 ccatgcccct gcatatcact ccgctaggcc tgctgctcct cattctgtac tgcctcatct 1860 caggcttgtc gtcagtgtac acagagctgc tcatgaagcg acagcggctg cccctggcac 1920 ttcagaacct cttcctctac acttttggtg tgcttctgaa tctaggtctg catgctggcg 1980 gcggctctgg cccaggcctc ctggaaggtt tctcaggatg ggcagcactc gtggtgctga 2040 gccaggcact aaatggactg ctcatgtctg ctgtcatgaa gcatggcagc agcatcacac 2100 gcctctttgt ggtgtcctgc tcgctggtgg tcaacgccgt gctctcagca gtcctgctac 2160 ggctgcagct cacagccgcc ttcttcctgg ccacattgct cattggcctg gccatgcgcc 2220 tgtactatgg cagccgctag tccctgacaa cttccaccct gattccggac cctgtagatt 2280 gggcgccacc accagatccc cctcccaggc cttcctccct ctcccatcag cagccctgta 2340 acaagtgcct tgtgagaaaa gctggagaag tgagggcagc caggttattc tctggaggtt 2400 ggtggatgaa ggggtacccc taggagatgt gaagtgtggg tttggttaag gaaatgctta 2460 ccatccccca cccccaacca agttcttcca gactaaagaa ttaaggtaac atcaatacct 2520 aggcctgaga aataacccca tccttgttgg gcagctccct gctttgtcct gcatgaacag 2580 agttgatgaa agtggggtgt gggcaacaag tggctttcct tgcctacttt agtcacccag 2640 cagagccact ggagctggct agtccagccc agccatggtg catgactctt ccataaggga 2700 tcctcaccct tccactttca tgcaagaagg cccagttgcc acagattata caaccattac 2760 ccaaaccact ctgacagtct cctccagttc cagcaatgcc tagagacatg ctccctgccc 2820 tctccacagt gctgctcccc acacctagcc tttgttctgg aaaccccaga gagggctggg 2880 cttgactcat ctcagggaat gtagcccctg ggccctggct taagccgaca ctcctgacct 2940 ctctgttcac cctgagggct gtcttgaagc ccgctaccca ctctgaggct cctaggaggt 3000 accatgcttc ccactctggg gcctgcccct gcctagcagt ctcccagctc ccaacagcct 3060 ggggaagctc tgcacagagt gacctgagac caggtacagg aaacctgtag ctcaatcagt 3120 gtctctttaa ctgcataagc aataagatct taataaagtc ttctaggctg tagggtggtt 3180 cctacaacca cagccaaaaa aaaa 3204 <210> 46 <211> 2763 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 139115CB1 <400> 46 tgcatttgct atgactttga ccggtccact gacaacgcaa tatgtttatc ggagaatatg 60 ggaagaaact ggcaactaca ctttttcatc tgatagcaat atttctgagt gtgaaaaaaa 120 caaaagcagc ccaatttttg cattccagga ggaagttcag aaaaaagtgt cacgttttaa 180 tctgcagatg gacataagtg gattaattcc tggtctagtg tctacattca tacttttgtc 240 tattagtgat cactacggac gaaaattccc tatgattttg tcttccgttg gtgctcttgc 300 aaccagcgtt tggctctgtt tgctttgcta ttttgccttt ccattccagc ttttgattgc 360 atctaccttc attggtgcat tttgtggcaa ttataccaca ttttggggag cttgctttgc 420 ctatatagtt gatcagtgta aagaacacaa acaaaaaaca attcgaatag ctatcattga 480 ctttctactt ggacttgtta ctggactaac aggactgtca tctggctatt ttattagaga 540 gctaggtttt gagtggtcgt ttctaattat tgctgtgtct cttgctgtta atttgatcta 600 tattttattt tttctcggag atccagtgaa agagtgttca tctcagaatg ttactatgtc 660 atgtagtgaa ggcttcaaaa acctatttta ccgaacttac atgcttttta agaatgcttc 720 tggtaagaga cgatttttgc tctgtttgtt actttttaca gtaatcactt atttttttgt 780 ggtaattggc attgccccaa tttttatcct ttatgaattg gattcaccac tctgctggaa 840 tgaagttttt ataggttatg gatcagcttt gggtagtgcc tcttttttga ctagtttcct 900 aggaatatgg cttttttctt attgtatgga agatattcat atggccttca ttgggatttt 960 taccacgatg acaggaatgg ctatgaccgc gtttgccagt acaacactga tgatgttttt 1020 agccagggtg ccgttccttt tcactattgt gccattctct gttctacggt ccatgttgtc 1080 aaaagtggtt cgttcgactg aacaaggtac cctgtttgct tgtattgctt tcttagaaac 1140 acttggagga gtcactgcag tttctacttt taatggaatt tactcagcca ctgttgcttg 1200 gtaccctggc ttcactttcc tgctgtctgc tggtctgtta ctacttccag ccatcagtct 1260 atgtgttgtc aagtgtacca gctggaatga gggaagctat gaacttctta tacaagaaga 1320 atccagtgaa gatgcttcag acaggtgact gtgatttaaa caaacaaaaa aaatctatga 1380 atgcacatat catataccat gacttctgaa gactataaat gaattccaca atcagtgctt 1440 cactgagaac caattttacc tatcttttct tctaaactga acagtcagag agacagctcc 1500 tggctttagc ttcttgtggt accacgcact ttgagcactt tgtgcgtatc atgcaatata 1560 cttgcaatac acagaacaaa tttcaaatac gcctcacttt tagacttaga agagaaacat 1620 taaaacttaa gggtgtaagg agggatcaag aaacttgata aggtcaaaag caataatctc 1680 tctgacatat tccaggctct tacactgaga ccaaagagaa atctttacct cagtttcttc 1740 atcagcagaa tgggtttctg gcctctctca gggataattt tgaaggcata atgaaaatta 1800 tgatgaatca ctcattggta ggaaaataat gatataagtt tcaaatatgt atgattttac 1860 ctatacttgg taatgctttg ttttatagag cctgttaagc tgctattgat agtcggagct 1920 tatatactgt gacttctgaa gactatacat gaattccaca atcagtgctt tgttgataca 1980 aaatccttaa aagggaggca ctttaaagaa tatgtatttt tcacttttct taatatgttt 2040 catcggtgac aggcatgata atatttctat atgtaatggg taattgggaa aaaatagatg 2100 ataaataaaa ttgctctaaa gaagttaaaa aactgaatga acagctaata ctggtataaa 2160 gtaactaatg tttggagcca acatttgttc cttgtgtcag caaaaggata ttcacattcc 2220 atgatccctg gctgagaatt ctgcctctag tctttcttac ccagctgttg tctatccttg 2280 ttcaattata aatactgcta agggcatttt taaaatacga tcttgtagtc cttaaatttg 2340 aatccgtcag cacggtcact cataggaaaa tgatcaaaca agcaagccag tcatgatttg 2400 act.ccttccc atctcatttc ttactgcctt acgctcatcc tgaggtccac cttggtctct 2460 aaaaacacca tgtgttctca tgcctccatg tcttttcaca cactgttcca tttgctcttc 2520 ctcccacatt acattgaaac tttcaagcct cagtcgaaac attgcttctt ctggatagca 2580 gccttcttga catccctcct cactccccag tccctacagg gcttccatag ctctttgtgt 2640 gcacttcgat cccagcattt tccatcgact tgtaattgtt tctgctacct gacaatcatc 2700 gccttgagta ctgggacaac ctttgattac tcattatatc ctcaataaat atttgttgaa 2760 cta 2763 <210> 47 <211> 1639 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1702940CB1 <400> 47 atcgcactga ggcttgagtc tgacttctct cccccacctg ctgtgccctt aaactgcaga 60 gatcggggcg ggggttgggg ggcaagcggc tcagatgggt tcaaaaaact ccccaggctc 120 aactctggtt ctgactgcct gagacatggg cagctgacac agcagacctt gaatcctgag 180 gatgtgaggc agggtatatc tgggaggccg gaggacgtgt ctggttatta cacagatgca 240 cagctggacg tgggatccac acagctcaga acagttggat cttgctcagt ctctgtcaga 300 ggaagatccc ttggacaaga ggaccctgcc ttggtgtgag agtgagggta gaggaagctg 360 gaacgagggt taaggaaaac cttccagtct ggacagtgac tggagagctc caaggaaagc 420 ccctcggtaa cccagccgct ggcaccatga acccagagag cagtatcttt attgaggatt 480 accttaagta tttccaggac caagtgagca gagagaatct gctacaactg ctgactgatg 540 atgaagcctg gaatggattc gtggctgctg ctgaactgcc cagggatgag gcagatgagc 600 tccgtaaagc tctgaacaag cttgcaagtc acatggtcat gaaggacaaa aaccgccacg 660 ataaagacca gcagcacagg cagtggtttt tgaaagagtt tcctcggttg aaaagggagc 720 ttgaggatca cataaggaag ctccgtgccc ttgcagagga ggttgagcag gtccacagag 780 gcaccaccat tgccaatgtg gtgtccaact ctgttggcac tacctctggc atcctgaccc 840 tcctcggcct gggtctggca cccttcacag aaggaatcag ttttgtgctc ttggacactg 900 gcatgggtct gggagcagca gctgctgtgg ctgggattac ctgcagtgtg gtagaactag 960 taaacaaatt gcgggcacga gcccaagccc gcaacttgga ccaaagcggc accaatgtag 1020 caaaggtgat gaaggagttt gtgggtggga acacacccaa tgttcttacc ttagttgaca 1080 attggtacca agtcacacaa gggattggga ggaacatccg tgccatcaga cgagccagag 1140 ccaaccctca gttaggagcg tatgccccac ccccgcatgt cattgggcga atctcagctg 1200 aaggcggtga acaggttgag agggttgttg aaggccccgc ccaggcaatg agcagaggaa 1260 ccatgatcgt gggtgcagcc actggaggca tcttgcttct gctggatgtg gtcagccttg 1320 catatgagtc aaagcacttg cttgaggggg caaagtcaga gtcagctgag gagctgaaga 1380 agcgggctca ggagctggag gggaagctca actttctcac caagatccat gagatgctgc 1440 agccaggcca agaccaatga ccccagagca gtgcagccac cagggcagaa atgccgggca 1500 caggccagga oaaaatgcag actttttttt ttttcaagtc tttgacgggg aagggagctc 1560 cgctttttcc cccagtaggg gtggcggggc ccaactctgg gccgtgtgaa cctcccgggg 1620 ggggggattc gattaacgc 1639 <210> 48 <221> 1600 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 1703342CB1 <400> 48 caaggcggcc caggacaggc aggggctgca cgcggtgaag aaaccaagac gcagagaggc 60 caagcccctt gccttgggtc acacagccaa aggaggcaga gccagaactc acaaccagat 120 ccagaggcaa cagggacatg gccacctggg acgaaaaggc agtcacccgc agggccaagg 180 tggctcccgc tgagaggatg agcaagttct taaggcactt cacggtcgtg ggagacgact 240 accatgcctg gaacatcaac tacaagaaat gggagaatga agaggaggag gaggaggagg 300 agcagccacc acccacacca gtctcaggcg aggaaggcag agctgcagcc cctgacgttg 360 cccctgcccc tggccccgca cccagggccc cccttgactt caggggcatg ttgaggaaac 420 tgttcagctc ccacaggttt caggtcatca tcatctgctt ggtggttctg gatgccctcc 480 tggtgcttgc tgagctcatc ctggacctga agatcatcca gcccgacaag aataactatg 540 ctgccatggt attccactac atgagcatca ccatcttggt cttttttatg atggagatca 600 tctttaaatt atttgtcttc cgcctggagt tctttcacca caagtttgag atcctggatg 660 ccgtcgtggt ggtggtctca ttcatcctcg acattgtcct cctgttccag gagcaccagt 720 ttgaggctct gggcctgctg attctgctcc ggctgtggcg ggtggcccgg atcatcaatg 780 ggattatcat ctcagttaag acacgttcag aacggcaact cttaaggtta aaacagatga 840 atgtacaatt ggccgccaag attcaacacc ttgagttcag ctgctctgag aaggaacaag 900 aaattgaaag acttaacaaa ctattgcgac agcatggact tcttggtgaa gtgaactaga 960 cccggaccag ctcccctcaa aaagaagaca ctgtctcatg ggcctgtgct gtcacgagag 1020 gaacagctgc ccctcctggg ccgcttggtg agaggtttgg tttgatacct ctgcctccct 1080 cctgccagca tggattctgg gtggacacag ccttgtggaa ggtccagtac caccaagagc 1140 tgcccatcca ctcccacccc acactgtatc aaatgtatca cattttctca tgttgaacac 1200 tttagcctta attgaaaatg agcaacaaag ctggacaatt gctagttgta tataaaattt 1260 aatctcaccg aatgtacagt tttcaaattt cacgtgtata ttaaggaact gatgcatctg 1320 agcattctga aagaaagaaa aagaagctac tttagctgcc accccattct agaaaagtct 1380 cttattttca agctgttcta aatagcttcg tctcagtttc cccaaaaggg gtacccaggc 1440 ccctcctctg tgtgccccag ctgcatcagc cagcttctag gtggctccat tgttttctgc 1500 cacctgacaa catttttcct caattactgt acaactactg tataaaataa aacaactact 1560 gtataaaata aactctctct tttccctgga aaaaaaaaaa 1600 <210> 49 <211> 2380 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1727529CB1 <400> 49 ctgagccatg gggggaaagc agcgggacga ggatgacgag gcctacggga agccagtcaa 60 atacgacccc tcctttcgag gccccatcaa gaacagaagc tgcacagatg tcatctgctg 120 cgtcctcttc ctgctcttca ttctaggtta catcgtggtg gggattgtgg cctggttgta 180 tggagacccc cggcaagtcc tctaccccag gaactctact ggggcctact gtggcatggg 240 ggagaacaaa gataagccgt atctcctgta cttcaacatc ttcagctgca tcctgtccag 300 caacatcatc tcagttgctg agaacggcct acagtgcccc acaccccagg tgtgtgtgtc 360 ctcctgcccg gaggacccat ggactgtggg aaaaaacgag ttctcacaga ctgttgggga 420 agtcttctat acaaaaaaca ggaacttttg tctgccaggg gtaccctgga atatgacggt 480 gatcacaagc ctgcaacagg aactctgccc cagtttcctc ctcccctctg ctccagctct 540 gggacgctgc tttccatgga ccaacattac tccaccggcg ctcccaggga tcaccaatga 600 caccaccata cagcagggga tcagcggtct tattgacagc ctcaatgccc gagacatcag 660 tgttaagatc tttgaagatt ttgcccagtc ctggtattgg attcttgttg ccctgggggt 720 ggctctggtc ttgagcctac tgtttatctt gcttctgcgc ctggtggctg ggcccctggt 780 gctggtgctg atcctgggag tgctgggcgt gctggcatac ggcatctact actgctggga 840 ggagtaccga gtgctgcggg acaagggcgc ctccatctcc cagctgggtt tcaccaccaa 900 cctcagtgcc taccagagcg tgcaggagac ctggctggcc gccctgatcg tgttggcggt 960 gcttgaagcc atcctgctgc tggtgctcat cttcctgcgg cagcggattc gtattgccat 1020 cgccctcctg aaggaggcca gcaaggctgt gggacagatg atgtctacca tgttctaccc 1080 actggtcacc tttgtcctcc tcctcatctg cattgcctac tgggccatga ctgctctgta 1140 cctggctaca tcggggcaac cccagtatgt gctctgggca tccaacatca gctcccccgg 1200 ctgtgagaaa gtgccaataa atacatcatg caaccccacg gcccaccttg tgaactcctc 1260 gtgcccaggg ctgatgtgcg tcttccaggg ctactcatcc aaaggcctaa tccaacgttc 1320 tgtcttcaat ctgcaaatct atggggtcct ggggctcttc tggaccctta actgggtact 1380 ggccctgggc caatgcgtcc tcgctggagc ctttgcctcc ttctactggg ccttccacaa 1440 gccccaggac atccctacct tccccttaat ctctgccttc atccgcacac tccgttacca 1500 cactgggtca ttggcatttg gagccctcat cctgaccctt gtgcagatag cccgggtcat 1560 cttggagtat attgaccaca agctcagagg agtgcagaac cctgtagccc gctgcatcat 1620 gtgctgtttc aagtgctgcc tctggtgtct ggaaaaattt atcaagttcc taaaccgcaa 1680 tgcatacatc atgatcgcca tctacgggaa gaatttctgt gtctcagcca aaaatgcgtt 1740 catgctactc atgcgaaaca ttgtcagggt ggtcgtcctg gacaaagtca cagacctgct 1800 gctgttcttt gggaagctgc tggtggtcgg aggcgtgggg gtcctgtcct tctttttttt 1860 ctccggtcgc atcccggggc tgggtaaaga ctttaagagc ccccacctca actattactg 1920 gctgcccatc atgacctcca tcctgggggc ctatgtcatc gccagcggct tcttcagcgt 1980 tttcggcatg tgtgtggaca cgctcttcct ctgcttcctg gaagacctgg agcggaacaa 2040 cggctccctg gaccggccct actacatgtc caagagcctt ctaaagattc tgggcaagaa 2100 gaacgaggcg cccccggaca acaagaagag gaagaagtga cagctccggc cctgatccag 2160 gactgcaccc cacccccacc gtccagccat ccaacctcac ttcgccttac aggtctccat 2220 tttgtggtaa aaaaaggttt taggccaggc gccgtggctc acgcctgtaa tccaacactt 2280 tgagaggctg aggcgggcgg atcacctgag tcaggagttc gagaccagcc tggccaacat 2340 ggtgaaacct ccgtctctat taaaaataca aaaattagcc 2380 <210> 50 <211> 3038 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 2289333CB1 <400> 50 aggggcaggg aggcgggcac caggcgcggg tccctccggg caggcgaggt aggcctgggc 60 ctgacgccgg ccacgcagcg gcgggagagt gagcactcgg gcggcggcgt cctggagacc 120 cgcgagagat ggaagcggcg gcgacgccgg cggctgccgg ggcggcgagg cgcgaggagc 180 tagatatgga tgtaatgagg cccttgataa atgagcagaa ttttgatggg acatcagatg 240 aagaacatga gcaagagctt ctgcctgttc agaagcatta ccaacttgat gatcaagagg 300 gcatttcatt tgtacaaact cttatgcacc ttcttaaagg aaatattgga actggccttt 360 taggacttcc attggcaata aaaaatgcag gcatagtgct tggaccaatc agccttgtgt 420 ttataggaat tatttctgtt cactgtatgc acatattggt acgttgcagt cactttctat 480 gtctgaggtt taaaaagtca acattaggtt atagtgacac tgtgagcttt gctatggaag 540' tgagtccttg gagttgtctt cagaagcaag cagcatgggg gcggagtgtg gttgactttt 600 ttctggtgat aacacagctg ggattctgta gtgtttatat tgtcttctta gctgaaaatg 660 tgaaacaagt tcatgaagga ttcctggaga gtaaagtgtt tatttcaaat agtaccaatt 720 catcaaaccc ttgtgagaga agaagtgttg acctaaggat atatatgctt tgctttcttc 780 catttataat tcttttggtc ttcattcgtg aactaaagaa tctatttgta ctttcattcc 840 ttgccaacgt ttccatggct gtcagtcttg tgataattta ccagtatgtt gtcaggaaca 900 tgccagatcc ccacaacctt ccaatagtgg ctggttggaa gaaataccca ctcttttttg,960 gtactgctgt atttgctttt gaaggcatag gagtggtcct tccactggaa aaccaaatga 2020 aagaatcaaa gcgtttccct caagcgttga atattggcat ggggattgtt acaactttgt 1080 atgtaacatt agctacttta ggatatatgt gtttccatga tgaaatcaaa ggcagcataa 1140 ctttaaatct tccccaagat gtatggttat atcaatcagt gaaaattcta tattcctttg 1200 gcatttttgt gacatattca attcagttct atgttccagc agagatcatt atccctggga 1260 tcacatccaa atttcatact aaatggaagc aaatctgtga atttgggata agatccttct 1320 tggttagtat tacttgtgcc ggagcaattc ttattcctcg tttagacatt gtgatttcct 1380 tcgttggagc tgtgagcagc agcacattgg ccctaatcct gccacctttg gttgaaattc 1440 ttacattttc gaaggaacat tataatatat ggatggtcct gaaaaatatt tctatagcat 1500 tcactggagt tgttggcttc ttattaggta catatataac tgttgaagaa attatttatc 1560 ctactcccaa agttgtagct ggcactccac agagtccttt tctaaatttg aattcaacat 1620 gcttaacatc tggtttgaaa tagtaaaagc agaatcatga gtcttctatt tttgtcccat 1680 ttctgaaaat tatcaagata actagtaaaa tacattgcta tatacataaa aatggtaaca 1740 aactctgttt tctttggcac gatattaata ttttggaagt aatcataact ctttaccagt 1800 agtggtaaac ctatgaaaaa tccttgcttt taagtgttag caatagttca aaaaattaag 1860 ttctgaaaat tgaaaaaatt aaaatgtaaa aaaattaaag aataaaaata cttctattat 1920 tcttttatct cagtaagaaa taccttaacc aagatatctc tcttttatgc tactcttttg 1980 ccactcactt gagaacagaa taggatttca acaataagag aataaaataa gaacatgtat 2040 aacaaaaagc tctctccaga tcatccctgt gaatgccaaa gtaaacttta tgtacagtgt 2100 aaaaaaaaaa aatctcagtt atgtttttat tagccaaatt ctaatgattg gctcctggaa 2160 gtatagaaaa ctcccattaa cataatataa gcatcagaaa attgcaaaca ctagaattaa 2220 ttttacactc taatggtagt tgatcttcat agtcaagagg cactgttcaa gatcatgact 2280 tagtgtttca atgaaatttg acaagggact ttaaaactta tccagtgcaa ctcccttgtt 2340 tttcgtcaga ggaaaaggag gcctagaaag gttaagtaac ttggtcgaga ccactcagcc 2400 ttgagatcaa gaaaacctaa tcttctgact cccaggccag gatgttttat ttctcacatc 2460 atgtccaaga aaaagaataa attatgttca gcttaatttt agtgttgaat ctatttgatt 2520 atattttaat actttgaaaa tgaatgtgtg atttttaata gtatatgtga cctgagcaga 2580 aaatcaggga actccaagaa gcctacactg tggccatata aacctcagca agagaaagaa 2640 gctatgttct tttaaaacag aatagagacc gcttgctggt gaaactcctg gctagtaaga 2700 tgtgtgtcta gctatactat ttgtggcttg agctttttta attattacct tcctttcctg 2760 agttttgtag gcaccacatt cctgaatggc agaaaataga cacctcagaa aacggaggat 2820 ttgtggactc tttccagccc tgtggctttt cttatcacag ccttttattt attatgagca 2880 gaataaaaga atcagctagg tgtggtggtc tgtgcttata atcccagcta ctctggagga 2940 taagttggga ggatcacttg agaggccagg agcttgagac cagcctgggc agcatagtga 3000 gacctcgact ctataaaaca taaaaaaaaa aaaaaaaa 3038 <210> 51 <211> 2608 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 2720354CB1 <400> 51 taggctaatt ttttttacag acacgatttc gccacgttgg ccaggctggt cttgaactcc 60 tgacctcaag tgatccaccc acctcagcct cccaaagtgt tgggattaca ggcgtgagcc 120 actgcacctg gccaggctca tcactttttg cgcctattgc ctcgaagcca gtctctgatg 180 ggacattagg gcaggggccc ttcagcctag tctgggacat gggccgctca ctcagcagta 240 tgacaagcat cacctggaga acgggccagt ctcaggaggt cgttcatgcc ccactggcag 300 tgcactgtgc agccatagtg taaacaagag gcttaacctg aactggtctg agatcttggg 360 gaccccctac cctgtctcca gcagcctgtc cctttagctg tttgcctact ggcaccccat 420 cctgagaagg catagatacc cggcccaccc tgccctggaa ttacaaaagt cttagactgt 480 gcctgagtgc ccggcctcct tgggagaccc tcctaggcag cctaagcacc agacccggga 540 gctgggtgct ctggtcctgc ctgcctgcct ctcactggac cctctccttc caggtacggc 600 ttcaggtcca gagcgtggag aagcctcagt accgcgggac gttgcactgc ttcaagtcca 660 tcatcaagca agagagcgtg ctgggcctgt acaagggcct gggctcgccg ctcatggggc 720 tcaccttcat caacgcgctg gtgttcgggg tgcagggcaa caccctccgg gccctgggcc 780 acgactcgcc cctcaaccag ttcctggcag gtgcggcggc gggcgccatc cagtgcgtca 840 tctgctgccc catggagctg gccaagacgc ggctgcagct gcaggacgcg ggcccagcgc 900 gcacctacaa gggctcgctg gactgcctcg cgcagatcta cgggcacgag ggtctgcgtg 960 gcgtcaaccg gggcatggtg tccacgttgc tgcgtgagac tcccagcttc ggcgtctact 1020 tcctcaccta tgacgctctc acgcgggcgc tgggctgcga gccgggcgac cgcctgctgg 1080 tgcccaagct gctgttggcg ggcggtacgt caggcatcgt gtcctggctc tctacctatc 1140 ctgtggacgt ggtcaagtcg cggctgcagg cggacggact gcggggcgcc ccgcgctacc 1200 gcggcatcct ggactgcgtg caccagagct accgcgccga gggctggcgc gtcttcacac 1260 gggggctggc gtccacgctg ctgcgcgcct tccccgtcaa cgctgccacc ttcgccaccg 1320 tcacggtggt gctcacctac gcgcgcggcg aggaggccgg gcccgagggc gaggctgtgc 2380 ccgccgcccc tgcggggcct gccctggcgc agccctccag cctgtgacgc tcaccccgcc 1440 ctccttcccc agggctcctt ctcagaaacc tgggacataa attggcccct gagtcgattg 1500 ccctgcttcc tgctgggatg ctgcgagctg tggagtctat cagatgtggg ctgaattttg 1560 ctgatcagct gggtagtttt ggccgagaac tgcacttgcc tcagtgttct catctatgaa 1620 ataaggaccc tcatgcccac actgtagagt cacgaagctc agagattatt cccagcagca 1680 gecagcacct ggcctggctg aggccattgc accgttatcc tggaaactga ggcagacact 1740 ccagcccctt tctgggatcc tggccacgtc attgtgctcc tgccctgcag gctggctccc 1800 gggggtctct gatggccaac caaggggcca cccagggacc tctaactcca cacatcctcc 1860 acccgggggg gtggtgggcc acccctctgg tctgtgttag ggacagagga aaacttggtg 1920 tgcctcctgg tgtcacagaa ctggatcctc tgcatacccc agcttctcca catgccactg 1980 ctaggggtac cccagctgct gccactcctg ctggagggtg aactggggac cctgcaccct 2040 ccgggaagcc atggagtctg ctggaggcac catatcagcc tgcgggacta gggtggggag 2100 caaacaggcc agcggtggag gtctggacag ttcaagtgtg atgcagctgt ggcaaggaga 2160 aatccttccg cctctgggcc tcaggctgcc tgtccataaa atggggacat ggccagctga 2220 cggacaactg agtctccggc ccacctacca ccgccagcca ggatccccca aagtgtgcag 2280 agggctcagc agagaacagt atgggacccc ctcaccaggc ctggaacacc tccagccaca 2340 aagaagccaa aggtcagtcc ctctgctccc cagcaaacgg tgcctcccag gcattctcag 2400 tgccagggct tcatccctgt gaaggcacag ggcctgctag tgggcacagg ggtggctagt 2460 tggggcctgg ggcagaggag ggctgcacca ggcgtcctgg ggaatgtgct cagtgaagac 2520 gacactgggc tttgcacagc ctggtgtcgc tgtacagaaa ctgtcaaggg aataaagtgt 2580 tctttgtttt ttaaaaaaaa aaaaaaaa 2608 <210> 52 <211> 3804 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 3038193CB1 <400> 52 ccctttcctg tcactggcta ctaccactcc caaccctcct caaagccgcc ggagcaaccc 60 ccaggtcttt actttacaat cggcaatttg acttgctctg ctgcatgtct ggagggacca 120 aggaaagtgt ggagacgctc caaggattag gtgatcggag cttgaaaaga aaaaaagcca 180 aacaaataaa caaaacccac ccaccctaac aaatatgagg ctgctggaga gaatgaggaa 240 agactggttc atggtcggaa tagtgctggc gatcgctgga gctaaactgg agccgtccat 300 aggggtgaat gggggaccac tgaagccaga aataactgta tcctacattg ctgttgcaac 360 aatattcttt aacagtggac tatcattgaa aacagaggag ctgaccagtg ctttggtgca 420 tctaaaactg catcttttta ttcagatctt tactcttgca ttcttcccag caacaatatg 480 gctttttctt cagcttttat caatcacacc catcaacgaa tggcttttaa aaggtttgca 540 gacagtaggt tgcatgcctc cgcctgtgtc ttctgcagtg attttaacca aggcagttgg 600 tggaaatgag ggcatcgtta taacacccct gctcctgctg ctttttcttg gttcatcttc 660 ttctgtgcct ttcacatcta ttttttctca gctttttatg actgttgtgg ttcctctcat 720 cattggacag attgtccgaa gatacatcaa ggattggctt gagagaaaga agcctccttt 780 tggtgctatc agcagcagtg tactcctcat gatcatctac acaacattct gtgacacgtt 840 ctctaaccca aatattgacc tggataaatt cagccttgtt ctcatactgt tcataatatt 900 ttctatccag ctgagtttta tgcttttaac tttcatcttt tcaacaagga ataattcggg 960 tttcacacca gcagacacag tggctatcat tttctgttct acacacaaat cccttacatt 1020 gggaattccg atgctgaaga tcgtgtttgc aggctatgag catctctctt taatatctgt 1080 acccttgctc atctaccacc cagctcagat ccttctggga agtgtgttgg tgccaacaat 1140 caagtcttgg atggtatcaa ggcagaagaa actactccaa accagggggc cactggctaa 1200 cttgaataat ccagaaggct tggaatatct atccatcaaa tttgggcatt aaaataaata 1260 ccaagagtcc atcctccagg gagtgaagct gacaaggccg acagtataac aaaggaggtg 1320 gactttctgt agcaatgtat atatgtacag gattgtacat actagcaatt ctgaagactt 1380 gtacttgtga atgttgcctc aatgcatatt ttattttttt acacaaaaat atgagatcct 1440 gtttaagtgc cttaaaatgt atttgacaag agcgttattt ccacaatatg ctttgttgat 150.0 tactgccagg ggtggtacaa tatttggggg ttaattttgc tttcctaatg caggaatcag 1560 tcatggtaag tgacaaaaag caaacatgct ttccctgcag cacctttgtg taatacaacc 1620 ctatagtagt tactgtaatg tttgaaatga ggtcacacca tcaggaaaat gcccttctga 1680 tgacagtgaa aatttccaaa gtcttattca tgcatacttt gatttactgt gtgattcttt 1740 ttttctacga ctgtgacatg cctcttcctt atcaactcag caggggtcat agatcgaata 1800 gatgctgaaa agcgtaagat atatgcattc cttgacatca tttttaaaga cattccttca 1860 aatagtttcc acacagaaat tcctcactcc cattatgaga gattgtggtt atatgtctta 1920 aatttattat aagctgcttc aaagaaaggg tctgaatgtt tgaattatga gtgaaatcat 1980 gtgaaatttt gagttaaact ctgtgatttg attttcaggg tctttaaaat atatcttaat 2040 atcttcttcc tctttattca ataatttctg tcttgcactt acacactcat aacagccaaa 2100 tatgaggcac aaaaatgtta caatcagttt gaaagcagca tcaattaatg gtagattcta 2160 ttcacattcc acaacccaga ccaaattttt ttcctattac gcagatgtgc tgagcacttt 2220 ccagattgcc cctgttggcc aaaagcagcc tgttacatcc tggaattaag cacacttaag 2280 gtatttgaga caatttatta atgaaaattt ccttggcaga tttgacaaat gttggcaata 2340 tttttttaaa agttaaatca tattgctttc atgaataaat gaaaatataa aggtcatgga 2400 tgcaaacaaa tgttacatat acacattctg tctctccaga tgaaaagaac atgcaaaacc 2460 atttaataac caaaatatca agtaaaatta gttcccaacg gggcagcagc tttcaaatga 2520 gtgtccaata tttgcttctg ctatagctgc aagaactgta actggaccca agtagagaat 2580 gaagccacgt atagaactac gagaacactt ttctgtgttt cccccatgcc gtcctgtcac 2640 atcctcttac acgtcctctc ttgatttgat agacaatatt ggcatcctgg gtctcactga 2700 ggccgtgcta tgtcctcagc agctgttttt gttgtttcgt tattatgccc acaacaaaaa 2760 atcattcctt agaaactcac caagtttatc tactgtgtaa atttatatta ttgttactac 2820 caggtctcat cttttgtcaa tgtcattgaa taaatttcat aagagttatt ctcagtgtga 2880 attttaaggc taatgccaga tcctgcaaaa atctatgcta accaggctgt agtacacact 2940 gttataaaga attttacttg tgtctaaaac tacagtaatt ttgcttaggt aattgtgctt 3000 acctatggag cacaggaagg ctcttaggtt ttgttcctac aagtttcttt gaattttgga 3060 gtaaatggaa gtgtctgtct gtctgtcatc tatctgccct atcataaaaa tctttctccc 3120 taacattaaa atactgatcc ccgcccccaa cttatctacc tctattgtct aacacctata 3180 gtaggtgtga tcatgggata aaattcaact gaaaatgcta tgataacatt ttatcgtttg 3240 ctttaaaaat gtgctttgtt ttcaaataat ctttacatag tgaactttgg tggcgttagt 3300 gatatgttta tgcctatttc ttttttttac acaaattcct tggcatattt tttcataaag 3360 aacaaaaaat aaaatcaaaa tttattttta attcatgctt attgggattt aattattcag 3420 agcttaaaat attttgttat gtttatacac tgtaaagcta tctgttttat gcatttgttt 3480 tgtctaaatg tatttatgaa agaaatacat tagattatat ttatgtttac tcatttttcc 3540 acctggattt tttttaatgg ttgttacaaa attagatttt ttaatgggta ataatgttgg 3600 tattttcatg ttttttctta gtattaaaat ttttgtgggt tttttaaaat ttttccctat 3660 tctgttaaaa attaacacac ctctagctaa tgttcagtgt ttgtgctaaa taccaaattt 3720 tttcaaaagg attggttaag tcataaagtg gattatttat gatgactgga agatgaaaat 3780 aattatatga ttaaacaaag aatg 3804 <220> 53 <211> 1894 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte Ib No: 3460979CB1 <220>
<221> unsure <222> 1651 <223> a, t, c, g, or other <400> 53 acggatcact agtatgcggc gcagtgtgct ggaaagggaa caaacatggc cgctctggcg 60 cccgtcggct cccccgcctc ccgcggtcct aggctggccg cgggcctccg gctgctccca 120 atgctgggtt tgctgcagtt gctggccgag cctggcctgg gccgcgtcca tcacctggca 180 ctcaaggatg atgtgaggca taaagttcat ctgaacacct ttggcttctt caaggatggg 240 tacatggtgg tgaatgtcag tagcctctca ctgaatgagc ctgaagacaa ggatgtgact 300 attggattta gcctagaccg tacaaagaat gatggctttt cttcttacct ggatgaagat 360 gtgaattact gtattttaaa gaaacagtct gtctctgtca cccttttaat cctagacatc 420 tccagaagtg aggtaagagt aaagtctcca ccagaagctg gtacccagtt accaaagatc 480 atcttcagca gggatgagaa agtccttggt cagagccagg agcctaatgt taaccctgct 540 tcagcaggca accagaccca gaagacacaa gatggtggaa agtctaaaag aagtacagtg 600 gattcaaagg ccatgggaga gaaatccttt tctgttcata ataatggtgg ggcagtgtca 660 tttcagtttt tctttaacat cagcactgat gaccaagaag gcctttacag tctttatttt 720;
cataaatgcc ttggaaaaga attgccaagt gacaagttta cattcagcct tgatattgag 780 atcacagaga agaatcctga cagctacctc tcagcaggag aaattcctct ccccaaatta 840 tacatctcaa tggccttttt cttctttctt tctgggacca tctggattca tatccttcga 900 aaacgacgga atgatgtatt taaaatccac tggctgatgg cggcccttcc tttcaccaag 960 tctctttcct tggtgttcca tgcaattgac taccactaca tctcctccca gggcttccct 1020 atcgaaggct gggctgttgt gtactacata actcaccttt tgaaaggggc gctactcttc 1080 atcaccattg cactcattgg cactggctgg gctttcatta agcacatcct ttctgataaa 1140 gacaaaaaga tcttcatgat tgtcattcca ctccaggtcc tggcaaatgt agcctacatc 1200 atcatagagt ccaccgagga gggcacgact gaatatggct tgtggaagga ctctctattt 1260 ctggtcgacc tgttgtgttg tggtgccatc ctcttcccag tggtgtggtc aatcagacat 1320 ttacaagaag catcagcaac agatggaaaa gctgctatta acttagcaaa gctgaaactt 1380 ttcagacatt attacgtctt gattgtgtgt tacatatact tcactaggat cattgcattt 1440 ctcctcaaac tcgctgttcc attccagtgg aagtggctct accagctcct ggatgaaacg 1500 gccacactgg tcttctttgt tctaacgggg tataaattcc gtccggcttc agataacccc 1560 tacctacaac tttctcagga agaagaagac ttggaaatgg agtccgtgta agaaatcttt 1620 cttccctctt ccttagccct gaaccctttg nctaacacaa agcagcacag tgtgaatcga 1680 gccggctggt ctcagcattt cgtggctgca ggggtgggtc ctctatattt agcagaaggg 1740 accggcactg gagcccaagg ggtcggtctg gttgaaggca agatttggca accatactgg 1800 gctgtgccgg aaaaggaaag ggggggccaa aaaacaattg gggccggcgt caaaaaaccg 1860 ggcgaacaag agaaaaagcg ggcccaggag aaag 1894 <210> 54 <211> 1668 <212> DNA
<213> Homo Sapiens <220>

<221> misc_feature <223> Incyte ID No: 7472200CB1 <400> 54 atgacactgg tttactttcc tccttcaaag cttcagcagc agcagcagcc atcgagatcc 60 agtcgcctgg cccaacagtt ggcccaatcc tcctggcagc tggccctgcg ctttggcaaa 120 cggaccacta tccacggcct ggacaggctg cttagtgcca aggccagtcg atgggagcga 180 ttcgtctggc tgtgcacctt tgtgagtgcc ttcctgggcg cggtgtacgt ttgcctgatt 240 ctctccgccc gctacaacgc cgcccacttc cagacggtgg tggatagcac gcggtttccg 300 gtttaccgca taccatttcc ggtcataacg atctgcaacc ggaatcgcct caactggcaa 360 cgcctggcgg aggcgaagtc aagattcctg gccaacggca gcaactccgc ccagcaggag 420 ctcttcgagc tgattgtggg cacctacgac gatgcttact tcggtcactt tcagtccttc 480 gagcgattgc gcaaccagcc aacggagctg ctcaactatg tcaatttcag ccaggtggtg 540 gattttatga cctggcgctg caacgagctg ctcgcggaat gcctgtggcg ccaccatgcc 600 tacgactgct gcgagatccg ctcgaagcgg cgcagcaaga acggcttgtg ctgggctttc 660 aactcgctgg agacggaaga gggcaggcgg atgcagctgc tcgatcccat gtggccctgg 720 cgtactgggt cggcgggtcc catgagcgcc ctctccgtgc gtgttctcat ccagcccgcg 780 aagcactggc cggggcacag ggagacgaat gccatgaagg gcatcgatgt catggttacc 840 gagccatttg tgtggcacaa caatccgttc ttcgtggccg cgaacacgga gacgaccatg 900 gagatcgaac ccgtcatcta cttctatgac aacgacaccc ggggagttcg ctccgaccag 960 cgccagtgcg tcttcgatga tgagcacaac agcaaggatt tcaagtcgct gcaaggatac 1020 gtttacatga ttgaaaactg tcagtccgag tgccatcagg agtacttggt gcgctattgc 1080 aactgcacaa tggacctact gtttccaccg gacctgctca tctactccca caatcccggc 1140 gagaaggagt tcgttcgcaa ccaatttcag ggaatgtcct gcaagtgctt ccgcaactgc 1200 tactccctca actacatcag cgatgtccgg cccgccttcc tgccaccgga tgtgtacgca 1260 aacaactcct atgtggacct ggatgtgcac tttcgcttcg agaccattat ggtctatcgc 1320 accagcctcg tcttcggctg ggtggactta atggttagct ttggaggaat tgccggtctt 1380 tttcttggct gctccctaat tagtggcatg gaactggcct atttcctgtg cattgaggtg 1440 ccggcctttg ggctggatgg actgcgtcga aggtggaagg ctcgacggca gatggatctg 1500 ggcgtaaccg tgcccacgcc cactttgaac tttcaacaaa ccacgcccag tcagctgatg 1560 gagaactaca ttatgcaact gaaggctgag aaggcgcaac agcagaaggc gaactttcaa 1620 aactggcacc gcataacatt tgctcaaaag catgttattg gcaagtga 1668

Claims (28)

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:1-27, 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:1-27, c) a biologically active fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-27, and d) an immunogenic fragment of an amino acid sequence selected from the group consisting of SEQ ID NO:1-27.

2. An isolated polypeptide of claim 1 selected from the group consisting of SEQ ID NO:1-27.

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

4. An isolated polynucleotide encoding a polypeptide of claim 2.

5. An isolated polynucleotide of claim 4 selected from the group consisting of SEQ ID
NO:28-54.

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

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

8. A transgenic organism comprising a recombinant polynucleotide of claim 6.

9. 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.

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

11. 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:28-54, 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:28-54, c) a polynucleotide sequence complementary to a), d) a polynucleotide sequence complementary to b), and e) an RNA equivalent of a)-d).

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

13. A method for detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide of claim 11, the method comprising:
a) hybridizing the sample with a probe comprising at least 20 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 or fragments thereof, and b) detecting the presence or absence of said hybridization complex, and, optionally, if present, the amount thereof.

14. A method of claim 13, wherein the probe comprises at least 60 contiguous nucleotides.

15. A method for detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide of claim 11, the method comprising:
a) amplifying said target polynucleotide or fragment thereof using polymerase chain reaction amplification, and b) detecting the presence or absence of said amplified target polynucleotide or fragment thereof, and, optionally, if present, the amount thereof.

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

17. A composition of claim 16, wherein the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NO:1-27.

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

19. 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.

20. A composition comprising an agonist compound identified by a method of claim 19 and a pharmaceutically acceptable excipient.

21. A method for treating a disease or condition associated with decreased expression of functional TRICH, comprising administering to a patient in need of such treatment a composition of claim 20.

22. 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.

23. A composition comprising an antagonist compound identified by a method of claim 22 and a pharmaceutically acceptable excipient.

24. A method for treating a disease or condition associated with overexpression of functional TRICH, comprising administering to a patient in need of such treatment a composition of claim 23.

25. A method of screening for a compound that specifically binds to the polypeptide of claim 1, said method comprising the steps of:
a) combining the polypeptide of claim 1 with at least one test compound under suitable conditions, and b) detecting binding of the polypeptide of claim 1 to the test compound, thereby identifying a compound that specifically binds to the polypeptide of claim 1.

26. A method of screening for a compound that modulates the activity of the polypeptide of claim 1, said method comprising:
a) combining the polypeptide of claim 1 with at least one test compound under conditions permissive for the activity of the polypeptide of claim 1, b) assessing the activity of the polypeptide of claim 1 in the presence of the test compound, and c) comparing the activity of the polypeptide of claim 1 in the presence of the test compound with the activity of the polypeptide of claim 1 in the absence of the test compound, wherein a change in the activity of the polypeptide of claim 1 in the presence of the test compound is indicative of a compound that modulates the activity of the polypeptide of claim 1.

27. A method for screening a compound for effectiveness in altering expression of a target polynucleotide, wherein said target polynucleotide comprises a sequence of claim 5, the method comprising:
a) exposing a sample comprising the target polynucleotide to a compound, under conditions suitable for the expression of the target polynucleotide, b) detecting altered expression of the target polynucleotide, and c) comparing the expression of the target polynucleotide in the presence of varying amounts of the compound and in the absence of the compound.

28. A method for assessing toxicity of a test compound, said method comprising:
a) treating a biological sample containing nucleic acids with the test compound;
b) hybridizing the nucleic acids of the treated biological sample with a probe comprising at least 20 contiguous nucleotides of a polynucleotide of claim 11 under conditions whereby a specific hybridization complex is formed between said probe and a target polynucleotide in the biological sample, said target polynucleotide comprising a polynucleotide sequence of a polynucleotide of claim 11 or fragment thereof;
c) quantifying the amount of hybridization complex; and d) comparing the amount of hybridization complex in the treated biological sample with the amount of hybridization complex in an untreated biological sample, wherein a difference in the amount of hybridization complex in the treated biological sample is indicative of toxicity of the test compound.