CN1276010A - CBFBGA09: a human SL15 homolog - Google Patents

Abstract

GBFBGA09 polypeptides and polynucleotides and methods for producing such polypeptides by recombinant techniques are disclosed. Also disclosed are methods for utilizing CBFBGA09 polypeptides and polynucleotides in the design of protocols for the treatment of deficiencies in mannose metabolism, among others, and diagnostic assays for such conditions.

Description

CBFBGA09: a human SL15 homolog

CBFBGA09: A HUMAN SL15 HOMOLOG

FIELD OF INVENTION

This invention relates to newly identified polynucleotides, polypeptides encoded by them and to the use of such polynucleotides and polypeptides, and to their production. More particularly, the polynucleotides and polypeptides of the present invention relate to the SLI 5 family, hereinafter referred to as CBFBGA09. The invention also relates to inhibiting or activating the action of such polynucleotides and polypeptides.

BACKGROUND OF THE INVENTION

SL15, a suppressor of the Lecl5 and Lec35 CHO mutant cells, can correct the abnormalities in intracellular mannose metabolism. Therefore, SLI 5 may participate in mannose metabolism. This indicates that the family has an established, proven history as therapeutic targets. Clearly there is a need for identification and characterization of further members of the family which can play a role in preventing, ameliorating or correcting dysfunctions or diseases, including, but not limited to, metabolic disorders.

SUMMARY OF THE INVENTION In one aspect, the invention relates to CBFBGA09 polypeptides and recombinant materials and methods for their production. Another aspect of the invention relates to methods for using such CBFBGA09 polypeptides and polynucleotides. Such uses include the treatment of etabolic disorders, among others. In still another aspect, the invention relates to methods to identify agonists and antagonists using the materials provided by the invention, and treating conditions associated with CBFBGA09 imbalance with the identified compounds. Yet another aspect of the invention relates to diagnostic assays for detecting diseases associated with inappropriate CBFBGA09 activity or levels.

DESCRIPTION OF THE INVENTION Definitions The following definitions are provided to facilitate understanding of certain terms used frequently herein.

"CBFBGA09" refers, among others, generally to a polypeptide having the amino acid sequence set forth in SEQ ID NO: 2 or an allelic variant thereof.

CONFIRMATION COP* "CBFBGA09 activity or CBFBGA09 polypeptide activity" or "biological activity of the CBFBGA09 or CBFBGA09 polypeptide" refers to the metabolic or physiologic function of said CBFBGA09 including similar activities or improved activities or these activities with decreased undesirable side-effects. Also included are antigenic and immunogenic activities of said CBFBGA09.

"CBFBGA09 gene" refers to a polynucleotide having the nucleotide sequence set forth in SEQ ID NO:l or allelic variants thereof and/or their complements.

"Antibodies" as used herein includes polyclonal and monoclonal antibodies, chimeric, single chain, and humanized antibodies, as well as Fab fragments, including the products of an Fab or other immunoglobulin expression library.

"Isolated" means altered "by the hand of man" from the natural state. If an "isolated" composition or substance occurs in nature, it has been changed or removed from its original environment, or both. For example, a polynucleotide or a polypeptide naturally present in a living animal is not "isolated," but the same polynucleotide or polypeptide separated from the coexisting materials of its natural state is "isolated", as the term is employed herein.

"Polynucleotide" generally refers to any polyribonucleotide or polydeoxribonucleotide, which may be unmodified RNA or DNA or modified RNA or DNA. "Polynucleotides" include, without limitation single- and double-stranded DNA, DNA that is a mixture of single- and double- stranded regions, single- and double-stranded RNA, and RNA that is mixture of single- and double-stranded regions, hybrid molecules comprising DNA and RNA that may be single-stranded or, more typically, double-stranded or a mixture of single- and double-stranded regions. In addition, "polynucleotide" refers to triple-stranded regions comprising RNA or DNA or both RNA and DNA. The term polynucleotide also includes DNAs or RNAs containing one or more modified bases and DNAs or RNAs with backbones modified for stability or for other reasons. "Modified" bases include, for example, tritylated bases and unusual bases such as inosine. A variety of modifications has been made to DNA and RNA; thus, "polynucleotide" embraces chemically, enzymatically or metabolically modified forms of polynucleotides as typically found in nature, as well as the chemical forms of DNA and RNA characteristic of viruses and cells. "Polynucleotide" also embraces relatively short polynucleotides, often referred to as oligonucleotides. "Polypeptide" refers to any peptide or protein comprising two or more amino acids joined to each other by peptide bonds or modified peptide bonds, i.e., peptide isosteres. "Polypeptide" refers to both short chains, commonly referred to as peptides, oligopeptides or oligomers, and to longer chains, generally referred to as proteins. Polypeptides may contain amino acids other than the 20 gene-encoded amino acids. "Polypeptides" include amino acid sequences modified either by natural processes, such as posttranslational processing, or by chemical modification techniques which are well known in the art. Such modifications are well described in basic texts and in more detailed monographs, as well as in a voluminous research literature. Modifications can occur anywhere in a polypeptide, including the peptide backbone, the amino acid side-chains and the amino or carboxyl termini. It will be appreciated that the same type of modification may be present in the same or varying degrees at several sites in a given polypeptide. Also, a given polypeptide may contain many types of modifications. Polypeptides may be branched as a result of ubiquitination, and they may be cyclic, with or without branching. Cyclic, branched and branched cyclic polypeptides may result from posttranslation natural processes or may be made by synthetic methods. Modifications include acetylation, acylation, ADP-ribosylation, amidation, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphotidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent cross-links, formation of cystine, formation of pyroglutamate, formylation, gamma- carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, transfer-RNA mediated addition of amino acids to proteins such as arginylation, and ubiquitination. See, for instance, PROTEINS - STRUCTURE AND MOLECULAR PROPERTIES, 2nd Ed., T. E. Creighton, W. H. Freeman and Company, New York, 1993 and Wold, F., Posttranslational Protein Modifications: Perspectives and Prospects, pgs. 1-12 in POSTTRANSLATIONAL COVALENT MODIFICATION OF PROTEINS, B. C. Johnson, Ed., Academic Press, New York, 1983; Seifter et al, "Analysis for protein modifications and nonprotein cofactors", Meth Enzymol (1990) 182:626-646 and Rattan et al, "Protein Synthesis: Posttranslational Modifications and Aging", Ann NYAcadSci (1992) 663:48-62. "Variant" as the term is used herein, is a polynucleotide or polypeptide that differs from a reference polynucleotide or polypeptide respectively, but retains essential properties. A typical variant of a polynucleotide differs in nucleotide sequence from another, reference polynucleotide. Changes in the nucleotide sequence of the variant may or may not alter the amino acid sequence of a polypeptide encoded by the reference polynucleotide. Nucleotide changes may result in amino acid substitutions, additions, deletions, fusions and truncations in the polypeptide encoded by the reference sequence, as discussed below. A typical variant of a polypeptide differs in amino acid sequence from another, reference polypeptide. Generally, differences are limited so that the sequences of the reference polypeptide and the variant are closely similar overall and, in many regions, identical. A variant and reference polypeptide may differ in amino acid sequence by one or more substitutions, additions, deletions in any combination. A substituted or inserted amino acid residue may or may not be one encoded by the genetic code. A variant of a polynucleotide or polypeptide may be a naturally occurring such as an allelic variant, or it may be a variant that is not known to occur naturally. Non-naturally occurring variants of polynucleotides and polypeptides may be made by mutagenesis techniques or by direct synthesis.

"Identity" is a measure of the identity of nucleotide sequences or amino acid sequences. In general, the sequences are aligned so that the highest order match is obtained. "Identity" per se has an art-recognized meaning and can be calculated using published techniques. See, e.g.: (COMPUTATIONAL MOLECULAR BIOLOGY, Lesk, A.M., ed., Oxford University Press, New York, 1988; BIOCOMPUTING: INFORMATICS AND GENOME PROJECTS, Smith, D.W., ed., Academic Press, New York, 1993; COMPUTER ANALYSIS OF SEQUENCE DATA, PART I, Griffin, A.M., and Griffin. H.G., eds., Humana Press, New Jersey, 1994; SEQUENCE ANALYSIS IN MOLECULAR BIOLOGY, von Heinje, G., Academic Press, 1987; and SEQUENCE ANALYSIS PRIMER, Gribskov, M. and Devereux, J., eds., M Stockton Press, New York, 1991). While there exist a number of methods to measure identity between two polynucleotide or polypeptide sequences, the term "identity" is well known to skilled artisans (Carillo, H., and Lipton, D., S1AM J Applied Math (1988) 48:1073). Methods commonly employed to determine identity or similarity between two sequences include, but are not limited to, those disclosed in Guide to Huge Computers, Martin J. Bishop, ed., Academic Press, San Diego, 1994, and Carillo, H., and Lipton, D., SUM J Applied Math (1988) 48:1073. Methods to determine identity and similarity are codified in computer programs. Preferred computer program methods to determine identity and similarity between two sequences include, but are not limited to, GCS program package (Devereux, J., et al, Nucleic Acids Research (1984) 12(1):387), BLASTP, BLASTN, FASTA (Atschul, S.F. et al, JMolec Biol (1990) 215:403). As an illustration, by a polynucleotide having a nucleotide sequence having at least, for example, 95% "identity" to a reference nucleotide sequence of SEQ ID NO: 1 is intended that the nucleotide sequence of the polynucleotide is identical to the reference sequence except that the polynucleotide sequence may include up to five point mutations per each 100 nucleotides of the reference nucleotide sequence of SEQ ID NO: 1. In other words, to obtain a polynucleotide having a nucleotide sequence at least 95% identical to a reference nucleotide sequence, up to 5% of the nucleotides in the reference sequence may be deleted or substituted with another nucleotide, or a number of nucleotides up to 5% of the total nucleotides in the reference sequence may be inserted into the reference sequence. These mutations of the reference sequence may occur at the 5 or 3 terminal positions of the reference nucleotide sequence or anywhere between those terminal positions, interspersed either individually among nucleotides in the reference sequence or in one or more contiguous groups within the reference sequence.

Similarly, by a polypeptide having an amino acid sequence having at least, for example, 95% "identity" to a reference amino acid sequence of SEQ ID NO:2 is intended that the amino acid sequence of the polypeptide is identical to the reference sequence except that the polypeptide sequence may include up to five amino acid alterations per each 100 amino acids of the reference amino acid of SEQ ID NO: 2. In other words, to obtain a polypeptide having an amino acid sequence at least 95% identical to a reference amino acid sequence, up to 5% of the amino acid residues in the reference sequence may be deleted or substituted with another amino acid, or a number of amino acids up to 5% of the total amino acid residues in the reference sequence may be inserted into the reference sequence. These alterations of the reference sequence may occur at the amino or carboxy terminal positions of the reference amino acid sequence or anywhere between those terminal positions, interspersed either individually among residues in the reference sequence or in one or more contiguous groups within the reference sequence.

Polypeptides of the Invention

In one aspect, the present invention relates to CBFBGA09 polypeptides (or CBFBGA09 proteins). The CBFBGA09 polypeptides include the polypeptide of SEQ ID NO:2; as well as polypeptides comprising the amino acid sequence of SEQ ID NO: 2; and polypeptides comprising the amino acid sequence which have at least 80% identity to that of SEQ ID NO:2 over its entire length, and still more preferably at least 90% identity, and even still more preferably at least 95% identity to SEQ ID NO: 2. Furthermore, those with at least 97-99% are highly preferred. Also included within CBFBGA09 polypeptides are polypeptides having the amino acid sequence which have at least 80% identity to the polypeptide having the amino acid sequence of SEQ ID NO:2 over its entire length, and still more preferably at least 90% identity, and still more preferably at least 95% identity to SEQ ID NO:2. Furthermore, those with at least 97-99% are highly preferred. Preferably CBFBGA09 polypeptide exhibit at least one biological activity of CBFBGA09.

The CBFBGA09 polypeptides may be in the form of the "mature" protein or may be a part of a larger protein such as a fusion protein. It is often advantageous to include an additional amino acid sequence which contains secretory or leader sequences, pro-sequences, sequences which aid in purification such as multiple histidine residues, or an additional sequence for stability during recombinant production.

Fragments of the CBFBGA09 polypeptides are also included in the invention. A fragment is a polypeptide having an amino acid sequence that entirely is the same as part, but not all, of the amino acid sequence of the aforementioned CBFBGA09 polypeptides. As with CBFBGA09 polypeptides, fragments may be "free-standing," or comprised within a larger polypeptide of which they form a part or region, most preferably as a single continuous region. Representative examples of polypeptide fragments of the invention, include, for example, fragments from about amino acid number 1-20, 21-40, 41-60, 61- 80, 81-100, and 101 to the end of CBFBGA09 polypeptide. In this context "about" includes the particularly recited ranges larger or smaller by several, 5, 4, 3, 2 or 1 amino acid at either extreme or at both extremes.

Preferred fragments include, for example, truncation polypeptides having the amino acid sequence of CBFBGA09 polypeptides, except for deletion of a continuous series of residues that includes the amino terminus, or a continuous series of residues that includes the carboxyl terminus or deletion of two continuous series of residues, one including the amino terminus and one including the carboxyl terminus. Also preferred are fragments characterized by structural or functional attributes such as fragments that comprise alpha-helix and alpha-helix forming regions, beta-sheet and beta-sheet- forming regions, turn and tum-forming regions, coil and coil-forming regions, hydrophilic regions, hydrophobic regions, alpha amphipathic regions, beta amphipathic regions, flexible regions, surface- forming regions, substrate binding region, and high antigenic index regions. Other preferred fragments are biologically active fragments. Biologically active fragments are those that mediate CBFBGA09 activity, including those with a similar activity or an improved activity, or with a decreased undesirable activity. Also included are those that are antigenic or immunogenic in an animal, especially in a human. Preferably, all of these polypeptide fragments retain the biological activity of the CBFBGA09, including antigenic activity. Variants of the defined sequence and fragments also form part of the present invention. Preferred variants are those that vary from the referents by conservative amino acid substitutions — i.e., those that substitute a residue with another of like characteristics. Typical such substitutions are among Ala, Val, Leu and He; among Ser and Thr; among the acidic residues Asp and Glu; among Asn and Gb; and among the basic residues Lys and Arg; or aromatic residues Phe and Tyr. Particularly preferred are variants in which several, 5-10, 1-5, or 1-2 a ino acids are substituted, deleted, or added any combination.

The CBFBGA09 polypeptides of the vention can be prepared b any suitable manner. Such polypeptides bclude isolated naturally occurring polypeptides, recombinantly produced polypeptides, synthetically produced polypeptides, or polypeptides produced by a combination of these methods. Means for preparing such polypeptides are well understood b the art. Polynucleotides of the Invention

Another aspect of the bvention relates to CBFBGA09 polynucleotides. CBFBGA09 polynucleotides bclude isolated polynucleotides which encode the CBFBGA09 polypeptides and fragments, and polynucleotides closely related thereto. More specifically, CBFBGA09 polynucleotide of the bvention bclude a polynucleotide comprisbg the nucleotide sequence contabed b SEQ ID NO: 1 encoding a CBFBGA09 polypeptide of SEQ ID NO: 2, and polynucleotide having the particular sequence of SEQ ID NO: 1. CBFBGA09 polynucleotides further bclude a polynucleotide comprisbg a nucleotide sequence that has at least 80% identity over its entire length to a nucleotide sequence encodbg the CBFBGA09 polypeptide of SEQ ID NO:2, and a polynucleotide comprisbg a nucleotide sequence that is at least 80% identical to of SEQ ID NO: 1 over its entire length. In this regard, polynucleotides at least 90% identical are particularly preferred, and those with at least 95% are especially preferred. Furthermore, those with at least 97% are highly preferred and those with at least 98-99% are most highly preferred, with at least 99% bebg the most preferred. Also bcluded under CBFBGA09 polynucleotides are a nucleotide sequence which has sufficient identity to a nucleotide sequence contained in SEQ ID NO: 1 to hybridize under conditions useable for amplification or for use as a probe or marker. The invention also provides polynucleotides which are complementary to such CBFBGA09 polynucleotides.

CBFBGA09 of the bvention is structurally related to other proteins of the family, as shown by the results of sequencbg the cDNA of Table 1 (SEQ ID NO:l) encodbg human CBFBGA09. The cDNA sequence of SEQ ED NO: 1 contains an open readbg frame (nucleotide number 17 to 757) encodbg a polypeptide of 247 ambo acids of SEQ ID NO:2. The arnbo acid sequence of Table 2 (SEQ ID NO:2) has about 87.9% identity (usbg FASTA) b 247 ambo acid residues with hamster SL15 (F.E. Ware et al., J. Biol. Chem. 271:13935-13938,1996). The nucleotide sequence of Table 1 (SEQ ID NO: 1) has about 48% identity (usbg FASTA) b 1407 nucleotide residues with hamster SL15 (F.E. Ware et al., J. Biol. Chem. 271 : 13935-13938, 1996). Thus, CBFBGA09 polypeptides and polynucleotides of the present bvention are expected to have, bter alia, similar biological functions/properties to their homologous polypeptides and polynucleotides, and their utility is obvious to anyone skilled b the art.

Table l^a

GAGCTAGCTTTGCAATATGGCGGCCGAGGCGGACGGACCGCTTAAACGGCTGCTCGTGCCGATTCTTTT ACCTGAGAAATGCTACGACCAACTTTTCGTTCAGTGGGACTTGCTTCACGTCCCCTGCCTCAAGATTCT CCTCAGCAAAGGCCTGGGGCTGGGCATTGTGGCTGGCTCACTTCTAGTAAAGCTGCCCCAGGTGTTTAA AATCCGGGGAGCCAAGAGTGCTGAAGGGTTGAGTCTCCAGTCTGTAATGCTGGAGCTAGTGGCATTGAC TGGGACCATGGTCTACAGCATCACTAACAACTTCCCATTCAGCTCTTGGGGTGAAGCCTTATTCCTGAT GCTCCAGACGATCACCATCTGCTTCCTGGTCATGCACTACAGAGGACAGACTGTGAAAGGTGTCGCTTT CCTCGCTTGCTACGGCCTGGTCCTGCTGGTGCTTCTCTCACCTCTGACGCCCTTGACTGTAGTCACCCT GCTCCAGGCCTCCAATGTGCCTGCTGTGGTGGTGGGGAGGCTTCTCCAGGCAGCCACCAACTACCACAA CGGGTACACAGGCCAGCTCTCAGCCATCACAGTCTTCCTGCTGTTTGGGGGCTCCCTGGCCCGAATCTT CACTTCCATTCAGGAAACCGGAGATCCCCTGATGGCTGGGACCTTTGTGGTCTCCTCTCTCTGCAACGG CCTCATCGCCGCCCAGCTGCTCTTCTACTGGAATGCAAAGCCTCCCCACAAGCAGAAAAAGGCGCAGTA GAGCCAGCTACTGGAGTCATTCCGTTTCCACTCATTCACCCAACCTCAGGGTTCTCCCCATCTGAGCCA GCCTGCTGGTGTGACTTACTCATCCTCCATTCCTCTGCACTTGCAGACTTTCTGAGCCAGGGTTTTCTT TTAGTGGAAACAAATGGTTGATGGGATCCAGATCCTTTAGAAAAGGAGAGGATGGGGGTAGAGTCTCCC AAGCCAAAATTTTGACATTTGAGTGCTTTCGTAAGCCCTGTACATGTACTATTAATTCAGTCATTCAGC CAAGCCTCCTCCTCTAGCAGCAATTTCCAGCTGTTTAACACTATCCTGGGCAAATGTTTTACCCTGTCC TCCAGCCTCCCTGCTTCCCTTCTGGCCCTGGGGAAGACTTGAGTCTGGGACGGCCAGAGTTGGAGGGAC TGGGAGGCTGTGGCTGCCTCCCTCCCTCAGCCCGGCTGGGACTGTCTCCCGGACCCCAGTGCTGGGGTG GGGGAAGGGGGACGGAGAATGACTCAGGCAGGGCCCCAGGGTGGGATGAGGAGGTTCCTGCTCTGGCAG GTCTAGGCGGAAGGGAGTGGAGATGGGGCTGGTTGATCTGCTGCAGTGAGGGGAACAGATGGGACAATA AAGACTGGAGACTCAGTTGAATAATGC

A nucleotide sequence of a human CBFBGA09 (SEQ ID NO: 1).

Table 2^b

MAAEADGPLKRL VPI LPEKCYDQLFVQ D LHVPCLKILLSKG G GIVAGSLLVKLPQVFKIRGAK

SAEG SLQSVM E VA TGTMVYSIT-WFPFSS GEALFLMLQTITICPLV HYRGQTVKGVAF ACYG VLLVLLSP TPLTVV L QASIWPAVVVGRLLQAATNYHNGYTGQLSAITVFLLFGGSI-ARIFTSIQE

TGDP MAGTFWSSLCNGLIAAQL FYWNAKPP HKQKKAQ

An amino acid sequence of a human CBFBGA09 (SEQ ID NO: 2).

One polynucleotide of the present bvention encodbg CBFBGA09 may be obtabed usbg standard clonbg and screenbg, from a cDNA hbrary derived from mRNA b cells of human cord blood usbg the expressed sequence tag (EST) analysis (Adams, M.D., et al. Science (1991) 252:1651- 1656; Adams, M.D. et al, Nature, (1992) 355:632-634; Adams, M.D., et al, Nature (1995) 377 Supp:3-174). Polynucleotides of the bvention can also be obtabed from natural sources such as genomic DNA libraries or can be synthesized using well known and commercially available techniques. The nucleotide sequence encoding CBFBGA09 polypeptide of SEQ ID NO:2 may be identical to the polypeptide encoding sequence contained in Table 1 (nucleotide number 17 to 757 of SEQ ID NO: 1), or it may be a sequence, which as a result of the redundancy (degeneracy) of the genetic code, also encodes the polypeptide of SEQ ID NO:2. When the polynucleotides of the invention are used for the recombbant production of CBFBGA09 polypeptide, the polynucleotide may include the codbg sequence for the mature polypeptide or a fragment thereof, by itself; the codbg sequence for the mature polypeptide or fragment b readbg frame with other codbg sequences, such as those encodbg a leader or secretory sequence, a pre-, or pro- or prepro- proteb sequence, or other fusion peptide portions. For example, a marker sequence which facilitates purification of the fused polypeptide can be encoded. In certab preferred embodiments of this aspect of the bvention, the marker sequence is a hexa-histidbe peptide, as provided b the pQE vector (Qiagen, Inc.) and described b Gentz et al. , Proc Natl AcadSci USA (1989) 86:821- 824, or is an HA tag. The polynucleotide may also contab non-codbg 5 ' and 3 ' sequences, such as transcribed, non-translated sequences, splicbg and polyadenylation signals, ribosome bbdbg sites and sequences that stabilize mRNA.

Further preferred embodiments are polynucleotides encodbg CBFBGA09 variants comprise the ambo acid sequence CBFBGA09 polypeptide of Table 2 (SEQ ID NO:2) b which several, 5-10, 1-5, 1-3, 1-2 or 1 ambo acid residues are substituted, deleted or added, b any combination. The present bvention further relates to polynucleotides that hybridize to the hereb above- described sequences. In this regard, the present bvention especially relates to polynucleotides which hybridize under stringent conditions to the hereb above-described polynucleotides. As hereb used, the term "stringent conditions" means hybridization will occur only if there is at least 80%, and preferably at least 90%, and more preferably at least 95%, yet even more preferably 97-99% identity between the sequences.

Polynucleotides of the bvention, which are identical or sufficiently identical to a nucleotide sequence contabed b SEQ ID NO: 1 or a fragment thereof, may be used as hybridization probes for cDNA and genomic DNA, to isolate full-length cDNAs and genomic clones encodbg CBFBGA09 polypeptide and to isolate cDNA and genomic clones of other genes (bcludbg genes encodbg homologs and orthologs from species other than human) that have a high sequence similarity to the CBFBGA09 gene. Such hybridization techniques are known to those of skill b the art. Typically these nucleotide sequences are 80% identical, preferably 90% identical, more preferably 95% identical to that of the referent. The probes generally will comprise at least 15 nucleotides. Preferably, such probes will have at least 30 nucleotides and may have at least 50 nucleotides. Particularly preferred probes will range between 30 and 50 nucleotides.

In one embodiment, to obtab a polynucleotide encodbg CBFBGA09 polypeptide, bcludbg homologs and orthologs from species other than human, comprises the steps of screenbg an appropriate hbrary under stbgent hybridization conditions with a labeled probe havbg the SEQ ID NO: 1 or a fragment thereof; and isolating full-length cDNA and genomic clones containbg said polynucleotide sequence. Thus b another aspect, CBFBGA09 polynucleotides of the present bvention further bclude a nucleotide sequence comprisbg a nucleotide sequence that hybridize under stringent condition to a nucleotide sequence havbg SEQ ID NO: 1 or a fragment thereof. Also bcluded with CBFBGA09 polypeptides are polypeptide comprisbg ambo acid sequence encoded by nucleotide sequence obtabed by the above hybridization condition. Such hybridization techniques are well known to those of skill b the art. Stringent hybridization conditions are as defined above or, alternatively, conditions under overnight cubation at 42°C b a solution comprisbg: 50% formamide, 5xSSC (150mM NaCl, 15mM trisodium citrate), 50 mM sodium phosphate (pH7.6), 5x Denhardt's solution, 10 % dextran sulfate, and 20 microgram/ml denatured, sheared salmon sperm DNA, followed by washbg the filters b 0. lx SSC at about 65°C.

The polynucleotides and polypeptides of the present bvention may be employed as research reagents and materials for discovery of treatments and diagnostics to animal and human disease.

Vectors, Host Cells, Expression The present bvention also relates to vectors which comprise a polynucleotide or polynucleotides of the present bvention, and host cells which are genetically engbeered with vectors of the bvention and to the production of polypeptides of the bvention by recombinant techniques. Cell-free translation systems can also be employed to produce such protebs usbg RNAs derived from the DNA constructs of the present bvention. For recombinant production, host cells can be genetically engbeered to bcorporate expression systems or portions thereof for polynucleotides of the present bvention. Introduction of polynucleotides bto host cells can be effected by methods described b many standard laboratory manuals, such as Davis et al. , BASIC METHODS IN MOLECULAR BIOLOGY (1986) and Sambrook et al., MOLECULAR CLONING: A LABORATORY MANUAL, 2nd Ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1989) such as calcium phosphate transfection, DEAE-dextran mediated transfection, transvection, microbjection, cationic lipid-mediated transfection, electroporation, transduction, scrape loading, ballistic btroduction or infection.

Representative examples of appropriate hosts bclude bacterial cells, such as streptococci, staphylococci, E. coli, Streptomyces and Bacillus subtilis cells; fungal cells, such as yeast cells and Aspergillus cells; insect cells such as Drosophila S2 and Spodoptera Sf9 cells; animal cells such as CHO, COS, HeLa, C127, 3T3, BHK, HEK 293 and Bowes melanoma cells; and plant cells.

A great variety of expression systems can be used. Such systems bclude, among others, chromosomal, episomal and virus-derived systems, e.g., vectors derived from bacterial plasmids, from bacteriophage, from transposons, from yeast episomes, from insertion elements, from yeast chromosomal elements, from viruses such as baculoviruses, papova viruses, such as SV40, vaccinia viruses, adenoviruses, fowl pox viruses, pseudorabies viruses and retroviruses, and vectors derived from combinations thereof, such as those derived from plasmid and bacteriophage genetic elements, such as cosmids and phagemids. The expression systems may contab control regions that regulate as well as engender expression. Generally, any system or vector suitable to mabtab, propagate or express polynucleotides to produce a polypeptide b a host may be used. The appropriate nucleotide sequence may be inserted bto an expression system by any of a variety of well-known and routine techniques, such as, for example, those set forth b Sambrook et al., MOLECULAR CLONING, A LABORATORY MANUAL (supra). For secretion of the translated proteb bto the lumen of the endoplasmic reticulum, bto the periplasmic space or bto the extracellular environment, appropriate secretion signals may be bcorporated bto the desired polypeptide. These signals may be endogenous to the polypeptide or they may be heterologous signals.

If the CBFBGA09 polypeptide is to be expressed for use b screenbg assays, generally, it is preferred that the polypeptide be produced at the surface of the cell. In this event, the cells may be harvested prior to use in the screening assay. If CBFBGA09 polypeptide is secreted into the medium, the medium can be recovered in order to recover and purify the polypeptide; if produced intracellularly, the cells must first be lysed before the polypeptide is recovered. CBFBGA09 polypeptides can be recovered and purified from recombinant cell cultures by well-known methods bcludbg ammonium sulfate or ethanol precipitation, acid extraction, anion or cation exchange chromatography, phosphocellulose chromatography, hydrophobic bteraction chromatography, affinity chromatography, hydroxylapatite chromatography and lectin chromatography. Most preferably, high performance liquid chromatography is employed for purification. Well known techniques for refolding proteins may be employed to regenerate active conformation when the polypeptide is denatured during isolation and or purification.

Diagnostic Assays

This bvention also relates to the use of CBFBGA09 polynucleotides for use as diagnostic reagents. Detection of a mutated form of CBFBGA09 gene associated with a dysfunction will provide a diagnostic tool that can add to or define a diagnosis of a disease or susceptibility to a disease which results from under-expression, over-expression or altered expression of CBFBGA09. Individuals carrybg mutations b the CBFBGA09 gene may be detected at the DNA level by a variety of techniques.

Nucleic acids for diagnosis may be obtabed from a subject's cells, such as from blood, urine, saliva, tissue biopsy or autopsy material. The genomic DNA may be used directly for detection or may be amplified enzymatically by usbg PCR or other amplification techniques prior to analysis. RNA or cDNA may also be used b similar fashion. Deletions and insertions can be detected by a change b size of the amplified product b comparison to the normal genotype. Pobt mutations can be identified by hybridizbg amplified DNA to labeled CBFBGA09 nucleotide sequences. Perfectly matched sequences can be distinguished from mismatched duplexes by RNase digestion or by differences b melting temperatures. DNA sequence differences may also be detected by alterations b electrophoretic mobility of DNA fragments b gels, with or without denaturing agents, or by direct DNA sequencbg. See, e.g., Myers et al. , Science (1985) 230: 1242. Sequence changes at specific locations may also be revealed by nuclease protection assays, such as RNase and S 1 protection or the chemical cleavage method. See Cotton et al , Proc Natl Acad Sci USA ( 1985) 85 : 4397-4401. In another embodiment, an array of oligonucleotides probes comprisbg CBFBGA09 nucleotide sequence or fragments thereof can be constructed to conduct efficient screenbg of e.g., genetic mutations. Array technology methods are well known and have general applicability and can be used to address a variety of questions b molecular genetics bcludbg gene expression, genetic linkage, and genetic variability. (See for example: M.Chee et al., Science, Vol 274, pp 610-613 (1996)).

The diagnostic assays offer a process for diagnosbg or determbbg a susceptibility to metabolic disorders through detection of mutation b the CBFBGA09 gene by the methods described.

In addition, metabolic disorders can be diagnosed by methods comprisbg determbbg from a sample derived from a subject an abnormally decreased or increased level of CBFBGA09 polypeptide or CBFBGA09 mRNA. Decreased or increased expression can be measured at the RNA level using any of the methods well known in the art for the quantitation of polynucleotides, such as, for example, PCR, RT-PCR, RNase protection, Northern blotting and other hybridization methods. Assay techniques that can be used to deterrnbe levels of a protein, such as an CBFBGA09 polypeptide, b a sample derived from a host are well-known to those of skill b the art. Such assay methods bclude radiobimunoassays, competitive-bbdbg assays, Western Blot analysis and ELISA assays.

Thus in another aspect, the present bvention relates to a diagonostic kit for a disease or suspectability to a disease, particularly metabolic disorders, which comprises:

(a) a CBFBGA09 polynucleotide, preferably the nucleotide sequence of SEQ ID NO: 1, or a fragment thereof ;

(b) a nucleotide sequence complementary to that of (a);

(c) a CBFBGA09 polypeptide, preferably the polypeptide of SEQ ID NO: 2, or a fragment thereof; or

(d) an antibody to a CBFBGA09 polypeptide, preferably to the polypeptide of SEQ ID NO: 2. It will be appreciated that in any such kit, (a), (b), (c) or (d) may comprise a substantial component.

Chromosome Assays The nucleotide sequences of the present bvention are also valuable for chromosome identification. The sequence is specifically targeted to and can hybridize with a particular location on an bdividual human chromosome. The mappbg of relevant sequences to chromosomes according to the present bvention is an important first step b correlating those sequences with gene associated disease. Once a sequence has been mapped to a precise chromosomal location, the physical position of the sequence on the chromosome can be correlated with genetic map data. Such data are found, for example, b V. McKusick, Mendelian Inheritance b Man (available on lbe through Johns Hopkins University Welch Medical Library). The relationship between genes and diseases that have been mapped to the same chromosomal region are then identified through linkage analysis (cobheritance of physically adjacent genes). The differences in the cDNA or genomic sequence between affected and unaffected individuals can also be determined. If a mutation is observed in some or all of the affected individuals but not in any normal individuals, then the mutation is likely to be the causative agent of the disease.

Antibodies

The polypeptides of the bvention or their fragments or analogs thereof, or cells expressbg them can also be used as immunogens to produce antibodies immunospecific for the CBFBGA09 polypeptides. The term "immunospecific" means that the antibodies have substantiall greater affinity for the polypeptides of the bvention than their affinity for other related polypeptides b the prior art.

Antibodies generated against the CBFBGA09 polypeptides can be obtabed by administering the polypeptides or epitope-bearing fragments, analogs or cells to an animal, preferably a nonhuman, using routine protocols. For preparation of monoclonal antibodies, any technique which provides antibodies produced by continuous cell lbe cultures can be used. Examples bclude the hybridoma technique (Kohler, G. and Milstein, C, Nature (1975) 256:495-497), the trioma technique, the human B-cell hybridoma technique (Kozbor et al. , Immunology Today (1983) 4:72) and the EBV-hybridoma technique (Cole et al, MONOCLONAL ANTIBODIES AND CANCER THERAPY, pp. 77-96, Alan R. Liss, Inc., 1985). Techniques for the production of sbgle chab antibodies (U.S. Patent No. 4,946,778) can also be adapted to produce sbgle chab antibodies to polypeptides of this bvention. Also, transgenic mice, or other organisms bcludbg other mammals, may be used to express humanized antibodies.

The above-described antibodies may be employed to isolate or to identify clones expressbg the polypeptide or to purify the polypeptides by affinity chromatography.

Antibodies against CBFBGA09 polypeptides may also be employed to treat metabolic disorders, among others.

Vaccines Another aspect of the invention relates to a method for inducing an immunological response in a mammal which comprises inoculating the mammal with CBFBGA09 polypeptide, or a fragment thereof, adequate to produce antibody and/or T cell immune response to protect said animal from metabolic disorders, among others. Yet another aspect of the invention relates to a method of inducing immunological response in a mammal which comprises, delivering CBFBGA09 polypeptide via a vector directing expression of CBFBGA09 polynucleotide in vivo in order to induce such an immunological response to produce antibody to protect said animal from diseases.

Further aspect of the invention relates to an immunological/vaccbe formulation (composition) which, when introduced into a mammalian host, induces an immunological response in that mammal to a CBFBGA09 polypeptide wherein the composition comprises a CBFBGA09 polypeptide or CBFBGA09 gene. The vaccine formulation may further comprise a suitable carrier. S ce CBFBGA09 polypeptide may be broken down in the stomach, it is preferably administered parenterally (bcludbg subcutaneous, intramuscular, intravenous, intradermal etc. bjection). Formulations suitable for parenteral admbistration include aqueous and non-aqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation instonic with the blood of the recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents or thickening agents. The formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampoules and vials and may be stored b a freeze-dried condition requiring only the addition of the sterile liquid carrier immediately prior to use. The vaccine formulation may also include adjuvant systems for enhancing the immunogenicity of the formulation, such as oil-in water systems and other systems known the art. The dosage will depend on the specific activity of the vaccine and can be readily determined by routine experimentation. Screening Assays

The CBFBGA09 polypeptide of the present bvention may be employed b a screenbg process for compounds which activate (agonists) or inhibit activation of (antagonists, or otherwise called inhibitors) the CBFBGA09 polypeptide of the present bvention. Thus, polypeptides of the bvention may also be used to assess identify agonist or antagonists from, for example, cells, cell-free preparations, chemical libraries, and natural product mixtures. These agonists or antagonists may be natural or modified substrates, ligands, receptors, enzymes, etc., as the case may be, of the polypeptide of the present bvention; or may be structural or functional mimetics of the polypeptide of the present bvention. See Coligan eta , Current Protocols in Immunology l(2):Chapter 5 (1991). CBFBGA09 polypeptides are responsible for many biological functions, bcludbg many pathologies.

Accordingly, it is desirous to find compounds and drugs which stimulate CBFBGA09 polypeptide on the one hand and which can inhibit the function of CBFBGA09 polypeptide on the other hand. In general, agonists are employed for therapeutic and prophylactic purposes for such conditions as metabolic disorders. Antagonists may be employed for a variety of therapeutic and prophylactic purposes for such conditions as metabolic disorders.

In general, such screenbg procedures may bvolve usbg appropriate cells which express the CBFBGA09 polypeptide or respond to CBFBGA09 polypeptide of the present bvention. Such cells bclude cells from mammals, yeast, Drosophila or E. coli. Cells which express the CBFBGA09 polypeptide (or cell membrane contabbg the expressed polypeptide) or respond to CBFBGA09 polypeptide are then contacted with a test compound to observe bbdbg, or stimulation or inhibition of a functional response. The ability of the cells which were contacted with the candidate compounds is compared with the same cells which were not contacted for CBFBGA09 activity.

The assays may simply test bbdbg of a candidate compound wherein adherence to the cells bearing the CBFBGA09 polypeptide is detected by means of a label directly or bdirectly associated with the candidate compound or in an assay volvbg competition with a labeled competitor. Further, these assays may test whether the candidate compound results in a signal generated by activation of the CBFBGA09 polypeptide, using detection systems appropriate to the cells bearing the CBFBGA09 polypeptide. Inhibitors of activation are generally assayed b the presence of a known agonist and the effect on activation by the agonist by the presence of the candidate compound is observed.

Further, the assays may simply comprise the steps of mixbg a candidate compound with a solution contabbg a CBFBGA09 polypeptide to form a mixture, measuring CBFBGA09 activity b the mixture, and comparing the CBFBGA09 activity of the mixture to a standard. The CBFBGA09 cDNA, protein and antibodies to the protein may also be used to configure assays for detectbg the effect of added compounds on the production of CBFBGA09 mRNA and protein in cells. For example, an ELISA may be constructed for measuring secreted or cell associated levels of CBFBGA09 proteb using monoclonal and polyclonal antibodies by standard methods known in the art, and this can be used to discover agents which may inhibit or enhance the production of CBFBGA09 (also called antagonist or agonist, respectively) from suitably manipulated cells or tissues.

The CBFBGA09 protein may be used to identify membrane bound or soluble receptors, if any, through standard receptor binding techniques known in the art. These include, but are not limited to, ligand binding and crosslinking assays in which the CBFBGA09 is labeled with a radioactive isotope (eg 1251), chemically modified (eg biotbylated), or fused to a peptide sequence suitable for detection or purification, and incubated with a source of the putative receptor (cells, cell membranes, cell supernatants, tissue extracts, bodily fluids). Other methods include biophysical techniques such as surface plasmon resonance and spectroscopy. In addition to bebg used for purification and cloning of the receptor, these binding assays can be used to identify agonists and antagonists of CBFBGA09 which compete with the bind g of CBFBGA09 to its receptors, if any. Standard methods for conducting screenbg assays are well understood in the art. Examples of potential CBFBGA09 polypeptide antagonists bclude antibodies or, b some cases, oligonucleotides or proteins which are closely related to the ligands, substrates, receptors, enzymes, etc., as the case may be, of the CBFBGA09 polypeptide, e.g., a fragment of the ligands, substrates, receptors, enzymes, etc.; or small molecules which bbd to the polypetide of the present bvention but do not elicit a response, so that the activity of the polypeptide is prevented.

Thus m another aspect, the present invention relates to a screening kit for identifying agonists, antagonists, ligands, receptors, substrates, enzymes, etc. for CBFBGA09 polypeptides; or compounds which decrease or enhance the production of CBFBGA09 polypeptides, which comprises:

(a) a CBFBGA09 polypeptide, preferably that of SEQ ID NO:2;

(b) a recombbant cell expressing a CBFBGA09 polypeptide, preferably that of SEQ ID NO:2;

(c) a cell membrane expressing a CBFBGA09 polypeptide; preferably that of SEQ ID NO: 2; or (d) antibody to a CBFBGA09 polypeptide, preferably that of SEQ ID NO: 2.

It will be appreciated that in any such kit, (a), (b), (c) or (d) may comprise a substantial component. Prophylactic and Therapeutic Methods

This bvention provides methods of treating abnormal conditions such as metabolic disorders related to both an excess of and insufficient amounts of CBFBGA09 polypeptide activity.

If the activity of CBFBGA09 polypeptide is b excess, several approaches are available. One approach comprises administering to a subject an inhibitor compound (antagonist) as herebabove described along with a pharmaceutically acceptable carrier b an amount effective to inhibit the function of the CBFBGA09 polypeptide, such as, for example, by blockbg the bbdbg of ligands, substrates, receptors, enzymes, etc., or by bhibitbg a second signal, and thereby alleviating the abnormal condition. In another approach, soluble forms of CBFBGA09 polypeptides still capable of bbd g the ligand, substrate, enzymes, receptors, etc. in competition with endogenous CBFBGA09 polypeptide may be administered. Typical embodiments of such competitors comprise fragments of the CBFBGA09 polypeptide.

In still another approach, expression of the gene encoding endogenous CBFBGA09 polypeptide can be inhibited using expression blockbg techniques. Known such techniques involve the use of antisense sequences, either internally generated or separately administered. See, for example, O'Connor, J Neurochem (1991) 56:560 b Oligodeoxynucleotides as Antisense Inhibitors of Gene Expression. CRC Press, Boca Raton, FL (1988). Alternatively, oligonucleotides which form triple helices with the gene can be supplied. See, for example, Lee et al, Nucleic Acids Res (1979) 6:3073; Cooney et al, Science (1988) 241:456; Dervan et al, Science (1991) 251:1360. These oligomers can be administered per se or the relevant oligomers can be expressed in vivo.

For treating abnormal conditions related to an under-expression of CBFBGA09 and its activity, several approaches are also available. One approach comprises administering to a subject a therapeutically effective amount of a compound which activates CBFBGA09 polypeptide, i.e., an agonist as described above, b combbation with a pharmaceutically acceptable carrier, to thereby alleviate the abnormal condition. Alternatively, gene therapy may be employed to effect the endogenous production of CBFBGA09 by the relevant cells in the subject. For example, a polynucleotide of the bvention may be engbeered for expression b a replication defective retroviral vector, as discussed above. The refroviral expression construct may then be isolated and btroduced bto a packagbg cell transduced with a retroviral plasmid vector contabbg RNA encodbg a polypeptide of the present bvention such that the packagbg cell now produces infectious viral particles containbg the gene of bterest. These producer cells may be administered to a subject for engbeering cells in vivo and expression of the polypeptide in vivo. For overview of gene therapy, see Chapter 20, Gene Therapy and other Molecular Genetic-based Therapeutic Approaches, (and references cited thereb) b Human Molecular Genetics, T Strachan and A P Read, BIOS Scientific Publishers Ltd (1996). Another approach is to administer a therapeutic amount of CBFBGA09 polypeptides b combination with a suitable pharmaceutical carrier.

Formulation and Administration Peptides, such as the soluble form of CBFBGA09 polypeptides, and agonists and antagonist peptides or small molecules, may be formulated b combination with a suitable pharmaceutical carrier. Such formulations comprise a therapeutically effective amount of the polypeptide or compound, and a pharmaceutically acceptable carrier or excipient. Such carriers bclude but are not limited to, salbe, buffered salbe, dextrose, water, glycerol, ethanol, and combinations thereof. Formulation should suit the mode of administration, and is well withb the skill of the art. The bvention further relates to pharmaceutical packs and kits comprisbg one or more contabers filled with one or more of the bgredients of the aforementioned compositions of the bvention.

Polypeptides and other compounds of the present bvention may be employed alone or b conjunction with other compounds, such as therapeutic compounds. Preferred forms of systemic administration of the pharmaceutical compositions bclude bjection, typically by btravenous bjection. Other bjection routes, such as subcutaneous, btramuscular, or btraperitoneal, can be used. Alternative means for systemic administration bclude transmucosal and transdermal administration usbg penetrants such as bile salts or fusidic acids or other detergents. In addition, if properly formulated b enteric or encapsulated formulations, oral -idministration may also be possible. Administration of these compounds may also be topical and/or localized, b the form of salves, pastes, gels and the like.

The dosage range required depends on the choice of peptide, the route of administration, the nature of the formulation, the nature of the subject's condition, and the judgment of the attending practitioner. Suitable dosages, however, are b the range of 0.1-100 μg/kg of subject. Wide variations b the needed dosage, however, are to be expected b view of the variety of compounds available and the differing efficiencies of various routes of administration. For example, oral administration would be expected to require higher dosages than administration by btravenous bjection. Variations b these dosage levels can be adjusted usbg standard empirical routines for optimization, as is well understood b the art. Polypeptides used b treatment can also be generated endogenously b the subject, b treatment modalities often referred to as "gene therapy" as described above. Thus, for example, cells from a subject may be engbeered with a polynucleotide, such as a DNA or RNA, to encode a polypeptide ex vivo, and for example, by the use of a retroviral plasmid vector. The cells are then btroduced bto the subject. All publications, includbg but not limited to patents and patent applications, cited in this specification are hereb bcorporated by reference as if each mdividual publication were specifically and individually bdicated to be incorporated by reference hereb as though fully set forth.

SEQUENCE LISTING

(1) GENERAL INFORMATION

(i) APPLICANT: MAO, MAO

WANG, YA-XIN

(ii) TITLE OF THE INVENTION: CBFBGA09: A HUMAN SL15 HOMOLOG

(iii) NUMBER OF SEQUENCES: 2

(iv) CORRESPONDENCE ADDRESS: (A) ADDRESSEE: RATNER & PRESTIA

(B) STREET: P.O. BOX 980

(C) CITY: VALLEY FORGE

(D) STATE: PA

(E) COUNTRY: USA (F) ZIP: 19482

(v) COMPUTER READABLE FORM:

(A) MEDIUM TYPE: Diskette

(B) COMPUTER: IBM Compatible (C) OPERATING SYSTEM: DOS

(D) SOFTWARE: FastSEQ for Windows Version 2.0

(vi) CURRENT APPLICATION DATA:

(A) APPLICATION NUMBER: TO BE ASSIGNED (B) FILING DATE:

(C) CLASSIFICATION: UNKNOWN

(vii) PRIOR APPLICATION DATA: (A) APPLICATION NUMBER: (B) FILING DATE:

(viii) ATTORNEY/AGENT INFORMATION: (A) NAME: PRESTIA, PAUL F

(B) REGISTRATION NUMBER: 23,031

(C) REFERENCE/DOCKET NUMBER: GP-70311

(ix) TELECOMMUNICATION INFORMATION: (A) TELEPHONE: 610-407-0700

(B) TELEFAX: 610-407-0701

(C) TELEX: 846169

(2) INFORMATION FOR SEQ ID NO:l:

(i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 1407 base pairs (B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear (ii) MOLECULE TYPE: cDNA

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:l:

GAGCTAGCTT TGCAATATGG CGGCCGAGGC GGACGGACCG CTTAAACGGC TGCTCGTGCC 60

GATTCTTTTA CCTGAGAAAT GCTACGACCA ACTTTTCGTT CAGTGGGACT TGCTTCACGT 120

CCCCTGCCTC AAGATTCTCC TCAGCAAAGG CCTGGGGCTG GGCATTGTGG CTGGCTCACT 180 TCTAGTAAAG CTGCCCCAGG TGTTTAAAAT CCGGGGAGCC AAGAGTGCTG AAGGGTTGAG 240

TCTCCAGTCT GTAATGCTGG AGCTAGTGGC ATTGACTGGG ACCATGGTCT ACAGCATCAC 300

TAACAACTTC CCATTCAGCT CTTGGGGTGA AGCCTTATTC CTGATGCTCC AGACGATCAC 360

CATCTGCTTC CTGGTCATGC ACTACAGAGG ACAGACTGTG AAAGGTGTCG CTTTCCTCGC 420

TTGCTACGGC CTGGTCCTGC TGGTGCTTCT CTCACCTCTG ACGCCCTTGA CTGTAGTCAC 480 CCTGCTCCAG GCCTCCAATG TGCCTGCTGT GGTGGTGGGG AGGCTTCTCC AGGCAGCCAC 540

CAACTACCAC AACGGGTACA CAGGCCAGCT CTCAGCCATC ACAGTCTTCC TGCTGTTTGG 600

GGGCTCCCTG GCCCGAATCT TCACTTCCAT TCAGGAAACC GGAGATCCCC TGATGGCTGG 660

GACCTTTGTG GTCTCCTCTC TCTGCAACGG CCTCATCGCC GCCCAGCTGC TCTTCTACTG 720

GAATGCAAAG CCTCCCCACA AGCAGAAAAA GGCGCAGTAG AGCCAGCTAC TGGAGTCATT 780 CCGTTTCCAC TCATTCACCC AACCTCAGGG TTCTCCCCAT CTGAGCCAGC CTGCTGGTGT 840

GACTTACTCA TCCTCCATTC CTCTGCACTT GCAGACTTTC TGAGCCAGGG TTTTCTTTTA 900

GTGGAAACAA ATGGTTGATG GGATCCAGAT CCTTTAGAAA AGGAGAGGAT GGGGGTAGAG 960

TCTCCCAAGC CAAAATTTTG ACATTTGAGT GCTTTCGTAA GCCCTGTACA TGTACTATTA 1020

ATTCAGTCAT TCAGCCAAGC CTCCTCCTCT AGCAGCAATT TCCAGCTGTT TAACACTATC 1080 CTGGGCAAAT GTTTTACCCT GTCCTCCAGC CTCCCTGCTT CCCTTCTGGC CCTGGGGAAG 1140

ACTTGAGTCT GGGACGGCCA GAGTTGGAGG GACTGGGAGG CTGTGGCTGC CTCCCTCCCT 1200

CAGCCCGGCT GGGACTGTCT CCCGGACCCC AGTGCTGGGG TGGGGGAAGG GGGACGGAGA 1260

ATGACTCAGG CAGGGCCCCA GGGTGGGATG AGGAGGTTCC TGCTCTGGCA GGTCTAGGCG 1320

GAAGGGAGTG GAGATGGGGC TGGTTGATCT GCTGCAGTGA GGGGAACAGA TGGGACAATA 1380 AAGACTGGAG ACTCAGTTGA ATAATGC 1407

(2) INFORMATION FOR SEQ ID NO: 2:

(i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 247 amino acids

(B) TYPE: amino acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear (ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 2:

Met Ala Ala Glu Ala Asp Gly Pro Leu Lys Arg Leu Leu Val Pro lie 1 5 10 15

Leu Leu Pro Glu Lys Cys Tyr Asp Gin Leu Phe Val Gin Trp Asp Leu

20 25 30

Leu His Val Pro Cys Leu Lys lie Leu Leu Ser Lys Gly Leu Gly Leu 35 40 45 Gly He Val Ala Gly Ser Leu Leu Val Lys Leu Pro Gin Val Phe Lys 50 55 60

He Arg Gly Ala Lys Ser Ala Glu Gly Leu Ser Leu Gin Ser Val Met 65 70 75 80

Leu Glu Leu Val Ala Leu Thr Gly Thr Met Val Tyr Ser He Thr Asn 85 90 95

Asn Phe Pro Phe Ser Ser Trp Gly Glu Ala Leu Phe Leu Met Leu Gin

100 105 110

Thr He Thr He Cys Phe Leu Val Met His Tyr Arg Gly Gin Thr Val 115 120 125 Lys Gly Val Ala Phe Leu Ala Cys Tyr Gly Leu Val Leu Leu Val Leu 130 135 140

Leu Ser Pro Leu Thr Pro Leu Thr Val Val Thr Leu Leu Gin Ala Ser 145 150 155 160

Asn Val Pro Ala Val Val Val Gly Arg Leu Leu Gin Ala Ala Thr Asn 165 170 175

Tyr His Asn Gly Tyr Thr Gly Gin Leu Ser Ala He Thr Val Phe Leu

180 185 190

Leu Phe Gly Gly Ser Leu Ala Arg He Phe Thr Ser He Gin Glu Thr 195 200 205 Gly Asp Pro Leu Met Ala Gly Thr Phe Val Val Ser Ser Leu Cys Asn 210 215 220

Gly Leu He Ala Ala Gin Leu Leu Phe Tyr Trp Asn Ala Lys Pro Pro 225 230 235 240

His Lys Gin Lys Lys Ala Gin 245

Claims (1)

1. What is claimed is:

   1. An isolated polynucleotide comprisbg a nucleotide sequence that has at least 80% identity over its entire length to a nucleotide sequence encodbg the CBFBGA09 polypeptide of SEQ ID NO:2; or a nucleotide sequence complementary to said isolated polynucleotide.

   2. The polynucleotide of claim 1 wherein said polynucleotide comprises the nucleotide sequence contained in SEQ ID NO: 1 encodbg the CBFBGA09 polypeptide of SEQ ED N02.

   3. The polynucleotide of claim 1 wherein said polynucleotide comprises a nucleotide sequence that is at least 80% identical to that of SEQ ID NO: 1 over its entire length.

   4. The polynucleotide of claim 3 which is polynucleotide of SEQ ID NO: 1.

   5. The polynucleotide of claim 1 which is DNA or RNA.

   6. A DNA or RNA molecule comprising an expression system, wherein said expression system is capable of producbg a CBFBGA09 polypeptide comprisbg an ambo acid sequence, which has at least 88% identity with the polypeptide of SEQ ID NO:2 when said expression system is present b a compatible host cell.

   7. A host cell comprising the expression system of claim 6.

   8. A process for producing a CBFBGA09 polypeptide comprisbg culturing a host of claim 7 under conditions sufficient for the production of said polypeptide and recovering the polypeptide from the culture.

   9. A process for producbg a cell which produces a CBFBGA09 polypeptide thereof comprisbg transforming or transfecting a host cell with the expression system of claim 6 such that the host cell, under appropriate culture conditions, produces a CBFBGA09 polypeptide.

   10. A CBFBGA09 polypeptide comprising an ambo acid sequence which is at least 88% identical to the amino acid sequence of SEQ ID NO:2 over its entire length.

   11. The polypeptide of claim 10 which comprises the amino acid sequence of SEQ ID NO:2.

   12. An antibody immunospecific for the CBFBGA09 polypeptide of claim 10.

   13. A method for the treatment of a subject in need of enhanced activity or expression of CBFBGA09 polypeptide of claim 10 comprising:

   (a) administering to the subject a therapeutically effective amount of an agonist to said polypeptide; and/or

   (b) providbg to the subject an isolated polynucleotide comprisbg a nucleotide sequence that has at least 80% identity to a nucleotide sequence encodbg the CBFBGA09 polypeptide of SEQ ED NO:2 over its entire length; or a nucleotide sequence complementary to said nucleotide sequence b a form so as to effect production of said polypeptide activity in vivo.

   14. A method for the treatment of a subject havbg need to inhibit activity or expression of CBFBGA09 polypeptide of claim 10 comprising: (a) administering to the subject a therapeutically effective amount of an antagonist to said polypeptide; and/or

   (b) administering to the subject a nucleic acid molecule that inhibits the expression of the nucleotide sequence encoding said polypeptide; and/or

   (c) admbistering to the subject a therapeutically effective amount of a polypeptide that competes with said polypeptide for its ligand, substrate , or receptor.

   15. A process for diagnos g a disease or a susceptibility to a disease in a subject related to expression or activity of CBFBGA09 polypeptide of claim 10 b a subject comprisbg:

   (a) determining the presence or absence of a mutation in the nucleotide sequence encodbg said CBFBGA09 polypeptide in the genome of said subject; and/or

   (b) analyzing for the presence or amount of the CBFBGA09 polypeptide expression in a sample derived from said subject.

   16. A method for identifying compounds which inhibit (antagonize) or agonize the CBFBGA09 polypeptide of claim 10 which comprises:

   (a) contacting a candidate compound with cells which express the CBFBGA09 polypeptide (or cell membrane expressbg CBFBGA09 polypeptide) or respond to CBFBGA09 polypeptide; and

   (b) observbg the bbdbg, or stimulation or inhibition of a functional response; or comparing the ability of the cells (or cell membrane) which were contacted with the candidate compounds with the same cells which were not contacted for CBFBGA09 polypeptide activity.

   17. An agonist identified by the method of claim 16.

   18. An antagonist identified by the method of claim 16.

   19. A recomb ant host cell produced by a method of Claim 9 or a membrane thereof expressing a CBFBGA09 polypeptide.