The present invention is directed to the MMSC2 gene, its protein product and the use of the protein to (i) detect mutant MMAC1 proteins, (ii) screen for drugs which can be used for suppressing tumor growth and (iii) identify proteins which interact with the MMAC1 gene or are involved in the tumor suppression pathway of the MMAC1 gene.

BACKGROUND OF THE INVENTION
 The present invention is directed to the MMSC2 gene, its protein product
 and the use of the protein to (i) detect mutant MMAC1 proteins, (ii)
 screen for drugs which can be used for suppressing tumor growth and (iii)
 identify proteins which interact with the MMAC1 gene or are involved in
 the tumor suppression pathway of the MMAC1 gene.
 The publications and other materials used herein to illuminate the
 background of the invention or provide additional details respecting the
 practice, are incorporated by reference, and for convenience are
 respectively grouped in the appended List of References.
 A number of genetic alterations are involved in the oncogenesis of
 glioblastoma multiforme, including inactivation of p53, p16, RB,
 amplification of the gene encoding epidermal growth factor receptor and
 several other molecular alterations (Louis & Gusella, 1995). However the
 most common genetic alteration is the deletion of large regions or an
 entire copy of chromosome 10 (Fults et al., 1990; Rahseed et al., 1992).
 Recently, the tumor suppressor gene MMAC1 (Steck et al., 1997), also known
 as PTEN (Li et al., 1997) or TEP1 (Li & Sun, 1997) was mapped to 10q23 and
 shown to be mutated in 17-24% of xenografited and primary glioblastomas,
 14% of breast cancer samples and 25% of kidney carcinomas (Steck et al.,
 1997). The mutation frequency in established cell lines of these tumor
 types is somewhat higher. In addition to this predicted involvement in
 sporadic cancer, germ-line MMAC1 mutations have been detected in two
 autosomal dominant disorders, Cowden disease (Nelen et al., 1997; Liaw et
 al., 1997), a syndrome that confers an elevated risk for tumors of breast,
 thyroid and skin, and Bannayan-Zonana syndrome (Marsh et al., 1997), a
 condition characterized by macrocephaly, lipomas, intestinal hamartomatous
 polyps, vascular malformations and some skin disorders. Mutations of MMAC1
 in primary endometrial carcinomas (Kong et al., 1997) and in juvenile
 polyposis coli (Olschwang et al., 1998) have also been seen.
 The predicted protein product of the MMAC1 gene has several regions of
 homology with other proteins. The MMAC1 protein has an amino terminal
 domain with extensive homology to tensin, a protein that interacts with
 actin filaments at focal adhesions, and with auxilin, a protein involved
 in synaptic vesicle transport. The MMAC1 protein also has a region with
 extensive homology to protein tyrosine phosphatases (Steck et al., 1997;
 Li et al., 1997). Mutations of MMAC1 in tumors, its cytoplasmic
 localization (Li & Sun, 1997) and its intrinsic phosphatase activity (Li &
 Sun, 1997; Myers et al., 1997) suggested that its activity could be
 important in some aspect of tumor progression, possibly to counteract the
 oncogenic effect of a specific protein tyrosine kinase. In addition, MMAC1
 is rapidly down-regulated by TGF.beta. in cells sensitive to its cell
 growth and cell adhesion regulatory properties (Li & Sun, 1997).
 Experiments on glioma cell growth have shown that MMAC1 is a protein
 phosphatase that exhibits functional and specific growth-suppressing
 activity. In such experiments, the introduction of HA-tagged MMAC1 into
 glioma cells containing endogenous mutant alleles caused growth
 suppression, but was without effect in cells containing wild-type MMAC1
 (Furnari et al., 1997). The ectopic expression of MMAC1alleles, which
 carried mutations found in primary tumors and have been shown or are
 expected to inactivate its phosphatase activity, caused little growth
 suppression (Furnari et al., 1997). Although these activities of MMAC1 are
 known, the mechanisms of tumor suppression by MMAC1 and the interaction of
 the MMAC1 protein with other proteins are not well understood.
 Many cytosolic signaling proteins and cytoskeletal proteins are composed of
 modular units of small protein-protein interactive domains that allow
 reversible and regulated assembly into larger protein complexes. These
 domains include the Src-homology SH2 and SH3 domains (Schlessinger, 1994;
 Pawson, 1994), pleckstrin-homology (PH) domains (Lemmon et al., 1996;
 Shaw, 1996), phosphotyrosine-binding (PTB) domains (Harrison, 1996; van
 der Greer & Pawson, 1995; Kavanaugh et al., 1995) and postsynaptic density
 protein, disc-large, zo-1 (PDZ) domains (Woods & Bryant, 1991; Dho et al.,
 1992; Woods & Bryant, 1993; Keimedy, 1995; Kornau et al., 1995). So far,
 PDZ domains have been found in more than 50 proteins (Tsunoda et al.,
 1997), and many proteins have multiple PDZ domains (Pawson & Scott, 1997).
 For a review of PDZ domains, as well as the other protein-protein
 interactive domains, see Pawson & Scott (1997).
 A distinguishing feature of PDZ domains is their recognition of short
 peptides at the carboxyl terminal end of proteins. For example, one family
 of PDZ domains selected peptides with the consensus motif
 Glu-(Ser/Thr)-Xaa-(Val/Ile) (SEQ ID NO: 1) at the carboxy terminus whereas
 a second family of PDZ domains selected peptides with hydrophobic or
 aromatic side chains at the carboxy terminal three residues (Songyang et
 al., 1997). The presence of multiple PDZ domains in proteins may have at
 least two important consequences. An individual PDZ-containing protein
 could bind several subunits of a particular channel thereby inducing
 channel aggregations. Furthermore, the individual domains of a protein can
 have distinct binding specificities thereby inducing the formation of
 clusters that contain heterogeneous groups of proteins.
 One example of this latter consequence of multiple PDZ domains is the InaD
 protein which contains five PDZ domains and acts as a scaffolding protein
 to organize the light-activated signaling events in Drosophila (Shieh &
 Zhu, 1996; Tsunoda et al., 1997). InaD associates through distinct PDZ
 domains with a calcium channel(TRP), phospholipase C-.beta. (the target of
 rhodopsin-activated heterotrimeric guanine nucleotide-binding protein
 (Gq.alpha.) and protein kinase C.
 Two further properties of PDZ domains or proteins which contain them may
 expand their potential activities. First, some PDZ domains may bind
 internal peptide sequences and, indeed, have a propensity to undergo
 homotypic or heterotypic interactions with other PDZ domains (Brenman et
 al., 1996). Second, proteins with PDZ domains frequently contain other
 interaction modules, including SH3 and LIM domains, and catalytic elements
 such a tyrosine phosphatase or nitric oxide synthase domains. PDZ domains
 may therefore both coordinate the localization and clustering of receptors
 and channels, and provide a bridge to the cytoskeleton or intracellular
 signaling pathways.
 It is desired to determine the mechanisms of tumor suppression for MMAC1
 and to identify proteins which interact with the MMAC1 protein. Such
 proteins can be used to assay for mutated MMAC1 proteins and/or screen
 potential drugs for suppressing tumor growth and/or identify additional
 proteins which interact with MMAC 1.
 SUMMARY OF THE INVENTION
 The present invention is directed to the MMSC2 gene, its protein product
 and the use of the protein to (i) detect mutant MMAC1 proteins, (ii)
 screen for drugs which can be used for suppressing tumor growth and (iii)
 identify proteins which interact with the MMAC1 gene or are involved in
 the tumor suppression pathway of the MAMC1 gene.
 Using yeast two-hybrid screening, it has been found MMAC1 binds to a
 protein herein named MMSC2. The nucleotide sequence is set forth as SEQ ID
 NO:2, and the amino acid sequence is set forth as SEQ ID NO:3. It has been
 found that MMSC2 has 11 PDZ domains and that one or more of these domains
 interacts specifically with the three carboxyl terminal amino acids of
 MMAC1. Specifically, it has been found that PDZ domain numbers 7,10 and 13
 interact with MMAC1, with 7 appearing stronger. Since MMSC2 contains 11
 PDZ domains and interacts with MMAC1, a known tumor suppressor having a
 region of homology with protein tyrosine phosphatases, MMSC2 acts as a
 scaffolding protein in a common biochemical pathway with MMAC 1. These
 characteristics indicate that the interaction between MMAC 1 and MMSC2 is
 required for the tumor suppressor activity of MMAC1.

SUMMARY OF SEQUENCE LISTING
 SEQ ID NO:1 is a consensus motif to which one family of PDZ domains
 interacts. SEQ ID NO:2 is the nucleotide sequence for the MMSC2 gene. SEQ
 ID NO:3 is the amino acid sequence for the MMSC2 protein. SEQ ID NO:4 is
 the 15 C-terminal amino acids of MMAC1. SEQ ID NO:5 is primer 9BP-1 F1.
 SEQ ID NO:6 is primer 9BP-1 R4. SEQ ID NO:7 is primer 9BP-1 #1. SEQ ID
 NO:8 is primer 9BP-1 #2. SEQ ID NO:9 is primer 9BP-1 #NO:10 is primer
 9BP-1 #7. SEQ ID NO:11 is the SH3 binding peptide. SEQ ID NO:12 is the
 MMAC1 binding peptide. SEQ ID NOs:13-72 are primers for PCR amplification
 of the MMSC2 gene.
 DETAILED DESCRIPTION OF THE INVENTION
 The present invention is directed to the MMSC2 gene, its protein product
 and the use of the protein to (i) detect mutant MMAC1 proteins, (ii)
 screen for drugs which can be used for suppressing tumor growth and (iii)
 identify proteins which interact with the MMAC1 gene or are involved in
 the tumor suppression pathway of the MMAC1 gene.
 Using yeast two-hybrid screening, it has been found MMAC1 binds to a
 protein herein named MMSC2. The nucleotide sequence is set forth as SEQ ID
 NO:2, and the amino acid sequence is set forth as SEQ ID NO:3. It has been
 found MMSC2 has 11 PDZ domains and that one or more of these domains
 interacts specifically with the three carboxyl terminal amino acids of
 MMAC1. Specifically, it has been found that PDZ domain numbers 7, 10 and
 13 interact with MMAC 1 with 7 appearing stronger. Since MMSC2 contains 11
 PDZ domains and interacts with MMAC1, a known tumor suppressor having a
 region of homology with protein tyrosine phosphatases, MMSC2 acts as a
 scaffolding protein in a common biochemical pathway with MMAC1. These
 characteristics indicate that the interaction between MMAC1 and MMSC2 is
 required for the tumor suppressor activity of MMAC1.
 The evidence presented herein shows that the function of MMSC2 is to make a
 scaffold that binds to MMAC1, the phosphatase substrate(s), and the
 (probably oncogene) tyrosine kinase(s). Thus, a valuable drug will be one
 that can prevent binding of either the substrate(s) or the tyrosine
 kinases(s) to MMSC2.
 The yeast two-hybrid screening assay described herein identified five
 clones encoding bonafide MMAC1-interacting proteins. These clones were
 named PDZBN2B, PDZBN3A, PDZBN5B, PDZBN18D, and pdzk4. Comparison of the
 sequences of these clones suggested that they were all partial cDNAs
 derived from the same novel gene. A search of GenBank with these sequences
 revealed strong sequence similarity with a partial mouse cDNA sequence
 called 9ORF binding protein 1 (9BP-1)(GenBank Accession #AF000168).
 Several rounds of cDNA library screening were required to identify cDNA
 clones that could be assembled into the full length MMAC2 sequence. In the
 first round, a 509 base pair(bp) probe was developed from the 5' end of
 clone PDZBN2B using the primers 9BP-1 F1 and 9BP-1 R4. This probe was used
 to screen a human placental cDNA library and a human prostate cDNA
 library. Two of the informative clones obtained were p118a(placental) and
 pr63(prostate). A search of GenBank with this additional sequence yielded
 an additional human EST (GenBank Accesion #C75629). For the second round
 of cDNA library screening, a 202 bp probe was developed from the 5' end of
 this EST using primers 9BP-1 #1 and 9BP-1 #2. This probe was used to
 screen a human prostate cDNA library; two of the informative clones
 obtained were clone #10 and clone #3. For the third round of cDNA library
 screening, a 172 bp probe was developed from the 5' end of clone #3 using
 primers 9BP-1 #5 and 9BP-1 #7 and used to screen a human prostate cDNA
 library. One of the resulting clones, clone #6, yielded the start codon
 and part of the 5' UTR, including in-frame upstream stop codons. The
 nucleotide sequence for MMSC2 is set forth in SEQ ID NO:2 with the amino
 acid sequence of the encoded protein set forth in SEQ ID NO:3.
 As previously noted, SEQ ID NO:2 sets forth the nucleotide sequence for
 MMSC2. However, it has been found that the mRNA for MMSC2 is subject to
 alternate splicing On the basis of the sequence for MMSC2, genomic clones
 have been isolated and are being analyzed to determine splice junctions
 and alternate splicing for the mRNA. In addition, the PDZ domains of MMSC2
 are analyzed in the yeast two-hybrid assay to identify other proteins
 which interact with MMSC2 and consequently are involved in the MMAC1 tumor
 suppressor pathway.
 Since MMSC2 is an MMAC1 interacting protein that is involved in tumor
 suppression activity in the MMAC1 pathway, mutations in the MMSC2 gene
 which affect the interaction of MMSC2 with MMAC 1 or affect the
 interaction of other proteins with MMAC1 as a result of the scaffolding
 effect of MMSC2 will interfere with the MMAC1 tumor suppressor pathway and
 lead to tumorigenesis. Thus, an additional aspect of the present invention
 is the screening of MMSC2 for such mutations using conventional
 techniques. Such methods may further comprise the step of amplifying a
 portion of the MMAC2 gene, and may further include a step of providing a
 set of polynucleotides which are primers for amplification of said portion
 of the MMSC2 gene. The method is useful for identifying mutations for use
 in either diagnosis of cancer or prognosis of cancer. Since such variants
 can now be detected earlier, i.e., before symptoms appear, and more
 definitively, better treatment options will be available in those
 individuals identified as having harmful mutations in MMSC2.
 The present invention is directed to the determination that the MMSC2 binds
 to the C-terminal region of MMAC1 and is involved in a common pathway with
 MMAC1 which is a known tumor suppressor. Since many of the mutations in
 MMAC1 are frameshift or nonsense mutations which consequently alter the
 C-terminus of MMAC1, MMSC2 can be used to assay for normal or mutated MMAC
 1 proteins using conventional techniques.
 Finally, the present invention is directed to a method for screening drug
 candidates to identify drugs useful for treating or preventing cancer.
 Drug screening is performed by expressing mutant MMSC2 and assaying the
 effect of a drug candidate on the binding of MMSC2 with MMAC 1. Similarly,
 one can test the effect of a drug candidate on the binding of wild-type
 MMSC2 with a mutant MMAC1. Such assays can be performed in vitro or in
 vivo, such as in oocytes, mammalian cells or transgenic animals. Other
 assays may test the ability of a drug, wherein the drug may be, e.g., a
 peptide, to replace the activity of MMSC2 such that the drug plus MMAC1
 will work in concert similar to the normal wild-type interactions of MMSC2
 and MMAC1. Again, similar assays may be performed to screen for drugs
 which replace a mutant MMAC1 and will bind to wild-type MMSC2 to replace
 the MMAC1 function which is lacking as a result of a mutated MMAC1.
 According to the diagnostic and prognostic method of the present invention,
 alteration of the wild-type MMSC2 gene is detected. In addition, the
 method can be performed by detecting the wild- type MMSC2 gene and
 confirming the lack of a cause of cancer as a result of this locus.
 "Alteration of a wild-type gene" encompasses all forms of mutations
 including deletions, insertions and point mutations in the coding and
 noncoding regions. Deletions may be of the entire gene or of only a
 portion of the gene. Point mutations may result in stop codons, frameshift
 mutations or amino acid substitutions. Somatic mutations are those which
 occur only in certain tissues and are not inherited in the germline.
 Germline mutations can be found in any of a body's tissues and are
 inherited. Point mutational events may occur in regulatory regions, such
 as in the promoter of the gene, leading to loss or diminution of
 expression of the mRNA. Point mutations may also abolish proper RNA
 processing, leading to loss of expression of the MMSC2 gene product, or to
 a decrease in mRNA stability or translation efficiency.
 Useful diagnostic techniques include, but are not limited to fluorescent in
 situ hybridization (FISH), direct DNA sequencing, PFGE analysis, Southern
 blot analysis, single stranded conformation analysis (SSCA), RNase
 protection assay, allele-specific oligonucleotide (ASO), dot blot
 analysis, hybridization using nucleic acid modified with gold
 nanoparticles and PCR-SSCP, as discussed in detail further below. Also
 useful is the recently developed technique of DNA microchip technology.
 The presence of cancer due to a germline mutation at this locus may be
 ascertained by testing any tissue of a human for mutations of the MMSC2
 gene. For example, a person who has inherited a germline MMSC2 mutation,
 especially one which alters the interaction of MMSC2 with MMAC1, would be
 prone to develop cancer. This can be determined by testing DNA from any
 tissue of the person's body. Most simply, blood can be drawn and DNA
 extracted from the cells of the blood. In addition, prenatal diagnosis can
 be accomplished by testing fetal cells, placental cells or amniotic cells
 for mutations of the MMAC2 gene. Alteration of a wild-type MMSC2 allele,
 whether, for example, by point mutation or deletion, can be detected by
 any of the means discussed herein.
 There are several methods that can be used to detect DNA sequence
 variation. Direct DNA sequencing, either manual sequencing or automated
 fluorescent sequencing can detect sequence variation. Another approach is
 the single-stranded conformation polymorphism assay (SSCP) (Orita et al.,
 1989). This method does not detect all sequence changes, especially if the
 DNA fragment size is greater than 200 bp, but can be optimized to detect
 most DNA sequence variation. The reduced detection sensitivity is a
 disadvantage, but the increased throughput possible with SSCP makes it an
 attractive, viable alternative to direct sequencing for mutation detection
 on a research basis. The fragments which have shifted mobility on SSCP
 gels are then sequenced to determine the exact nature of the DNA sequence
 variation. Other approaches based on the detection of mismatches between
 the two complementary DNA strands include clamped denaturing gel
 electrophoresis (CDGE) (Sheffield et al., 1991), heteroduplex analysis
 (HA) (White et al., 1992) and chemical mismatch cleavage (CMC) (Grompe et
 al., 1989). None of the methods described above will detect large
 deletions, duplications or insertions, nor will they detect a regulatory
 mutation which affects transcription or translation of the protein. Other
 methods which might detect these classes of mutations such as a protein
 truncation assay or the asymmetric assay, detect only specific types of
 mutations and would not detect missense mutations. A review of currently
 available methods of detecting DNA sequence variation can be found in a
 recent review by Grompe (1993). Once a mutation is known, an allele
 specific detection approach such as allele specific oligonucleotide (ASO)
 hybridization can be utilized to rapidly screen large numbers of other
 samples for that same mutation. Such a technique can utilize probes which
 are labeled with gold nanoparticles to yield a visual color result
 (Elghanian et al., 1997).
 A rapid preliminary analysis to detect polymorphisms in DNA sequences can
 be performed by looking at a series of Southern blots of DNA cut with one
 or more restriction enzymes, preferably with a large number of restriction
 enzymes. Each blot contains a series of normal individuals and a series of
 cancer cases. Southern blots displaying hybridizing fragments differing in
 length from control DNA when probed with sequences near or including the
 MMSC2 locus indicate a possible mutation. If restriction enzymes which
 produce very large restriction fragments are used, then pulsed field gel
 electrophoresis (PFGE) is employed.
 Detection of point mutations may be accomplished amplification, e.g., PCR,
 from genomic or cDNA and sequencing the amplified nucleic acid or by
 molecular cloning of the MMSC2 allele and sequencing the allele using
 techniques well known in the art.
 There are six well known methods for a more complete, yet still indirect,
 test for confirming the presence of a susceptibility allele: 1) single
 stranded conformation analysis (SSCP) (Orita et al., 1989); 2) denaturing
 gradient gel electrophoresis (DGGE) (Wartell et al., 1990; Sheffield et
 al., 1989); 3) RNase protection assays (Finkelstein et al., 1990; Kinszler
 et al., 1991); 4) allele-specific oligonucleotides (ASOs) (Conner et al.,
 1983); 5) the use of proteins which recognize nucleotide mismatches, such
 as the E. coli mutS protein (Modrich, 1991); and 6) allele-specific PCR
 (Rano and Kidd, 1989). For allele-specific PCR, primers are used which
 hybridize at their 3' ends to a particular MMSC2 mutation. If the
 particular mutation is not present, an amplification product is not
 observed. Amplification Refractory Mutation System (ARMS) can also be
 used, as disclosed in European Patent Application Publication No. 0332435
 and in Newton et al., 1989. Insertions and deletions of genes can also be
 detected by cloning, sequencing and amplification. In addition,
 restriction fragment length polymorphism (RFLP) probes for the gene or
 surrounding marker genes can be used to score alteration of an allele or
 an insertion in a polymorphic fragment. Such a method is particularly
 useful for screening relatives of an affected individual for the presence
 of the mutation found in that individual. Other techniques for detecting
 insertions and deletions as known in the art can be used.
 In the first three methods (SSCP, DGGE and RNase protection assay), a new
 electrophoretic band appears, SSCP detects a band which migrates
 differentially because the sequence change causes a difference in
 single-strand, intramolecular base pairing. RNase protection involves
 cleavage of the mutant polynucleotide into two or more smaller fragments.
 DGGE detects differences in migration rates of mutant sequences compared
 to wild-type sequences, using a denaturing gradient gel. In an
 allele-specific oligonucleotide assay, an oligonucleotide is designed
 which detects a specific sequence, and the assay is performed by detecting
 the presence or absence of a hybridization signal. In the mutS assay, the
 protein binds only to sequences that contain a nucleotide mismatch in a
 heteroduplex between mutant and wild-type sequences.
 Mismatches, according to the present invention, are hybridized nucleic acid
 duplexes in which the two strands are not 100% complementary. Lack of
 total homology may be due to deletions, insertions, inversions or
 substitutions. Mismatch detection can be used to detect point mutations in
 the gene or in its mRNA product. While these techniques are less sensitive
 than sequencing, they are simpler to perform on a large number of samples.
 An example of a mismatch cleavage technique is the RNase protection
 method. In the practice of the present invention, the method involves the
 use of a labeled riboprobe which is complementary to the human wild-type
 MMSC2 gene coding sequence. The riboprobe and either mRNA or DNA isolated
 from the person are annealed (hybridized) together and subsequently
 digested with the enzyme RNase A which is able to detect some mismatches
 in a duplex RNA structure. If a mismatch is detected by RNase A, it
 cleaves at the site of the mismatch. Thus, when the annealed RNA
 preparation is separated on an electrophoretic gel matrix, if a mismatch
 has been detected and cleaved by RNase A, an RNA product will be sceen
 which is smaller than the full length duplex RNA for the riboprobe and the
 mRNA or DNA. The riboprobe need not be the full length of the mRNA or gene
 but can be a segment of either. If the riboprobe comprises only a segment
 of the mRNA or gene, it will be desirable to use a number of these probes
 to screen the whole mRNA sequence for mismatches.
 In similar fashion, DNA probes can be used to detect mismatches, through
 enzymatic or chemical cleavage. See, e.g., Cotton et al., 1988; Shenk et
 al., 1975; Novack et al., 1986. Alternatively, mismatches can be detected
 by shifts in the electrophoretic mobility of mismatched duplexes relative
 to matched duplexes. See, e.g., Cariello, 1988. With either riboprobes or
 DNA probes, the cellular mRNA or DNA which might contain a mutation can be
 amplified using PCR (see below) before hybridization. Changes in DNA of
 the MMSC2 gene can also be detected using Southern hybridization,
 especially if the changes are gross rearrangements, such as deletions and
 insertions.
 DNA sequences of the MMSC2 gene which have been amplified by use of PCR may
 also be screened using allele-specific probes. These probes are nucleic
 acid oligomers, each of which contains a region of the gene sequence
 harboring a known mutation. For example, one oligomer may be about 30
 nucleotides in length, corresponding to a portion of the gene sequence. By
 use of a battery of such allele-specific probes, PCR amplification
 products can be screened to identify the presence of a previously
 identified mutation in the gene. Hybridization of allele-specific probes
 with amplified MMSC2 sequences can be performed, for example, on a nylon
 filter. Hybridization to a particular probe under high stringency
 hybridization conditions indicates the presence of the same mutation in
 the tissue as in the allele-specific probe.
 The newly developed technique of nucleic acid analysis via microchip
 technology is also applicable to the present invention. In this technique,
 literally thousands of distinct oligonucleotide probes are built up in an
 array on a silicon chip. Nucleic acid to be analyzed is fluorescently
 labeled and hybridized to the probes on the chip. It is also possible to
 study nucleic acid-protein interactions using these nucleic acid
 microchips. Using this technique one can determine the presence of
 mutations or even sequence the nucleic acid being analyzed or one can
 measure expression levels of a gene of interest. The method is one of
 parallel processing of many, even thousands, of probes at once and can
 tremendously increase the rate of analysis. Several papers have been
 published which use this technique. Some of these are Hacia et al., 1996;
 Shoemaker et al., 1996; Chee et al., 1996; Lockhart et al., 1996; DeRisi
 et al., 1996; Lipshutz et al., 1995. This method has already been used to
 screen people for mutations in the breast cancer gene BRCA1 (Hacia et al.,
 1996). This new technology has been reviewed in a news article in Chemical
 and Engineering News (Borman, 1996) and been the subject of an editorial
 (Nature Genetics, 1996). Also see Fodor (1997).
 The most definitive test for mutations in a candidate locus is to directly
 compare genomic MMSC2 sequences from patients with those from a control
 population. Alternatively, one could sequence messenger RNA after
 amplification, e.g., by PCR, thereby eliminating the necessity of
 determining the exon structure of the candidate gene.
 Mutations from patients falling outside the coding region of MMSC2 can be
 detected by examining the non-coding regions, such as introns and
 regulatory sequences near or within the genes. An early indication that
 mutations in noncoding regions are important may come from Northern blot
 experiments that reveal messenger RNA molecules of abnormal size or
 abundance in patients as compared to control individuals.
 Alteration of MMSC2 mRNA expression can be detected by any techniques known
 in the art. These include Northern blot analysis, PCR amplification and
 RNase protection. Diminished mRNA expression indicates an alteration of
 the wild-type gene. Alteration of wild-type genes can also be detected by
 screening for alteration of wild-type MMSC2 protein. For example,
 monoclonal antibodies immunoreactive with MMSC2 can be used to screen a
 tissue. Lack of cognate antigen would indicate a mutation. Antibodies
 specific for products of mutant alleles could also be used to detect
 mutant gene product. Such immunological assays can be done in any
 convenient formats known in the art. These include Western blots,
 immunohistochemical assays and ELISA assays. Any means for detecting an
 altered MMSC2 protein can be used to detect alteration of the wild-type
 MMSC2 gene. Functional assays, such as protein binding determinations, can
 be used. In addition, assays can be used which detect MMSC2 biochemical
 function. Finding a mutant MMSC2 gene product indicates alteration of a
 wild-type MMSC2 gene. One such binding assay is the binding of MMSC2 with
 wild-type MMAC1. Conversely, wild-type MMSC2 or the PDZ domain interacting
 with MMAC1 can be used in a protein binding assay or biochemical function
 assay to detect normal or mutant MMAC1 proteins, where the mutant proteins
 are proteins lacking a wild-type C-terminus.
 A mutant MMSC2 gene or gene product or a mutant MMAC1 can also be detected
 in other human body samples, such as serum, stool, urine and sputum. The
 same techniques discussed above for detection of mutant genes or gene
 products in tissues can be applied to other body samples. By screening
 such body samples, a simple early diagnosis can be achieved for cancer
 resulting from a mutation in the MMSC2 gene.
 The primer pairs of the present invention are useful for determination of
 the nucleotide sequence of a particular MMSC2 allele using PCR. The pairs
 of single-stranded DNA primers for MMSC2 can be annealed to sequences
 within or surrounding the MMSC2 gene in order to prime amplifying DNA
 synthesis of the gene itself. A complete set of these primers allows
 synthesis of all of the nucleotides of the gene coding sequences, i.e.,
 the exons. The set of primers preferably allows synthesis of both intron
 and exon sequences. Allele-specific primers can also be used. Such primers
 anneal only to particular MMSC2 mutant alleles, and thus will only amplify
 a product in the presence of the mutant allele as a template.
 In order to facilitate subsequent cloning of amplified sequences, primers
 may have restriction enzyme site sequences appended to their 5' ends.
 Alternatively, primers can also be prepared with 5' phosphoryl groups
 which will allow for blunt end coloning of amplied sequences. Thus, all
 nucleotides of the primers are derived from MMSC2 sequence or sequences
 adjacent to MMSC2, except for the few nucleotides necessary to form a
 restriction enzyme site. Such enzymes and sites are well known in the art.
 The primers themselves can be synthesized using techniques which are well
 known in the art. Generally, the primers can be made using oligonucleotide
 synthesizing machines which are commercially available. Given the sequence
 of MMSC2, design of particular primers is well within the skill of the
 art.
 The nucleic acid probes provided by the present invention are useful for a
 number of purposes. They can be used in Southern hybridization to genomic
 DNA and in the RNase protection method for detecting point mutations
 already discussed above. The probes can be used to detect PCR
 amplification products. They may also be used to detect mismatches with
 the MMSC2 gene or mRNA using other techniques Mutations which interfere
 with the function of the MMSC2 gene product are involved in the
 pathogenesis of cancer. Thus, the presence of an altered (or a mutant)
 MMSC2 gene which produces a protein having a loss of function, or altered
 function, directly increases the risk of cancer. In order to detect a
 MMSC2 gene mutation, a biological sample is prepared and analyzed for a
 difference between the sequence of the allele being analyzed and the
 sequence of the wild-type allele. Mutant MMSC2 alleles can be initially
 identified by any of the techniques described above. The mutant alleles
 are then sequenced to identify the specific mutation of the particular
 mutant allele. Alternatively, mutant alleles can be initially identified
 by identifying mutant (altered) proteins, using conventional techniques.
 The mutant alleles are then sequenced to identify the specific mutation
 for each allele. The mutations, especially those which lead to an altered
 function of the protein, are then used for the diagnostic and prognostic
 methods of the present invention.
 Definitions
 The present invention employs the following definitions.
 "Amplification of Polynucleotides" utilizes methods such as the polymerase
 chain reaction (PCR), ligation amplification (or ligase chain reaction,
 LCR) and amplification methods based on the use of Q-beta replicase. Also
 useful are strand displacement amplification (SDA), thermophilic SDA,
 nucleic acid sequence based amplification (3SR or NASBA) and repair chain
 reaction (RCR). These methods are well known and widely practiced in the
 art. See, e.g., U.S. Pat. Nos. 4,683,195 and 4,683,202 and Innis et al.,
 1990 (for PCR); Wu et al., 1989a and EP 320,308A (for LCR); U.S. Pat. Nos.
 5,270,184 and 5,455,166 and Walker et al., 1992 (for SDA); Spargo et al.,
 1996 (for thermophilic SDA) and U.S. Pat. No. 5,409,818, Fahy et al., 1991
 and Compton, 1991 for 3SR and NASBA. Reagents and hardware for conducting
 PCR are commercially available. Primers useful to amplify sequences from
 the MMSC2 region are preferably complementary to, and hybridize
 specifically to sequences in the MMSC2 region or in regions that flank a
 target region therein. MMSC2 sequences generated by amplification may be
 sequenced directly. Alternatively, but less desirably, the amplified
 sequence(s) may be cloned prior to sequence analysis. A method for the
 direct cloning and sequence analysis of enzymatically amplified genomic
 segments has been described by Scharf, 1986.
 "Analyte polynucleotide" and "analyte strand" refer to a single- or
 double-stranded polynucleotide which is suspected of containing a target
 sequence, and which may be present in a variety of types of samples,
 including biological samples.
 "Antibodies." The present invention also provides polyclonal and/or
 monoclonal antibodies and fragments thereof, and immunologic binding
 equivalents thereof, which are capable of specifically binding to the
 MMSC2 polypeptide and fragments thereof or to polynucleotide sequences
 from the MMSC2 region. The term "antibody" is used both to refer to a
 homogeneous molecular entity, or a mixture such as a serum product made up
 of a plurality of different molecular entities. Polypeptides may be
 prepared synthetically in a peptide synthesizer and coupled to a carrier
 molecule (e.g., keyhole limpet hemocyanin) and injected over several
 months into rabbits. Rabbit sera is tested for immunoreactivity to the
 MMSC2 polypeptide or fragment. Monoclonal antibodies may be made by
 injecting mice with the protein polypeptides, fusion proteins or fragments
 thereof. Monoclonal antibodies will be screened by ELISA and tested for
 specific immunoreactivity with MMSC2 polypeptide or fragments thereof.
 See, Harlow and Lane, 1988. These antibodies will be useful in assays as
 well as pharmaceuticals.
 Once a sufficient quantity of desired polypeptide has been obtained, it may
 be used for various purposes. A typical use is the production of
 antibodies specific for binding. These antibodies may be either polyclonal
 or monoclonal, and may be produced by in vitro or in vivo techniques well
 known in the art. For production of polyclonal antibodies, an appropriate
 target immune system, typically mouse or rabbit, is selected.
 Substantially purified antigen is presented to the immune system in a
 fashion determined by methods appropriate for the animal and by other
 parameters well known to immunologists. Typical sites for injection are in
 footpads, intramuscularly, intraperitoneally, or intradermally. Of course,
 other species may be substituted for mouse or rabbit. Polyclonal
 antibodies are then purified using techniques known in the art, adjusted
 for the desired specificity.
 An immunological response is usually assayed with an immunoassay. Normally,
 such immunoassays involve some purification of a source of antigen, for
 example, that produced by the same cells and in the same fashion as the
 antigen. A variety of immunoassay methods are well known in the art. See,
 e.g., Harlow and Lane, 1988, or Goding, 1986.
 Monoclonal antibodies with affinities of 10.sup.-8 M.sup.-1 or preferably
 10.sup.-9 to 10.sup.-10 M.sup.-1 or stronger will typically be made by
 standard procedures as described, e.g., in Harlow and Lane, 1988 or
 Goding, 1986. Briefly, appropriate animals will be selected and the
 desired immunization protocol followed. After the appropriate period of
 time, the spleens of such animals are excised and individual spleen cells
 fused, typically, to immortalized myeloma cells under appropriate
 selection conditions. Thereafter, the cells are clonally separated and the
 supernatants of each clone tested for their production of an appropriate
 antibody specific for the desired region of the antigen.
 Other suitable techniques involve in vitro exposure of lymphocytes to the
 antigenic polypeptidcs, or alternatively, to selection of libraries of
 antibodies in phage or similar vectors. See Huse et cal., 1989. The
 polypeptides and antibodies of the present invention may be used with or
 without modification. Frequently, polypeptides and antibodies will be
 labeled by joining, either covalently or non-covalently, a substance which
 provides for a detectable signal. A wide variety of labels and conjugation
 techniques are known and are reported extensively in both the scientific
 and patent literature. Suitable labels include radionuclides, enzymes,
 substrates, cofactors, inhibitors, fluorescent agents, chemiluminescent
 agents, magnetic particles and the like. Patents teaching the use of such
 labels include U.S. Pat. Nos. 3,817,837; 3,850,752; 3,939,350; 3,996,345;
 4,277,437; 4,275,149 and 4,366,241. Also, recombinant immunoglobulills may
 be produced (see U.S. Pat. No. 4,816,567).
 "Binding partner" refers to a molecule capable of binding a ligand molecule
 with high specificity, as for example, an antigen and an antigen-specific
 antibody or an enzyme and its inhibitor. In general, the specific binding
 partners must bind with sufficient affinity to immobilize the analyte
 copy/complementary strand duplex (in the case of polynucleotide
 hybridization) under the isolation conditions. Specific binding partners
 are known in the art and include, for example, biotin and avidin or
 streptavidin, IgG and protein A, the numerous, known receptor-ligand
 couples, and complementary polynucleotide strands. In the case of
 complementary polynucleotide binding partners, the partners are normally
 at least about 15 bases in length, and may be at least 40 bases in length.
 It is well recognized by those of skill in the art that lengths shorter
 than 15 (e.g., 8 bases), between 15 and 40, and greater than 40 bases may
 also be used. The polynucleotides may be composed of DNA, RNA, or
 synthetic nucleotide analogs. In addition, as disclosed herein, MMAC1 and
 PDZ binding peptides, as well as several other proteins, bind to or
 interact with MMSC2. Each of these proteins are also considered binding
 partners herein. Further binding partners can be identifed using, e.g.,
 the two-hybrid yeast screening assay as described herein.
 A "biological sample" refers to a sample of tissue or fluid suspected of
 containing an analyte polynucleotide or polypeptide from an individual
 including, but not limited to, e.g., plasma, serum, spinal fluid, lymph
 fluid, the external sections of the skin, respiratory, intestinal, and
 genitourinary tracts, tears, saliva, blood cells, tumors, organs, tissue
 and samples of in vitro cell culture constituents.
 "Encode". A polynucleotide is said to "encode" a polypeptide if, in its
 native state or when manipulated by methods well known to those skilled in
 the art, it can be transcribed and/or translated to produce the mRNA for
 and/or the polypeptide or a fragment thereof. The anti-sense strand is the
 complement of such a nucleic acid, and the encoding sequence can be
 deduced therefrom.
 "Isolated" or "substantially pure". An "isolated" or "substantially pure"
 nucleic acid (e.g., an RNA, DNA or a mixed polymer) is one which is
 substantially separated from other cellular components which naturally
 accompany a native human sequence or protein, e.g., ribosomes,
 polymerases, many other human genome sequences and proteins. The term
 embraces a nucleic acid sequence or protein which has been removed from
 its naturally occurring environment, and includes recombinant or cloned
 DNA isolates and chemically synthesized analogs or analogs biologically
 synthesized by heterologous systems.
 "MMSC2 Allele" refers to normal alleles of the MMSC2 locus that interact
 with MMAC1 as well as alleles of MMSC2 carrying variations that affect the
 interaction with MMAC1 and that cause cancer.
 "MMSC2 Locus", "MMSC2 Gene", "MMSC2 Nucleic Acids" or "MMSC2
 Polynucleotide" each refer to polynucleotides, all of which are in the
 MMSC2 region, that are likely to be expressed in normal tissue, certain
 alleles of which adversely affect the interaction with MMAC1 and result in
 cancer. The MMSC2 locus is intended to include coding sequences,
 intervening sequences and regulatory elements controlling transcription
 and/or translation. The MMSC2 locus is intended to include all allelic
 variations of the DNA sequence.
 These terms, when applied to a nucleic acid, refer to a nucleic acid which
 encodes a human MMSC2 polypeptide, fragment, homolog or variant,
 including, e.g., protein fusions or deletions. The nucleic acids of the
 present invention will possess a sequence which is either derived from, or
 substantially similar to a natural MMSC2-encoding gene or one having
 substantial homology with a natural MMSC2-encoding gene or a portion
 thereof.
 The MMSC2 gene or nucleic acid includes normal alleles of the MMSC2 gene,
 both silent alleles having no effect on the amino acid sequence of the
 MMSC2 polypeptide and alleles leading to amino acid sequence variants of
 the MMSC2 polypeptide that do not substantially affect its function. These
 terms also include alleles having one or more mutations which adversely
 affect the function of the MMSC2 polypeptide. A mutation may be a change
 in the MMSC2 nucleic acid sequence which produces a deleterious change in
 the amino acid sequence of the MMSC2 polypeptide, resulting in partial or
 complete loss of MMSC2 function, or may be a change in the nucleic acid
 sequence which results in the loss of effective MMSC2 expression or the
 production of aberrant forms of the MMSC2 polypeptide.
 The MMSC2 nucleic acid may be that shown in SEQ ID NO:2, or it may be an
 allele as described above, or a variant or derivative differing from that
 shown by a change which is one or more of addition, insertion, deletion
 and substitution of one or more nucleotides of the sequence shown. Changes
 to the nucleotide sequence may result in an amino acid change at the
 protein level, or not, as determined by the genetic code.
 Thus, nucleic acid according to the present invention may include a
 sequence different from the sequence shown in SEQ ID NO:2 yet encode a
 polypeptide with the same amino acid sequence as shown in SEQ ID NO:3.
 That is, nucleic acids of the present invention include sequences which
 are degenerate as a result of the genetic code. On the other hand, the
 encoded polypeptide may comprise an amino acid sequence which differs by
 one or more amino acid residues from the amino acid sequence shown in SEQ
 ID NO:3. Nucleic acid encoding a polypeptide which is an amino acid
 sequence variant, derivative or allele of the amino acid sequence shown in
 SEQ ID NO:3 is also provided by the present invention.
 The MMSC2 gene also refers to (a) any DNA sequence that (i) hybridizes to
 the complement of the DNA sequences that encode the amino acid sequence
 set forth in SEQ ID NO:3 under highly stringent conditions (Ausubel et
 al.) (ii) and encodes a gene product functionally equivalent to MMSC2, or
 (b) any DNA sequence that (i) hybridizes to the complement of the DNA
 sequences that encode the amino acid sequence set forth in SEQ ID NO:3
 under less stringent conditions, such as moderately stringent conditions
 (Ausubel et al.), and (ii) encodes a gene product functionally equivalent
 to MMSC2. The invention also includes nucleic acid molecules that are the
 complements of the sequences described herein.
 The polynuclcotide compositions of this invention include RNA, cDNA,
 genomic DNA, synthetic forms, and mixed polymers, both sense and antisense
 strands, and may be chemically or biochemically modified or may contain
 non-natural or derivatized nucleotide bases, as will be readily
 appreciated by those skilled in the art. Such modifications include, for
 example, labels, methylation, substitution of one or more of the naturally
 occurring nucleotides with an analog, internucleotide modifications such
 as uncharged linkages (e.g., methyl phosphonates, phosphotriesters,
 phosphoramidates, carbamates, etc.), charged linkages (e.g.,
 phosphorothioates, phosphorodithioates, etc.), pendent moieties (e.g.,
 polypeptides), intercalators (e.g., acridine, psoralen, etc.), chelators,
 alkylators, and modified linkages (e.g., alpha anomeric nucleic acids,
 etc.). Also included are synthetic molecules that mimic polynucleotides in
 their ability to bind to a designated sequence via hydrogen bonding and
 other chemical interactions. Such molecules are known in the art and
 include, for example, those in which peptide linkages substitute for
 phosphate linkages in the backbone of the molecule.
 The present invention provides recombinant nucleic acids comprising all or
 part of the MMSC2 region. The recombinant construct may be capable of
 replicating autonomously in a host cell. Alternatively, the recombinant
 construct may become integrated into the chromosomal DNA of the host cell.
 Such a recombinant polynucleotide comprises a polynucleotide of genomic,
 cDNA, RNA, semi-synthetic, or synthetic origin which, by virtue of its
 origin or manipulation, 1) is not associated with all or a portion of a
 polynucleotide with which it is associated in nature; 2) is linked to a
 polynucleotide other than that to which it is linked in nature; or 3) does
 not occur in nature. Where nucleic acid according to the invention
 includes RNA, reference to the sequence shown should be construed as
 reference to the RNA equivalent, with U substituted for T.
 Therefore, recombinant nucleic acids comprising sequences otherwise not
 naturally occurring are provided by this invention. Although the wild-type
 sequence may be employed, it will often be altered, e.g., by deletion,
 substitution or insertion. cDNA or genomic libraries of various types may
 be screened as natural sources of the nucleic acids of the present
 invention, or such nucleic acids may be provided by amplification of
 sequences resident in genomic DNA or other natural sources, e.g., by PCR.
 The choice of cDNA libraries normally corresponds to a tissue source which
 is abundant in mRNA for the desired proteins. Phage libraries are normally
 preferred, but other types of libraries may be used. Clones of a library
 are spread onto plates, transferred to a substrate for screening,
 denatured and probed for the presence of desired sequences.
 The DNA sequences used in this invention will usually comprise at least
 about five codons (15 nucleotides), more usually at least about 7-15
 codons, and most preferably, at least about 35 codons. One or more introns
 may also be present. This number of nucleotides is usually about the
 minimal length required for a successful probe that would hybridize
 specifically with a MMSC2-encoding sequence. In this context, oligomers of
 as low as 8 nucleotides, more generally 8-17 nucleotides, can be used for
 probes, especially in connection with chip technology.
 Techniques for nucleic acid manipulation are described generally, for
 example, in Sambrook et al., 1989 or Ausubel et al., 1992. Reagents useful
 in applying such techniques, such as restriction enzymes and the like, are
 widely known in the art and commercially available from such vendors as
 New England BioLabs, Boehringer Mannheim, Amersham, Promega, U. S.
 Biochemicals, New England Nuclear, and a number of other sources. The
 recombinant nucleic acid sequences used to produce fusion proteins of the
 present invention may be derived from natural or synthetic sequences. Many
 natural gene sequences are obtainable from various cDNA or from genomic
 libraries using appropriate probes. See, GenBank, National Institutes of
 Health.
 As used herein, a "portion" of the MMSC2 locus or region or allele is
 defined as having a minimal size of at least about eight nucleotides, or
 preferably about 15 nucleotides, or more preferably at least about 25
 nucleotides, and may have a minimal size of at least about 40 nucleotides.
 This definition includes all sizes in the range of 8-40 nucleotides as
 well as greater than 40 nucleotides. Thus, this definition includes
 nucleic acids of 8, 12, 15, 20, 25, 40, 60, 80, 100, 200, 300, 400, 500
 nucleotides, or nucleic acids having any number of nucleotides within
 these values (e.g., 9, 10, 11, 16, 23, 30, 38, 50, 72, 121, etc,
 nucleotides), or nucleic acids having more than 500 nucleotides, or any
 number of nucleotides between 500 and the number shown in SEQ ID NO:2. The
 present invention includes all novel nucleic acids having at least 8
 nucleotides derived from SEQ ID NO:2, its complement or functionally
 equivalent nucleic acid sequences. The present invention does not include
 nucleic acids which exist in the prior art. That is, the present invention
 includes all nucleic acids having at least 8 nucleotides derived from SEQ
 ID NO:2 with the proviso that it does not include nucleic acids existing
 in the prior art.
 "MMSC2 protein" or "MMSC2 polypeptide" refers to a protein or polypeptide
 encoded by the MAIMSC2 locus, variants or fragments thereof. The term
 "polypeptide" refers to a polymer of amino acids and its equivalent and
 does not refer to a specific length of the product; thus, peptides,
 oligopeptides and proteins are included within the definition of a
 polypeptide. This term also does not refer to, or exclude modifications of
 the polypeptide, for example, glycosylations, acetylations,
 phosphorylations, and the like. Included within the definition are, for
 example, polypeptides containing one or more analogs of an amino acid
 (including, for example, unnatural amino acids, etc.), polypeptides with
 substituted linkages as well as other modifications known in the art, both
 naturally and non-naturally occurring. Ordinarily, such polypeptides will
 be at least about 50% homologous to the native MMSC2 sequence, preferably
 in excess of about 90%, and more preferably at least about 95% homologous.
 Also included are proteins encoded by DNA which hybridize under high or
 low stringency conditions, to MMSC2-encoding nucleic acids and closely
 related polypeptides or proteins retrieved by antisera to the MMSC2
 protein(s).
 The MMSC2 polypeptide may be that shown in SEQ ID NO:3 which may be in
 isolated and/or purified form, free or substantially free of material with
 which it is naturally associated. The polypeptide may, if produced by
 expression in a prokaryotic cell or produced synthetically, lack native
 post-translational processing, such as glycosylation. Alternatively, the
 present invention is also directed to polypeptides which are sequence
 variants, alleles or derivatives of the MMSC2 polypeptide. Such
 polypeptides may have an amino acid sequence which differs from that set
 forth in SEQ ID NO:3 by one or more of addition, substitution, deletion or
 insertion of one or more amino acids. Preferred such polypeptides have
 MMSC2 function.
 Substitutional variants typically contain the exchange of one amino acid
 for another at one or more sites within the protein, and may be designed
 to modulate one or more properties of the polypeptide, such as stability
 against proteolytic cleavage, without the loss of other functions or
 properties. 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 involved.
 Preferred substitutions are ones which are conservative, that is, one
 amino acid is replaced with one of similar shape and charge. Conservative
 substitutions are well known in the art and typically include
 substitutions within the following groups: glycine, alanine; valine,
 isoleucine, leucine; aspartic acid, glutamic acid; asparagine, glutamine;
 serine, threonine; lysine, arginine; and tyrosine, phenylalanine.
 Certain amino acids may be substituted for other amino acids in a protein
 structure without appreciable loss of interactive binding capacity with
 structures such as, for example, antigen-binding regions of antibodies or
 binding sites on substrate molecules or binding sites on proteins
 interacting with the MMSC2 polypeptide. Since it is the interactive
 capacity and nature of a protein which defines that protein's biological
 functional activity, certain amino acid substitutions can be made in a
 protein sequence, and its underlying DNA coding sequence, and nevertheless
 obtain a protein with like properties. In making such changes, the
 hydrophathic index of amino acids may be considered. The importance of the
 hydrophobic amino acid index in conferring interactive biological function
 on a protein is generally understood in the art (Kyte & Doolittle, 1982).
 Alternatively, the substitution of like amino acids can be made
 effectively on the basis of hydrophilicity. The importance of
 hydrophilicity in conferring interactive biological function of a protein
 is generally understood in the art (U.S. Pat. No. 4,554,101). The use of
 the hydrophobic index or hydrophilicity in designing polypeptides is
 further discussed in U.S. Pat. No. 5,691,198.
 The length of polypeptide sequences compared for homology will generally be
 at least about 16 amino acids, usually at least about 20 residues, more
 usually at least about 24 residues, typically at least about 28 residues,
 and preferably more than about 35 residues.
 "Operably linked" refers to a juxtaposition wherein the components so
 described are in a relationship permitting them to function in their
 intended mainer. For instance, a promoter is operably linked to a coding
 sequence if the promoter affects its transcription or expression.
 The term "peptide mimetic" or "mimetic" is intended to refer to a substance
 which has the essential biological activity of the MMSC2 polypeptide. A
 peptide mimetic may be a peptide-containing molecule that mimics elements
 of protein secondary structure (Johnson et al., 1993). The underlying
 rationale behind the use of peptide mimetics is that the peptide backbone
 of proteins exists chiefly to orient amino acid side chains in such a way
 as to facilitate molecular interactions, such as those of antibody and
 antigen, enzyme and substrate or scaffolding proteins. A peptide mimetic
 is designed to permit molecular interactions similar to the natural
 molecule. A mimetic may not be a peptide at all, but it will retain the
 essential biological activity of natural MMSC2 polypeptide.
 "Probes". Polynucleotide polymorphisms associated with MMSC2 alleles which
 predispose to cancer are detected by hybridization with a polynucleotide
 probe which forms a stable hybrid with that of the target sequence, under
 highly stringent to moderately stringent hybridization and wash
 conditions. If it is expected that the probes will be perfectly
 complementary to the target sequence, high stringency conditions will be
 used. Hybridization stringency may be lessened if some mismatching is
 expected, for example, if variants are expected with the result that the
 probe will not be completely complementary. Conditions are chosen which
 rule out nonspecific/adventitious bindings, that is, which minimize noise.
 (It should be noted that throughout this disclosure, if it is simply
 stated that "stringent" conditions are used that is meant to be read as
 "high stringency" conditions are used.) Since such indications identify
 neutral DNA polymorphisms as well as mutations, these indications need
 further analysis to demonstrate detection of a MMSC2 susceptibility
 allele.
 Probes for MMSC2 alleles may be derived from the sequences of the MMSC2
 region, its cDNA, functionally equivalent sequences, or the complements
 thereof. The probes may be of any suitable length, which span all or a
 portion of the MMSC2 region, and which allow specific hybridization to the
 region. If the target sequence contains a sequence identical to that of
 the probe, the probes may be short, e.g., in the range of about 8-30 base
 pairs, since the hybrid will be relatively stable under even highly
 stringent conditions. If some degree of mismatch is expected with the
 probe, i.e., if it is suspected that the probe will hybridize to a variant
 region, a longer probe may be employed which hybridizes to the target
 sequence with the requisite specificity.
 The probes will include an isolated polynucleotide attached to a label or
 reporter molecule and may be used to isolate other polynucleotide
 sequences, having sequence similarity by standard methods. For techniques
 for preparing and labeling probes see, e.g., Sambrook et al., 1989 or
 Ausubel et al., 1992. Other similar polynucleotides may be selected by
 using homologous polynucleotides. Alternatively, polynucleotides encoding
 these or similar polypeptides may be synthesized or selected by use of the
 redundancy in the genetic code. Various codon substitutions may be
 introduced, e.g., by silent changes (thereby producing various restriction
 sites) or to optimize expression for a particular system. Mutations may be
 introduced to modify the properties of the polypeptide, perhaps to change
 the polypeptide degradation or turnover rate.
 Probes comprising synthetic oligonucleotides or other polynucleotides of
 the present invention may be derived from naturally occurring or
 recombinant single- or double-stranded polynucleotides, or be chemically
 synthesized. Probes may also be labeled by nick translation, Klenow
 fill-in reaction, or other methods known in the art.
 Portions of the polynucleotide sequence having at least about eight
 nucleotides, usually at least about 15 nucleotides, and fewer than about 9
 kb, usually fewer than about 1.0 kb, from a polynucleotide sequence
 encoding MMSC2 are preferred as probes. This definition therefore includes
 probes of sizes 8 nucleotides through 9000 nucleotides. Thus, this
 definition includes probes of 8, 12, 15, 20, 25, 40, 60, 80, 100, 200,
 300, 400, 500 nucleotides, or probes having any number of nucleotides
 within these values (e.g., 9, 10, 11, 16, 23, 30, 38, 50, 72, 121, etc,
 nucleotides), or probes having more than 500 nucleotides, or any number of
 nucleotides between 500 and the number shown in SEQ ID NO:2. The probes
 may also be used to determine whether mRNA encoding MMSC2 is present in a
 cell or tissue. The present invention includes all novel probes having at
 least 8 nucleotides derived from SEQ ID NO:2, its complement or
 functionally equivalent nucleic acid sequences. The present invention does
 not include probes which exist in the prior art. That is, the present
 invention includes all probes having at least 8 nucleotides derived from
 SEQ ID NO:2 with the proviso that it does not include probes existing in
 the prior art.
 Similar considerations and nucleotide lengths are also applicable to
 primers which may be used for the amplification of all or part of the
 MMSC2 gene. Thus, a definition for primers includes primers of 8, 12, 15,
 20, 25, 40, 60, 80, 100, 200, 300, 400, 500 nucleotides, or primers having
 any number of nucleotides within these values (e.g., 9, 10, 11, 16, 23,
 30, 38, 50, 72, 121, etc, nucleotides), or primers having more than 500
 nucleotides, or any number of nucleotides between 500 and 9000. The
 primers may also be used to determine whether mRNA encoding MMSC2 is
 present in a cell or tissue. The present invention includes all novel
 primers having at least 8 nucleotides derived from the MMSC2 locus for
 amplifying the MMSC2 gene, its complement or functionally equivalent
 nucleic acid sequences. The present invention does not include primers
 which exist in the prior art. That is, the present invention includes all
 primers having at least 8 nucleotides with the proviso that it does not
 include primers existing in the prior art.
 "Protein modifications or fragments" are provided by the present invention
 for MMSC2 polypeptides or fragments thereof which are substantially
 homologous to primary structural sequence but which include, e.g., in vivo
 or in vitro chemical and biochemical modifications or which incorporate
 unusual amino acids. Such modifications include, for example, acetylation,
 carboxylation, phosphorylation, glycosylation, ubiquitination, labeling,
 e.g., with radionuclides, and various enzymatic modifications, as will be
 readily appreciated by those well skilled in the art. A variety of methods
 for labeling polypeptides and of substituents or labels useful for such
 purposes are well known in the art, and include radioactive isotopes such
 as .sup.32 P, ligands which bind to labeled antiligands (e.g.,
 antibodies), fluorophores, chemiluminescent agents, enzymes, and
 antiligands which can serve as specific binding pair members for a labeled
 ligand. The choice of label depends on the sensitivity required, ease of
 conjugation with the primer, stability requirements, and available
 instrumentation. Methods of labeling polypeptides are well known in the
 art. See Sambrook et al., 1989 or Ausubel et al., 1992.
 Besides substantially full-length polypeptides, the present invention
 provides for biologically active fragments of the polypeptides.
 Significant biological activities include ligand-binding, immunological
 activity and other biological activities characteristic of MMSC2
 polypeptides. Immunological activities include both immunogenic function
 in a target immune system, as well as sharing of immunological epitopes
 for binding, serving as either a competitor or substitute antigen for an
 epitope of the MMSC2 protein. As used herein, "epitope" refers to an
 antigenic determinant of a polypeptide. An epitope could comprise three
 amino acids in a spatial conformation which is unique to the epitope.
 Generally, an epitope consists of at least five such amino acids, and more
 usually consists of at least 8-10 such amino acids. Methods of determining
 the spatial conformation of such amino acids are known in the art.
 For immunological purposes, tandem-repeat polypeptide segments may be used
 as immunogens, thereby producing highly antigenic proteins. Alternatively,
 such polypeptides will serve as highly efficient competitors for specific
 binding. Production of antibodies specific for MMSC2 polypeptides or
 fragments thereof is described below.
 The present invention also provides for fusion polypeptides, comprising
 MMSC2 polypeptides and fragments. Homologous polypeptides may be fusions
 between two or more MMSC2 polypeptide sequences or between the sequences
 of MMSC2 and a related protein. Likewise, heterologous fusions may be
 constructed which would exhibit a combination of properties or activities
 of the derivative proteins. For example, ligand-binding or other domains
 may be "swapped" between different new fusion polypeptides or fragments.
 Such homologous or heterologous fusion polypeptides may display, for
 example, altered strength or specificity of binding. Fusion partners
 include immunoglobulins, bacterial .beta.-galactosidase, trpE, protein A,
 .beta.-lactamase, .alpha.-amylase, alcohol dehydrogenase and yeast alpha
 mating factor. See Godowski et al., 1988.
 Fusion proteins will typically be made by either recombinant nucleic acid
 methods, as described below, or may be chemically synthesized. Techniques
 for the synthesis of polypeptides arc described, for example, in
 Merrifield, 1963.
 "Protein purification" refers to various methods for the isolation of the
 MMSC2 polypeptides from other biological material, such as from cells
 transformed with recombinant nucleic acids encoding MMSC2, and are well
 known in the art. For example, such polypeptides may be purified by
 immunoaffinity chromatography employing, e.g., the antibodies provided by
 the present invention. Various methods of protein purification are well
 known in the art, and include those described in Deutscher, 1990 and
 Scopes, 1982.
 The terms "isolated", "substantially pure", and "substantially homogeneous"
 are used interchangeably to describe a protein or polypeptide which has
 been separated from components which accompany it in its natural state. A
 monomeric protein is substantially pure when at least about 60 to 75% of a
 sample exhibits a single polypeptide sequence. A substantially pure
 protein will typically comprise about 60 to 90% W/W of a protein sample,
 more usually about 95%, and preferably will be over about 99% pure.
 Protein purity or homogeneity may be indicated by a number of means well
 known in the art, such as polyacrylamide gel electrophoresis of a protein
 sample, followed by visualizing a single polypeptide band upon staining
 the gel. For certain purposes, higher resolution may be provided by using
 HPLC or other means well known in the art which are utilized for
 purification.
 A MMSC2 protein is substantially free of naturally associated components
 when it is separated from the native contaminants which accompany it in
 its natural state. Thus, a polypeptide which is chemically synthesized or
 synthesized in a cellular system different from the cell from which it
 naturally originates will be substantially free from its naturally
 associated components. A protein may also be rendered substantially free
 of naturally associated components by isolation, using protein
 purification techniques well known in the art.
 A polypeptide produced as an expression product of an isolated and
 manipulated genetic sequence is an "isolated polypeptide," as used herein,
 even if expressed in a homologous cell type. Synthetically made forms or
 molecules expressed by heterologous cells are inherently isolated
 molecules.
 "Recombinant nucleic acid" is a nucleic acid which is not naturally
 occurring, or which is made by the artificial combination of two otherwise
 separated segments of sequence. This artificial combination is often
 accomplished by either chemical synthesis means, or by the artificial
 manipulation of isolated segments of nucleic acids, e.g., by genetic
 engineering techniques. Such is usually done to replace a codon with a
 redundant codon encoding the same or a conservative amino acid, while
 typically introducing or removing a sequence recognition site.
 Alternatively, it is performed to join together nucleic acid segments of
 desired functions to generate a desired combination of functions.
 "Regulatory sequences" refers to those sequences normally within 100 kb of
 the coding region of a locus, but they may also be more distant from the
 coding region, which affect the expression of the gene (including
 transcription of the gene, and translation, splicing, stability or the
 like of the messenger RNA).
 "Substantial homology or similarity". A nucleic acid or fragment thereof is
 "substantially homologous" ("or substantially similar") to another if,
 when optimally aligned (with appropriate nucleotide insertions or
 deletions) with the other nucleic acid (or its complementary strand),
 there is nucleotide sequence identity in at least about 60% of the
 nucleotide bases, usually at least about 70%, more usually at least about
 80%, preferably at least about 90%, and more preferably at least about
 95-98% of the nucleotide bases.
 Alternatively, substantial homology or (similarity) exists when a nucleic
 acid or fragment thereof will hybridize to another nucleic acid (or a
 complementary strand thereof) under selective hybridization conditions, to
 a strand, or to its complement. Selectivity of hybridization exists when
 hybridization which is substantially more selective than total lack of
 specificity occurs. Typically, selective hybridization will occur when
 there is at least about 55% homology over a stretch of at least about 14
 nucleotides, preferably at least about 65%, more preferably at least about
 75%, and most preferably at least about 90%. See, Kanehisa, 1984. The
 length of homology comparison, as described, may be over longer stretches,
 and in certain embodiments will often be over a stretch of at least about
 nine nucleotides, usually at least about 20 nucleotides, more usually at
 least about 24 nucleotides, typically at least about 28 nucleotides, more
 typically at least about 32 nucleotides, and preferably at least about 36
 or more nucleotides.
 Nucleic acid hybridization will be affected by such conditions as salt
 concentration, temperature, or organic solvents, in addition to the base
 composition, length of the complementary strands, and the number of
 nucleotide base mismatches between the hybridizing nucleic acids, as will
 be readily appreciated by those skilled in the art. Stringent temperature
 conditions will generally include temperatures in excess of 30.degree. C.,
 typically in excess of 37.degree. C., and preferably in excess of
 45.degree. C. Stringent salt conditions will ordinarily be less than 1000
 mM, typically less than 500 mM, and preferably less than 200 mM. However,
 the combination of parameters is much more important than the measure of
 any single parameter. The stringency conditions are dependent on the
 length of the nucleic acid and the base composition of the nucleic acid
 and can be determined by techniques well known in the art. See, e.g.,
 Wetmur and Davidson, 1968.
 Probe sequences may also hybridize specifically to duplex DNA under certain
 conditions to form triplex or other higher order DNA complexes. The
 preparation of such probes and suitable hybridization conditions are well
 known in the art.
 The terms "substantial homology" or "substantial identity", when referring
 to polypeptides, indicate that the polypeptide or protein in question
 exhibits at least about 30% identity with an entire naturally-occurring
 protein or a portion thereof, usually at least about 70% identity, more
 usually at least about 80% identity, preferably at least about 90%
 identity, and more preferably at least about 95% identity.
 Homology, for polypeptides, is typically measured using sequence analysis
 software. See, e.g., the Sequence Analysis Software Package of the
 Genetics Computer Group, University of Wisconsin Biotechnology Center, 910
 University Avenue, Madison, Wis. 53705. Protein analysis software matches
 similar sequences using measure of homology assigned to various
 substitutions, deletions and other modifications. Conservative
 substitutions typically include substitutions within the following groups:
 glycine, alanine; valine, isoleucine, leucine; aspartic acid, glutamic
 acid; asparagine, glutamine; serine, threonine; lysine, arginine; and
 phenylalanine, tyrosine.
 "Substantially similar function" refers to the function of a modified
 nucleic acid or a modified protein, with reference to the wild-type MMSC2
 nucleic acid or wild-type MMSC2 polypeptide. The modified polypeptide will
 be substantially homologous to the wild-type MMSC2 polypeptide and will
 have substantially the same function. The modified polypeptide may have an
 altered amino acid sequence and/or may contain modified amino acids. In
 addition to the similarity of function, the modified polypeptide may have
 other useful properties, such as a longer half-life. The similarity of
 function (activity) of the modified polypeptide may be substantially the
 same as the activity of the wild-type MMSC2 polypeptide. Alternatively,
 the similarity of function (activity) of the modified polypeptide may be
 higher than the activity of the wild-type MMSC2 polypeptide. The modified
 polypeptide is synthesized using conventional techniques, or is encoded by
 a modified nucleic acid and produced using conventional techniques. The
 modified nucleic acid is prepared by conventional techniques. A nucleic
 acid with a function substantially similar to the wild-type MMSC2 gene
 function produces the modified protein described above.
 A polypeptide "fragment," "portion" or "segment" is a stretch of amino acid
 residues of at least about five to seven contiguous amino acids, often at
 least about seven to nine contiguous amino acids, typically at least about
 nine to 13 contiguous amino acids and, most preferably, at least about 20
 to 30 or more contiguous amino acids.
 The polypeptides of the present invention, if soluble, may be coupled to a
 solid-phase support, e.g., nitrocellulose, nylon, column packing materials
 (e.g., Sepharose beads), magnetic beads, glass wool, plastic, metal,
 polymer gels, cells, or other substrates. Such supports may take the form,
 for example, of beads, wells, dipsticks, or membranes.
 "Target region" refers to a region of the nucleic acid which is amplified
 and/or detected. The term "target sequence" refers to a sequence with
 which a probe or primer will form a stable hybrid under desired
 conditions.
 The practice of the present invention employs, unless otherwise indicated,
 conventional techniques of chemistry, molecular biology, microbiology,
 recombinant DNA, genetics, and immunology. See, e.g., Maniatis et al.,
 1982; Sambrook et al., 1989; Ausubel et al., 1992; Glover, 1985; Anand,
 1992; Guthrie and Fink, 1991. A general discussion of techniques and
 materials for human gene mapping, including mapping of human chromosome 1,
 is provided, e.g., in White and Lalouel, 1988.
 Preparation of Recombinant or Chemically Synthesized Nucleic Acids;
 Vectors, Transformation, Host Cells
 Large amounts of the polynucleotides of the present invention may be
 produced by replication in a suitable host cell. Natural or synthetic
 polynucleotide fragments coding for a desired fragment will be
 incorporated into recombinant polynucleotide constructs, usually DNA
 constructs, capable of introduction into and replication in a prokaryotic
 or eukaryotic cell. Usually the polynucleotide constructs will be suitable
 for replication in a unicellular host, such as yeast or bacteria, but may
 also be intended for introduction to (with and without integration within
 the genome) cultured mammalian, plant, insect or other eukaryotic cell
 lines. The purification of nucleic acids produced by the methods of the
 present invention are described, e.g., in Sambrook et al., 1989 or Ausubel
 et al., 1992.
 The polynucleotides of the present invention may also be produced by
 chemical synthesis, e.g., by the phosphoramidite method described by
 Beaucage and Carruthers, 1981 or the triester method according to
 Matteucci and Caruthers, 1981, and may be performed on commercial,
 automated oligonucleotide synthesizers. A double-stranded fragment may be
 obtained from the single-stranded product of chemical synthesis either by
 synthesizing the complementary strand and annealing the strand together
 under appropriate conditions or by adding the complementary strand using
 DNA polymerase with an appropriate primer sequence.
 Polynucleotide constructs prepared for introduction into a prokaryotic or
 eukaryotic host may comprise a replication system recognized by the host,
 including the intended polynucleotide fragment encoding the desired
 polypeptide, and will preferably also include transcription and
 translational initiation regulatory sequences operably linked to the
 polypeptide encoding segment. Expression vectors may include, for example,
 an origin of replication or autonomously replicating sequence (ARS) and
 expression control sequences, a promoter, an enhancer and necessary
 processing information sites, such as ribosome-binding sites, RNA splice
 sites, polyadenylation sites, transcriptional terminator sequences, and
 mRNA stabilizing sequences. Such vectors may be prepared by means of
 standard recombinant techniques well known in the art and discussed, for
 example, in Sambrook et al., 1989 or Ausubel et al., 1992.
 An appropriate promoter and other necessary vector sequences will be
 selected so as to be functional in the host, and may include, when
 appropriate, those naturally associated with the MMSC2 gene. Examples of
 workable combinations of cell lines and expression vectors are described
 in Sambrook et al., 1989 or Ausubel et al., 1992; see also, e.g., Metzger
 et al., 1988. Many useful vectors are known in the art and may be obtained
 from such vendors as Stratagene, New England Biolabs, Promega Biotech, and
 others. Promoters such as the trp, lac and phage promoters, tRNA promoters
 and glycolytic enzyme promoters may be used in prokaryotic hosts. Useful
 yeast promoters include promoter regions for metallothionein,
 3-phosphoglycerate kinase or other glycolytic enzymes such as enolase or
 glyceraldehyde-3-phosphate dehydrbgenase, enzymes responsible for maltose
 and galactose utilization, and others. Vectors and promoters suitable for
 use in yeast expression are further described in Hitzeman et al. (EP
 73,675A). Appropriate non-native mammalian promoters might include the
 early and late promoters from SV40 (Fiers et al., 1978) or promoters
 derived from murine Molony leukemia virus, mouse tumor virus, avian
 sarcoma viruses, adenovirus II, bovine papilloma virus or polyoma. Insect
 promoters may be derived from baculovirus. In addition, the construct may
 be joined to an amplifiable gene (e.g., DHFR) so that multiple copies of
 the gene may be made. For appropriate enhancer and other expression
 control sequences, see also Enhancers and Eukaryotic Gene Expression, Cold
 Spring Harbor Press, Cold Spring Harbor, N.Y. (1983). See also, e.g., U.S.
 Pat. No. 5,691,198.
 While such expression vectors may replicate autonomously, they may also
 replicate by being inserted into the genome of the host cell, by methods
 well known in the art.
 Expression and cloning vectors will likely contain a selectable marker, a
 gene encoding a protein necessary for survival or growth of a host cell
 transformed with the vector. The presence of this gene ensures growth of
 only those host cells which express the inserts. Typical selection genes
 encode proteins that a) confer resistance to antibiotics or other toxic
 substances, e.g. ampicillin, neomycin, methotrexate, etc., b) complement
 auxotrophic deficiencies, or c) supply critical nutrients not available
 from complex media, e.g., the gene encoding D-alanine racemase for
 Bacilli. The choice of the proper selectable marker will depend on the
 host cell, and appropriate markers for different hosts are well known in
 the art.
 The vectors containing the nucleic acids of interest can be transcribed in
 vitro, and the resulting RNA introduced into the host cell by well-known
 methods, e.g., by injection (see, Kubo et al., 1988), or the vectors can
 be introduced directly into host cells by methods well known in the art,
 which vary depending on the type of cellular host, including
 electroporation; transfection employing calcium chloride, rubidium
 chloride calcium phosphate, DEAE-dextran, or other substances;
 microprojectile bombardment; lipofection; infection (where the vector is
 an infectious agent, such as a retroviral genome); and other methods. See
 generally, Sambrook et al., 1989 and Ausubel et al., 1992. The
 introduction of the polynucleotides into the host cell by any method known
 in the art, including, inter alia, those described above and in U.S. Pat.
 No. 5,691,198, will be referred to herein as "transformation." The cells
 into which have been introduced nucleic acids described above are meant to
 also include the progeny of such cells.
 Large quantities of the nucleic acids and polypeptides of the present
 invention may be prepared by expressing the MMSC2 nucleic acid or portions
 thereof in vectors or other expression vehicles in compatible prokaryotic
 or eukaryotic host cells. The most commonly used prokaryotic hosts are
 strains of Escherichia coli, although other prokaryotes, such as Bacillus
 subtilis or Pseudomonas may also be used.
 Mammalian or other eukaryotic host cells, such as those of yeast,
 filamentous fungi, plant, insect, or amphibian or avian species, may also
 be useful for production of the proteins of the present invention.
 Propagation of mammalian cells in culture is per se well known. See,
 Jakoby and Pastan (eds.), 1979. Examples of commonly used mammalian host
 cell lines are VERO and HeLa cells, Chinese hamster ovary (CHO) cells, and
 WI38, BHK, and COS cell lines. An example of a commonly used insect cell
 line is SF9. However, it will be appreciated by the skilled practitioner
 that other cell lines may be appropriate, e.g., to provide higher
 expression, desirable glycosylation patterns, or other features.
 Clones are selected by using markers depending on the mode of the vector
 construction. The marker may be on the same or a different DNA molecule,
 preferably the same DNA molecule. In prokaryotic hosts, the transformant
 may be selected, e.g., by resistance to ampicillin, tetracycline or other
 antibiotics. Production of a particular product based on temperature
 sensitivity may also serve as an appropriate marker.
 Prokaryotic or eukaryotic cells transformed with the polynucleotides of the
 present invention will be useful not only for the production of the
 nucleic acids and polypeptides of the present invention, but also, for
 example, in studying the characteristics of MMSC2 polypeptide.
 The probes and primers based on the MMSC2 gene sequence disclosed herein
 are used to identify homologous MMSC2 gene sequences and proteins in other
 species. These gene sequences and proteins are used in the
 diagnostic/prognostic, therapeutic and drug screening methods described
 herein for the species from which they have been isolated.
 Methods of Use: Drug Screening
 This invention is particularly useful for screening compounds by using the
 MMSC2 polypeptide or binding fragment thereof in any of a variety of drug
 screening techniques, such as those described herein and in published PCT
 application WO 97/02048. Since MMSC2 acts as a scaffold that binds to MMAC
 1, the phosphatase substrate(s) and the (probably oncogene) tyrosine
 kinase(s), a valuable drug candidate will be a drug that can prevent
 binding of either the substrate(s) or the tyrosine kinase(s) to MMSC2.
 The MMSC2 polypeptide or fragment employed in such a test may either be
 free in solution, affixed to a solid support, or borne on a cell surface.
 One method of drug screening utilizes eukaryotic or procaryotic host cells
 which are stably transformed with recombinant polynucleotides expressing
 the polypeptide or fragment, preferably in competitive binding assays.
 Such cells, either in viable or fixed form, can be used for standard
 binding assays. One may measure, for example, for the formation of
 complexes between a MMSC2 polypeptide or fragment and the agent being
 tested, or examine the degree to which the formation of a complex between
 a MMSC2 polypeptide or fragment and a known ligand, e.g., MMAC1, is aided
 or interfered with by the agent being tested.
 Thus, the present invention provides methods of screening for drugs
 comprising contacting such an agent with a MMSC2 polypeptide or fragment
 thereof and assaying (i) for the presence of a complex between the agent
 and the MMSC2 polypeptide or fragment, or (ii) for the presence of a
 complex between the MMSC2 polypeptide or fragment and a ligand, by methods
 well known in the art. In such competitive binding assays the MMSC2
 polypeptide or fragment is typically labeled. Free MMSC2 polypeptide or
 fragment is separated from that present in a protein:protein complex, and
 the amount of free (i.e., uncomplexed) label is a measure of the binding
 of the agent being tested to MMSC2 or its interference with or promotion
 of MMSC2:ligand binding, respectively. One may also measure the amount of
 bound, rather than free, MMSC2. It is also possible to label the ligand
 rather than the MMSC2 and to measure the amount of ligand binding to MMSC2
 in the presence and in the absence of the drug being tested.
 Another technique for drug screening provides high throughput screening for
 compounds having suitable binding affinity to the MMSC2 polypeptides and
 is described in detail in Geysen (published PCT application WO 84/03564).
 Briefly stated, large numbers of different small peptide test compounds
 are synthesized on a solid substrate, such as plastic pins or some other
 surface. The peptide test compounds are reacted with MMSC2 polypeptide and
 washed. Bound MMSC2 polypeptide is then detected by methods well known in
 the art.
 Purified MMSC2 can be coated directly onto plates for use in the
 aforementioned drug screening techniques. However, non-neutralizing
 antibodies to the polypeptide can be used to capture antibodies to
 immobilize the MMSC2 polypeptide on the solid phase.
 This invention also contemplates the use of competitive drug screening
 assays in which neutralizing antibodies capable of specifically binding
 the MMSC2 polypeptide compete with a test compound for binding to the
 MMSC2 polypeptide or fragments thereof. In this manner, the antibodies can
 be used to detect the presence of any peptide which shares one or more
 antigenic determinants of the MMSC2 polypeptide.
 The above screening methods are not limited to assays employing only MMSC2
 but are also applicable to studying MMSC2-protein complexes, e.g., the
 complex which occurs between MMSC2 and MMAC1. The effect of drugs on the
 activity of this complex, especially when either the MMSC2 or the MMSC2
 binding protein (e.g., MMAC1) contains a mutation, is analyzed.
 In accordance with these methods, the following assays are examples of
 assays which can be used for screening for drug candidates.
 A mutant MMSC2 (per se or as part of a fusion protein) is mixed with a
 wild-type protein (per se or as part of a fusion protein) to which
 wild-type MMSC2 binds. This mixing is performed in both the presence of a
 drug and the absence of the drug, and the amount of binding of the mutant
 MMSC2 with the wild-type protein is measured. If the amount of the binding
 is more in the presence of said drug than in the absence of said drug, the
 drug is a drug candidate for treating cancer resulting from a mutation in
 MMSC2. This assay is useful where the wild-type protein is a tumor
 suppressor, such as MMAC1.
 A wild-type MMSC2 (per se or as part of a fusion protein) is mixed with a
 wild-type protein (per se or as part of a fusion protein) to which
 wild-type MMSC2 binds. This mixing is performed in both the presence of a
 drug and the absence of the drug, and the amount of binding of the
 wild-type MMSC2 with the wild-type protein is measured. If the amount of
 the binding is more in the presence of said drug than in the absence of
 said drug, the drug is a drug candidate for treating cancer resulting from
 a mutation in MMSC2. This assay is useful where the wild-type protein is a
 tumor suppressor, such as MMAC1.
 A mutant MMSC2 (per se or as part of a fusion protein) is mixed with a
 wild-type protein (per se or as part of a fusion protein) to which
 wild-type MMSC2 binds. This mixing is performed in both the presence of a
 drug and the absence of the drug, and the amount of binding of the mutant
 MMSC2 with the wild-type protein is measured. If the amount of the binding
 is less in the presence of said drug than in the absence of said drug, the
 drug is a drug candidate for treating cancer resulting from a mutation in
 MMSC2. This assay is useful if the protein is an oncoprotein or a
 substrate of the oncoprotein.
 A wild-type MMSC2 (per se or as part of a fusion protein) is mixed with a
 wild-type protein (per se or as part of a fusion protein) to which
 wild-type MMSC2 binds. This mixing is performed in both the presence of a
 drug and the absence of the drug, and the amount of binding of the
 wild-type MMSC2 with the wild-type protein is measured. If the amount of
 the binding is less in the presence of said drug than in the absence of
 said drug, the drug is a drug candidate for treating cancer resulting from
 a mutation in MMSC2 or a cancer resulting from a mutation in MMAC1. This
 assay is useful if the protein is an oncoprotein or a substrate of the
 oncoprotein.
 A mutant protein, which as a wild-type protein binds to MMSC2 (per se or as
 part of a fusion protein) is mixed with a wild-type MMSC2 (per se or as
 part of a fusion protein). This mixing is performed in both the presence
 of a drug and the absence of the drug, and the amount of binding of the
 mutant protein with the wild-type MMSC2 is measured. If the amount of the
 binding is less in the presence of said drug than in the absence of said
 drug, the drug is a drug candidate for treating cancer resulting from a
 mutation in the gene encoding the protein.
 The polypeptide of the invention may also be used for screening compounds
 developed as a result of combinatorial library technology. Combinatorial
 library technology provides an efficient way of testing a potential vast
 number of different substances for ability to modulate activity of a
 polypeptide. Such libraries and their use are known in the art. The use of
 peptide libraries is preferred. See, for example, WO 97/02048.
 Briefly, a method of screening for a substance which modulates activity of
 a polypeptide may include contacting one or more test substances with the
 polypeptide in a suitable reaction medium, testing the activity of the
 treated polypeptide and comparing that activity with the activity of the
 polypeptide in comparable reaction medium untreated with the test
 substance or substances. A difference in activity between the treated and
 untreated polypeptides is indicative of a modulating effect of the
 relevant test substance or substances.
 Prior to or as well as being screened for modulation of activity, test
 substances may be screened for ability to interact with the polypeptide,
 e.g., in a yeast two-hybrid system (e.g., Bartel et al., 1993). This
 system may be used as a coarse screen prior to testing a substance for
 actual ability to modulate activity of the polypeptide. Alternatively, the
 screen could be used to screen test substances for binding to a MMSC2
 specific binding partner, such as MMAC1, or to find mimetics of the MMSC2
 polypeptide.
 Following identification of a substance which modulates or affects
 polypeptide activity, the substance may be investigated further.
 Furthermore, it may be manufactured and/or used in preparation, i.e.,
 manufacture or formulation, or a composition such as a medicament,
 pharmaceutical composition or drug. These may be administered to
 individuals.
 Thus, the present invention extends in various aspects not only to a
 substance identified using a nucleic acid molecule as a modulator of
 polypeptide activity, in accordancewith what is disclosed herein, but also
 a pharmaceutical composition, medicament, drug or other composition
 comprising such a substance, a method comprising administration of such a
 composition comprising such a substance, a method comprising
 administration of such a composition to a patient, e.g., for treatment
 (which may include preventative treatment) of cancer, use of such a
 substance in the manufacture of a composition for administration, e.g.,
 for treatment of cancer, and a method of making a pharmaceutical
 composition comprising admixing such a substance with a pharmaceutically
 acceptable excipient, vehicle or carrier, and optionally other
 ingredients. A substance identified using as a modulator of polypeptide
 function may be peptide or non-peptide in nature. Non-peptide "small
 molecules" are often preferred for many in vivo pharmaceutical uses.
 Accordingly, a mimetic or mimic of the substance (particularly if a
 peptide) may be designed for pharmaceutical use.
 The designing of mimetics to a known pharmaceutically active compound is a
 known approach to the development of pharmaceuticals based on a "lead"
 compound. This might be desirable where the active compound is difficult
 or expensive to synthesize or where it is unsuitable for a particular
 method of administration, e.g., peptides are unsuitable active agents for
 oral compositions as they tend to be quickly degraded by proteases in the
 alimentary canal. Mimetic design, synthesis and testing is generally used
 to avoid randomly screening large numbers of molecules for a target
 property.
 There are several steps commonly taken in the design of a mimetic from a
 compound having a given target property. First, the particular parts of
 the compound that are critical and/or important in determining the target
 property are determined. In the case of a peptide, this can be done by
 systematically varying the mino acid residues in the peptide, e.g., by
 substituting each residue in turn. Alanine scans of peptide are commonly
 used to refine such peptide motifs. These parts or residues constituting
 the active region of the compound are known as its "pharmacophore".
 Once the pharmacophore has been found, its structure is modeled according
 to its physical properties, e.g., stereochemistry, bonding, size and/or
 charge, using data from a range of sources, e.g., spectroscopic
 techniques, x-ray diffraction data and NMR. Computational analysis,
 similarity mapping (which models the charge and/or volume of a
 pharmacophore, rather than the bonding between atoms) and other techniques
 can be used in this modeling process.
 In a variant of this approach, the three-dimensional structure of the
 ligand and its binding partner are modeled. This can be especially useful
 where the ligand and/or binding partner change conformation on binding,
 allowing the model to take account of this in the design of the mimetic.
 A template molecule is then selected onto which chemical groups which mimic
 the pharmacophore can be grafted. The template molecule and the chemical
 groups grafted onto it can conveniently be selected so that the mimetic is
 easy to synthesize, is likely to be pharmacologically acceptable, and does
 not degrade in vivo, while retaining the biological activity of the lead
 compound. Alternatively, where the mimetic is peptide-based, further
 stability can be achieved by cyclizing the peptide, increasing its
 rigidity. The mimetic or mimetics found by this approach can then be
 screened to see whether they have the target property, or to what extent
 they exhibit it. Further optimization or modification can then be carried
 out to arrive at one or more final mimetics for in vivo or clinical
 testing.
 Methods of Use: Nucleic Acid Diagnosis and Diagnostic Kits
 In order to detect the presence of a MMSC2 allele predisposing an
 individual to cancer, a biological sample such as blood is prepared and
 analyzed for the presence or absence of susceptibility alleles of MMSC2.
 In order to detect the presence of cancer or as a prognostic indicator, a
 biological sample is prepared and analyzed for the presence or absence of
 mutant alleles of MMSC2. Results of these tests and interpretive
 information are returned to the health care provider for communication to
 the tested individual. Such diagnoses may be performed by diagnostic
 laboratories, or, alternatively, diagnostic kits are manufactured and sold
 to health care providers or to private individuals for self-diagnosis.
 Initially, the screening method involves amplification of the relevant
 MMSC2 sequences. In another preferred embodiment of the invention, the
 screening method involves a non-PCR based strategy. Such screening methods
 include two-step label amplification methodologies that arc well known in
 the art. Both PCR and non-PCR based screening strategies can detect target
 sequences with a high level of sensitivity.
 The most popular method used today is target amplification. Here, the
 target nucleic acid sequence is amplified with polymerases. One
 particularly preferred method using polymerase-driven amplification is the
 polymerase chain reaction (PCR). The polymerase chain reaction and other
 polymerase-driven amplification assays can achieve over a million-fold
 increase in copy number through the use of polymerase-driven amplification
 cycles. Once amplified, the resulting nucleic acid can be sequenced or
 used as a substrate for DNA probes.
 When the probes are used to detect the presence of the target sequences the
 biological sample to be analyzed, such as blood or serum, may be treated,
 if desired, to extract the nucleic acids. The sample nucleic acid may be
 prepared in various ways to facilitate detection of the target sequence,
 e.g., denaturation, restriction digestion, electrophoresis or dot
 blotting. The targeted region of the analyte nucleic acid usually must be
 at least partially single-stranded to form hybrids with the targeting
 sequence of the probe. If the sequence is naturally single-stranded,
 denaturation will not be required. However, if the sequence is
 double-stranded, the sequence will probably need to be denatured.
 Denaturation can be carried out by various techniques known in the art.
 Analyte nucleic acid and probe are incubated under conditions which promote
 stable hybrid formation of the target sequence in the probe with the
 putative targeted sequence in the analyte. The region of the probes which
 is used to bind to the analyte can be made completely complementary to the
 targeted region for MMSC2. Therefore, high stringency conditions are
 desirable in order to prevent false positives. However, conditions of high
 stringency are used only if the probes are complementary to regions of the
 chromosome which are unique in the genome. The stringency of hybridization
 is determined by a number of factors during hybridization and during the
 washing procedure, including temperature, ionic strength, base
 composition, probe length, and concentration of formamide. These factors
 are outlined in, for example, Maniatis et al., 1982 and Sambrook et al.,
 1989. Under certain circumstances, the formation of higher order hybrids,
 such as triplexes, quadraplexes, etc., may be desired to provide the means
 of detecting target sequences.
 Detection, if any, of the resulting hybrid is usually accomplished by the
 use of labeled probes. Alternatively, the probe may be unlabeled, but may
 be detectable by specific binding with a ligand which is labeled, either
 directly or indirectly. Suitable labels, and methods for labeling probes
 and ligands are known in the art, and include, for example, radioactive
 labels which may be incorporated by known methods (e.g., nick translation,
 random priming or kinasing), biotin, fluorescent groups, chemiluminescent
 groups (e.g., dioxetanes, particularly triggered dioxetanes), enzymes,
 antibodies, gold nanoparticles and the like. Variations of this basic
 scheme are known in the art, and include those variations that facilitate
 separation of the hybrids to be detected from extraneous materials and/or
 that amplify the signal from the labeled moiety. A number of these
 variations are reviewed in, e.g., Matthews and Kricka, 1988; Landegren et
 al., 1988; U.S. Pat. No. 4,868,105; and in EP 225,807A.
 As noted above, non-PCR based screening assays are also contemplated in
 this invention. This procedure hybridizes a nucleic acid probe (or an
 analog such as a methyl phosphonate backbone replacing the normal
 phosphodiester), to the low level DNA target. This probe may have an
 enzyme covalently linked to the probe, such that the covalent linkage does
 not interfere with the specificity of the hybridization. This
 enzyme-probe-conjugate-target nucleic acid complex can then be isolated
 away from the free probe enzyme conjugate and a substrate is added for
 enzyme detection. Enzymatic activity is observed as a change in color
 development or luminescent output resulting in a 10.sup.3 -10.sup.6
 increase in sensitivity. For an example relating to the preparation of
 oligodeoxynucleotide-alkaline phosphatase conjugates and their use as
 hybridization probes, see Jablonski et al., 1986.
 Two-step label amplification methodologies are known in the art. These
 assays work on the principle that a small ligand (such as digoxigenin,
 biotin, or the like) is attached to a nucleic acid probe capable of
 specifically binding MMSC2. Allele specific probes are also contemplated
 within the scope of this example and exemplary allele specific probes
 include probes encompassing the predisposing mutations of this disclosure.
 In one example, the small ligand attached to the nucleic acid probe is
 specifically recognized by an antibody-enzyme conjugate. In one embodiment
 of this example, digoxigenin is attached to the nucleic acid probe.
 Hybridization is detected by an antibody-alkaline phosphatase conjugate
 which turns over a chemiluminescent substrate. For methods for labeling
 nucleic acid probes according to this embodiment see Martin et al., 1990.
 In a second example, the small ligand is recognized by a second
 ligand-enzyme conjugate that is capable of specifically complexing to the
 first ligand. A well known embodiment of this example is the biotin-avidin
 type of interactions. For methods for labeling nucleic acid probes and
 their use in biotin-avidin based assays see Rigby et al., 1977 and Nguyen
 et al., 1992.
 It is also contemplated within the scope of this invention that the nucleic
 acid probe assays of this invention will employ a cocktail of nucleic acid
 probes capable of detecting MMSC2. Thus, in one example to detect the
 presence of MMSC2 in a cell sample, more than one probe complementary to
 the gene is employed and in particular the number of different probes is
 alternatively two, three, or five different nucleic acid probe sequences.
 In another example, to detect the presence of mutations in the MMSC2 gene
 sequence in a patient, more than one probe complementary to these genes is
 employed where the cocktail includes probes capable of binding to the
 allele-specific mutations identified in populations of patients with
 alterations in MMSC2. In this embodiment, any number of probes can be
 used, and will preferably include probes corresponding to the major gene
 mutations identified as predisposing an individual to cancer.
 Methods of Use: Peptide Diagnosis and Diagnostic Kits
 The presence of cancer can also be detected on the basis of the alteration
 of wild-type MMSC2 polypeptide. Such alterations can be determined by
 sequence analysis in accordance with conventional techniques. More
 preferably, antibodies (polyclonal or monoclonal) are used to detect
 differences in, or the absence of MMSC2 peptides. Techniques for raising
 and purifying antibodies are well known in the art and any such techniques
 may be chosen to achieve the preparations claimed in this invention. In a
 preferred embodiment of the invention, antibodies will immunoprecipitate
 MMSC2 proteins from solution as well as react with these proteins on
 Western or immunoblots of polyacrylamide gels. In another preferred
 embodiment, antibodies will detect MMSC2 proteins in paraffin or frozen
 tissue sections, using immunocytochemical techniques.
 Preferred embodiments relating to methods for detecting MMSC2 or its
 mutations include enzyme linked immunosorbent assays (ELISA),
 radioimmunoassays (RIA), immunoradiometric assays (IRMA) and
 immunoenzymatic assays (IEMA), including sandwich assays using monoclonal
 and/or polyclonal antibodies. Exemplary sandwich assays are described by
 David et al., in U.S. Pat. Nos. 4,376,110 and 4,486,530, hereby
 incorporated by reference.
 Alternatively, alterations in the MMSC2 sequence can be determined by
 detecting alterations in the interaction of MMSC2 with MMAC1 or the
 C-terminus of MMAC1. Wild-type MMAC1 or its C-terminus can be bound to a
 solid phase and the interaction with MMSC2 assayed by conventional
 techniques. Analogously, alterations in MMAC1 which affect its interaction
 with MMSC2 can be detected using wild-type MMSC2 or its PDZ domain which
 interacts with MMAC1 bound to a solid phase.
 Methods of Use: Rational Drug Design
 The goal of rational drug design is to produce structural analogs of
 biologically active polypeptides of interest or of small molecules with
 which they interact (e.g., agonists, antagonists, inhibitors) in order to
 fashion drugs which are, for example, more active or stable forms of the
 polypeptide, or which, e.g., enhance or interfere with the function of a
 polypeptide in vivo. See, e.g., Hodgson, 1991. In one approach, one first
 determines the three-dimensional structure of a protein of interest (e.g.,
 MMSC2 polypeptide) by x-ray crystallography, by computer modeling or most
 typically, by a combination of approaches. Less often, useful information
 regarding the structure of a polypeptide may be gained by modeling based
 on the structure of homologous proteins. An example of rational drug
 design is the development of HIV protease inhibitors (Erickson et al.,
 1990). In addition, peptides (e.g., MMSC2 polypeptide) are analyzed by an
 alanine scan (Wells, 1991). In this technique, an amino acid residue is
 replaced by Ala, and its effect on the peptide's activity is determined.
 Each of the amino acid residues of the peptide is analyzed in this manner
 to determine the important regions of the peptide.
 It is also possible to isolate a target-specific antibody, selected by a
 functional assay, and then to solve its crystal structure. In principle,
 this approach yields a pharmacore upon which subsequent drug design can be
 based. It is possible to bypass protein crystallography altogether by
 generating anti-idiotypic antibodies (anti-ids) to a functional,
 pharmacologically active antibody. As a mirror image of a mirror image,
 the binding site of the anti-ids would be expected to be an analog of the
 original receptor. The anti-id could then be used to identify and isolate
 peptides from banks of chemically or biologically produced banks of
 peptides. Selected peptides would then act as the pharmacore.
 Thus, one may design drugs which have, e.g., improved MMSC2 polypeptide
 activity or stability or which act as inhibitors, agonists, antagonists,
 etc. of MMSC2 polypeptide activity. By virtue of the availability of
 cloned MMSC2 sequence, sufficient amounts of the MMSC2 polypeptide may be
 made available to perform such analytical studies as x-ray
 crystallography. In addition, the knowledge of the MMSC2 protein sequence
 provided herein will guide those employing computer modeling techniques in
 place of, or in addition to x-ray crystallography.
 Methods of Use: Gene Therapy
 According to the present invention, a method is also provided of supplying
 wild-type MMSC2 function to a cell which carries a mutant MMSC2 allele.
 Supplying such a function should allow normal functioning of the recipient
 cells. The wild-type gene or a part of the gene may be introduced into the
 cell in a vector such that the gene remains extrachromosomal. In such a
 situation, the gene will be expressed by the cell from the
 extrachromosomal location. More preferred is the situation where the
 wild-type gene or a part thereof is introduced into the mutant cell in
 such a way that it recombines with the endogenous mutant gene present in
 the cell. Such recombination requires a double recombination event which
 results in the correction of the gene mutation. Vectors for introduction
 of genes both for recombination and for extrachromosomal maintenance are
 known in the art, and any suitable vector may be used. Methods for
 introducing DNA into cells such as electroporation, calcium phosphate
 co-precipitation and viral transduction are known in the art, and the
 choice of method is within the competence of the practitioner.
 As generally discussed above, the MMSC2 gene or fragment, where applicable,
 may be employed in gene therapy methods in order to increase the amount of
 the expression products of such gene in cells. It may also be useful to
 increase the level of expression of the MMSC2 gene even in those persons
 in which the mutant gene is expressed at a "normal" level, but the gene
 product is not fully functional.
 Gene therapy would be carried out according to generally accepted methods,
 for example, as described by Friedman (1991) or Culver (1996). Cells from
 a patient would be first analyzed by the diagnostic methods described
 above, to ascertain the production of MMSC2 polypeptide in the cells. A
 virus or plasmid vector (see further details below), containing a copy of
 the MMSC2 gene linked to expression control elements, is prepared. The
 vector may be capable of replicating inside the cells. Alternatively, the
 vector may be replication deficient and is replicated in helper cells for
 use in gene therapy. Suitable vectors are known, such as disclosed in U.S.
 Pat. No. 5,252,479, published PCT application WO 93/07282 and U.S. Pat.
 No. 5,691,198. The vector is then injected into the patient. If the
 transfected gene is not permanently incorporated into the genome of each
 of the targeted cells, the treatment may have to be repeated periodically.
 Gene transfer systems known in the art may be useful in the practice of the
 gene therapy methods of the present invention. These include viral and
 nonviral transfer methods. A number of viruses have been used as gene
 transfer vectors or as the basis for preparing gene transfer vectors,
 including papovaviruses (e.g., SV40, Madzak et al., 1992), adenovirus
 (Berkner, 1992; Berkner et al., 1988; Gorziglia and Kapikian, 1992;
 Quantin et al., 1992; Rosenfeld et al., 1992; Wilkinson et al., 1992;
 Stratford-Perricaudet et al., 1990; Schneider et al., 1998), vaccinia
 virus (Moss, 1992), adeno-associated virus (Muzyczka, 1992; Ohi et al.,
 1990; Russell & Hirata, 1998), herpesviruses including HSV and EBV
 (Margolskee, 1992; Johnson et al., 1992; Fink et al., 1992; Breakfield and
 Geller, 1987; Freese et al., 1990; Fink et al., 1996), lentiviruses
 (Naldini et al., 1996), vaccinia virus (Moss, 1996), Sindbis and Semliki
 Forest virus (Berglund et al., 1993), and retroviruses of avian
 (Brandyopadhyay and Temin, 1984; Petropoulos et al., 1992), murine
 (Miller, 1992; Miller et al., 1985; Sorge et al., 1984; Mann and
 Baltimore, 1985; Miller et al., 1988), and human origin (Shimada et al.,
 1991; Helseth et al., 1990; Page et al., 1990; Buchschacher and
 Panganiban, 1992). Most human gene therapy protocols have been based on
 disabled murine retroviruses, although adenovirus and adeno-associated
 virus are also being used.
 Nonviral gene transfer methods known in the art include chemical techniques
 such as calcium phosphate coprecipitation (Graham and van der Eb, 1973;
 Pellicer et al., 1980); mechanical techniques, for example microinjection
 (Anderson et al., 1980; Gordon et al., 1980; Brinster et al., 1981;
 Constantini and Lacy, 1981); membrane fusion-mediated transfer via
 liposomes (Feigner et al., 1987; Wang and Huang, 1989; Kaneda et al.,
 1989; Stewart et al., 1992; Nabel et al., 1990; Lim et al., 1992); and
 direct DNA uptake and receptor-mediated DNA transfer (Wolff et al., 1990;
 Wu et al., 1991; Zenke et al., 1990; Wu et al., 1989 b; Wolff et al.,
 1991; Wagner et al., 1990; Wagner et al., 1991; Cotten et al., 1990;
 Curiel et al., 1991a; Curiel et al., 1991b).
 In an approach which combines biological and physical gene transfer
 methods, plasmid DNA of any size is combined with a polylysine-conjugated
 antibody specific to the adenovirus hexon protein, and the resulting
 complex is bound to an adenovirus vector. The trimolecular complex is then
 used to infect cells. The adenovirus vector permits efficient binding,
 internalization, and degradation of the endosome before the coupled DNA is
 damaged. For other techniques for the delivery of adenovirus based vectors
 Schneider et al. (1998) and U.S. Pat. No. 5,691,198.
 Liposome/DNA complexes have been shown to be capable of mediating direct in
 vivo gene transfer. While in standard liposome preparations the gene
 transfer process is nonspecific, localized in vivo uptake and expression
 have been reported in tumor deposits, for example, following direct in
 situ administration (Nabel, 1992).
 Expression vectors in the context of gene therapy are meant to include
 those constructs containing sequences sufficient to express a
 polynucleotide that has been cloned therein. In viral expression vectors,
 the construct contains viral sequences sufficient to support packaging of
 the construct. If the polynucleotide encodes MMSC2, expression will
 produce MMSC2. If the polynucleotide encodes an antisense polynucleotide
 or a ribozyme, expression will produce the antisense polynucleotide or
 ribozyme. Thus in this context, expression does not require that a protein
 product be synthesized. In addition to the polynucleotide cloned into the
 expression vector, the vector also contains a promoter functional in
 eukaryotic cells. The cloned polynucleotide sequence is under control of
 this promoter. Suitable eukayotic promoters include those described above.
 The expression vector may also include sequences, such as selectable
 markers and other sequences described herein.
 Gene transfer techniques which target DNA directly to brain tissue is
 preferred. Receptor-mediated gene transfer, for example, is accomplished
 by the conjugation of DNA (usually in the form of covalently closed
 supercoiled plasmid) to a protein ligand via polylysine. Ligands are
 chosen on the basis of the presence of the corresponding ligand receptors
 on the cell surface of the target cell/tissue type. These ligand-DNA
 conjugates can be injected directly into the blood if desired and are
 directed to the target tissue where receptor binding and internalization
 of the DNA-protein complex occurs. To overcome the problem of
 intracellular destruction of DNA, coinfection with adenovirus can be
 included to disrupt endosome function.
 The therapy is as follows: patients who carry a MMSC2 susceptibility allele
 are treated with a gene delivery vehicle such that some or all of their
 brain precursor cells receive at least one additional copy of a functional
 normal MMSC2 allele, respectively. In this step, the treated individuals
 have reduced risk of cancer to the extent that the effect of the
 susceptible allele has been countered by the presence of the normal
 allele.
 Methods of Use: Peptide Therapy
 Peptides which have MMSC2 activity can be supplied to cells which carry a
 mutant or missing MMSC2 allele. Protein can be produced by expression of
 the cDNA sequence in bacteria, for example, using known expression
 vectors. Alternatively, MMSC2 polypeptide can be extracted from
 MMSC2-producing mammalian cells. In addition, the techniques of synthetic
 chemistry can be employed to synthesize MMSC2 protein. Any of such
 techniques can provide the preparation of the present invention which
 comprises the MMSC2 protein. The preparation is substantially free of
 other human proteins. This is most readily accomplished by synthesis in a
 microorganism or in vitro.
 Active MMSC2 molecules can be introduced into cells by microinjection or by
 use of liposomes, for example. Alternatively, some active molecules may be
 taken up by cells, actively or by diffusion. Supply of molecules with
 MMSC2 activity should lead to inhibition of cancer. Other molecules with
 MMSC2 activity (for example, peptides, drugs or organic compounds) may
 also be used to effect such an inhibition. Modified polypeptides having
 substantially similar function are also used for peptide therapy.
 Methods of Use: Transformed Hosts
 Animals for testing therapeutic agents can be selected after mutagenesis of
 whole animals or after treatment of germline cells or zygotes. Such
 treatments include insertion of mutant MMSC2 alleles, usually from a
 second animal species, as well as insertion of disrupted homologous genes.
 Alternatively, the endogenous MMSC2 gene of the animals may be disrupted
 by insertion or deletion mutation or other genetic alterations using
 conventional techniques (Capecchi, 1989; Valancius and Smithies, 1991;
 Hasty el al., 1991; Shinkai et al., 1992; Mombaerts et al., 1992; Philpott
 et al., 1992; Snouwaert et al., 1992; Donehower et al., 1992). After test
 substances have been administered to the animals, the presence of cancer
 must be assessed. If the test substance prevents or suppresses the
 appearance of cancer, then the test substance is a candidate therapeutic
 agent for treatment of cancer. These animal models provide an extremely
 important testing vehicle for potential therapeutic products.
 Methods of Use: Transgenic/Knockout Animals and Models
 In one embodiment of the invention, transgenic animals are produced which
 contain a functional transgene encoding a functional MMSC2 polypeptide or
 variants thereof. Transgenic animals expressing MMSC2 transgenes,
 recombinant cell lines derived from such animals and transgenic embryos
 may be useful in methods for screening for and identifying agents that
 induce or repress function of MMSC2. Transgenic animals of the present
 invention also can be used as models for studying indications such as
 cancers.
 In one embodiment of the invention, a MMSC2 transgene is introduced into a
 non-human host to produce a transgenic animal expressing a human or murine
 MMSC2 gene. The transgenic animal is produced by the integration of the
 transgene into the genome in a manner that permits the expression of the
 transgene. Methods for producing transgenic animals are generally
 described by Wagner and Hoppe (U.S. Pat. No. 4,873,191; which is
 incorporated herein by reference), Brinster et al. 1985; which is
 incorporated herein by reference in its entirety) and in "Manipulating the
 Mouse Embryo; A Laboratory Manual" 2nd edition (eds., Hogan, Beddington,
 Costantimi and Long, Cold Spring Harbor Laboratory Press, 1994; which is
 incorporated herein by reference in its entirety).
 It may be desirable to replace the endogenous MMSC2 by homologous
 recombination between the transgene and the endogenous gene; or the
 endogenous gene may be eliminated by deletion as in the preparation of
 "knock-out" animals. Typically, a MMSC2 gene flanked by genomic sequences
 is transferred by microinjection into a fertilized egg. The microinjected
 eggs are implanted into a host female, and the progeny are screened for
 the expression of the transgene. Transgenic animals may be produced from
 the fertilized eggs from a number of animals including, but not limited to
 reptiles, amphibians, birds, mammals, and fish. Within a particularly
 preferred embodiment, transgenic mice are generated which overexpress
 MMSC2 or express a mutant form of the polypeptide. Alternatively, the
 absence of a MMSC2 in "knock-out" mice permits the study of the effects
 that loss of MMSC2 protein has on a cell in vivo. Knock-out mice also
 provide a model for the development of MMSC2-related cancers.
 Methods for producing knockout animals are generally described by Shastry
 (1995, 1998) and Osterrieder and Wolf (1998). The production of
 conditional knockout animals, in which the gene is active until knocked
 out at the desired time is generally described by Feil et al. (1996),
 Gagneten et al. (1997) and Lobe and Nagy (1998). Each of these references
 is incorporated herein by reference.
 As noted above, transgenic animals and cell lines derived from such animals
 may find use in certain testing experiments. In this regard, transgenic
 animals and cell lines capable of expressing wild-type or mutant MMSC2 may
 be exposed to test substances. These test substances can be screened for
 the ability to enhance wild-type MMSC2 expression and or function or
 impair the expression or function of mutant MMSC2.
 Pharmaceutical Compositions and Routes of Administration
 The MMSC2 polypeptides, antibodies, peptides and nucleic acids of the
 present invention can be formulated in pharmaceutical compositions, which
 are prepared according to conventional pharmaceutical compounding
 techniques. See, for example, Remington's Pharmaceutical Sciences, 18th
 Ed. (1990, Mack Publishing Co., Easton, Pa.). The composition may contain
 the active agent or pharmaceutically acceptable salts of the active agent.
 These compositions may comprise, in addition to one of the active
 substances, a pharmaceutically acceptable excipient, carrier, buffer,
 stabilizer or other materials well known in the art. Such materials should
 be non-toxic and should not interfere with the efficacy of the active
 ingredient. The carrier may take a wide variety of forms depending on the
 form of preparation desired for administration, e.g., intravenous, oral,
 intrathecal, epineural or parenteral.
 For oral administration, the compounds can be formulated into solid or
 liquid preparations such as capsules, pills, tablets, lozenges, melts,
 powders, suspensions or emulsions. In preparing the compositions in oral
 dosage form, any of the usual pharmaceutical media may be employed, such
 as, for example, water, glycols, oils, alcohols, flavoring agents,
 preservatives, coloring agents, suspending agents, and the like in the
 case of oral liquid preparations (such as, for example, suspensions,
 elixirs and solutions); or carriers such as starches, sugars, diluents,
 granulating agents, lubricants, binders, disintegrating agents and the
 like in the case of oral solid preparations (such as, for example,
 powders, capsules and tablets). Because of their ease in administration,
 tablets and capsules represent the most advantageous oral dosage unit
 form, in which case solid pharmaceutical carriers are obviously employed.
 If desired, tablets may be sugar-coated or enteric-coated by standard
 techniques. The active agent can be encapsulated to make it stable to
 passage through the gastrointestinal tract while at the same time allowing
 for passage across the blood brain barrier. See for example, WO 96/11698.
 For parenteral administration, the compound may dissolved in a
 pharmaceutical carrier and administered as either a solution of a
 suspension. Illustrative of suitable carriers are water, saline, dextrose
 solutions, fructose solutions, ethanol, or oils of animal, vegetative or
 synthetic origin. The carrier may also contain other ingredients, for
 example, preservatives, suspending agents, solubilizing agents, buffers
 and the like. When the compounds are being administered intrathecally,
 they may also be dissolved in cerebrospinal fluid.
 The active agent is preferably administered in an therapeutically effective
 amount. The actual amount administered, and the rate and time-course of
 administration, will depend on the nature and severity of the condition
 being treated. Prescription of treatment, e.g. decisions on dosage,
 timing, etc., is within the responsibility of general practitioners or
 spealists, and typically takes account of the disorder to be treated, the
 condition of the individual patient, the site of delivery, the method of
 administration and other factors known to practitioners. Examples of
 techniques and protocols can be found in Remington 's Parmaceutical
 Sciences.
 Alternatively, targeting therapies may be used to deliver the active agent
 more specifically to certain types of cell, by the use of targeting
 systems such as antibodies or cell specific ligands. Targeting may be
 desirable for a variety of reasons, e.g. if the agent is unacceptably
 toxic, or if it would otherwise require too high a dosage, or if it would
 not otherwise be able to enter the target cells.
 Instead of administering these agents directly, they could be produced in
 the target cell, e.g. in a viral vector such as described above or in a
 cell based delivery system such as described in U.S. Pat. No. 5,550,050
 and published PCT application Nos. WO 92/19195, WO 94/25503, WO 95/01203,
 WO 95/05452, WO 96/02286, WO 96/02646, WO 96/40871, WO 96/40959 and WO
 97/12635. designed for implantation in a patient. The vector could be
 targeted to the specific cells to be treated, or it could contain
 regulatory elements which are more tissue specific to the target cells.
 The cell based delivery system is designed to be implanted in a patient's
 body at the desired target site and contains a coding sequence for the
 active agent. Alternatively, the agent could be administered in a
 precursor form for conversion to the active form by an activating agent
 produced in, or targeted to, the cells to be treated. See for example, EP
 425,731A and WO 90/07936.
 The identification of the association between the MMSC2 gene mutations and
 cancer permits the early presymptomatic screening of individuals to
 identify those at risk for developing cancer. To identify such
 individuals, MMSC2 alleles are screened for mutations either directly or
 after cloning the alleles. The alleles are tested for the presence of
 nucleic acid sequence differences from the normal allele using any
 suitable technique, including but not limited to, one of the following
 methods: fluorescent in situ hybridization (FISH), direct DNA sequencing,
 PFGE analysis, Southern blot analysis, single stranded conformation
 analysis (SSCP), linkage analysis, RNase protection assay, allele specific
 oligonucleotide (ASO), dot blot analysis and PCR-SSCP analysis. Also
 useful is the recently developed technique of DNA microchip technology.
 For example, either (1) the nucleotide sequence of both the cloned alleles
 and normal MMSC2 gene or appropriate fragment (coding sequence or genomic
 sequence) are determined and then compared, or (2) the RNA transcripts of
 the MMSC2 gene or gene fragment are hybridized to single stranded whole
 genomic DNA from an individual to be tested, and the resulting
 heteroduplex is treated with Ribonuclease A (RNase A) and run on a
 denaturing gel to detect the location of any mismatches. Two of these
 methods can be carried out according to the following procedures.
 The alleles of the MMSC2 gene in an individual to be tested are cloned
 using conventional techniques. For example, a blood sample is obtained
 from the individual. The genomic DNA isolated from the cells in this
 sample is partially digested to an average fragment size of approximately
 20 kb. Fragments in the range from 18-21 kb are isolated. The resulting
 fragments are ligated into an appropriate vector. The sequences of the
 clones are then determined and compared to the normal MMSC2 gene.
 Alternatively, polymerase chain reactions (PCRs) are performed with primer
 pairs for the 5' region or the exons of the MMSC2 gene. PCRs can also be
 performed with primer pairs based on any sequence of the normal MMSC2
 gene. For example, primer pairs for one of the introns can be prepared and
 utilized. Finally, RT-PCR can also be performed on the mRNA. The amplified
 products are then analyzed by single stranded conformation polymorphisms
 (SSCP) using conventional techniques to identify any differences and these
 are then sequenced and compared to the normal gene sequence.
 Individuals can be quickly screened for common MMSC2 gene variants by
 amplifying the individual's DNA using suitable primer pairs and analyzing
 the amplified product, e.g., by dot-blot hybridization using
 allele-specific oligonucleotide probes.
 The second method employs RNase A to assist in the detection of differences
 between the normal MMSC2 gene and defective genes. This comparison is
 performed in steps using small (.about.500 bp) restriction fragments of
 the MMSC2 gene as the probe. First, the MMSC2 gene is digested with a
 restriction enzyme(s) that cuts the gene sequence into fragments of
 approximately 500 bp. These fragments are separated on an electrophoresis
 gel, purified from the gel and cloned individually, in both orientations,
 into an SP6 vector (e.g., pSP64 or pSP65).
 The SP6-bascd plasmids containing inserts of the MMSC2 gene fragments are
 transcribed in vitro using the SP6 transcription system, well known in the
 art, in the presence of [.alpha.-.sup.32 P]GTP, generating radiolabeled
 RNA transcripts of both strands of the gene.
 Individually, these RNA transcripts are used to form heteroduplexes with
 the allelic DNA using conventional techniques. Mismatches that occur in
 the RNA:DNA heteroduplex, owing to sequence differences between the MMSC2
 fragment and the MMSC2 allele subclone from the individual, result in
 cleavage in the RNA strand when treated with RNase A. Such mismatches can
 be the result of point mutations or small deletions in the individual's
 allele. Cleavage of the RNA strand yields two or more small RNA fragments,
 which run faster on the denaturing gel than the RNA probe itself.
 Any differences which are found, will identify an individual as having a
 molecular variant of the MMSC2 gene and the consequent presence of cancer.
 These variants can take a number of forms. The most severe forms would be
 frame shift mutations or large deletions which would cause the gene to
 code for an abnormal protein or one which would significantly alter
 protein expression. Less severe disruptive mutations would include small
 in-frame deletions and nonconservative base pair substitutions which would
 have a significant effect on the protein produced, such as changes to or
 from a cysteine residue, from a basic to an acidic amino acid or vice
 versa, from a hydrophobic to hydrophilic amino acid or vice versa, or
 other mutations which would affect secondary or tertiary protein
 structure. Silent mutations or those resulting in conservative amino acid
 substitutions would not generally be expected to disrupt protein function.
 Genetic testing will enable practitioners to identify individuals at risk
 for cancer at, or even before, birth. Finally, this invention changes our
 understanding of the cause and treatment of cancer.
 EXAMPLES
 The present invention is further detailed in the following Examples, which
 are offered by way of illustration and are not intended to limit the
 invention in any manner. Standard techniques well known in the art or the
 techniques specifically described below are utilized.
 Example 1
 Identification of MMSC2
 A yeast two-hybrid assay was performed using conventional techniques, such
 as described by Fields and Song (1989), Chevray and Nathans (1992), Bartel
 et al. (1993) and Lee et al. (1995). Sequence encoding the C-terminal 15
 amino acids of MMAC1 (NEPFDEDQHTQITKV; SEQ ID NO:4) plus its stop codon
 was generated using an oligonucleotide synthesizer and was ligated to
 plasmid pGBT.C such that the coding sequence of MMAC1 was in-frame with
 coding sequence for the Gal4p DNA-binding domain. This plasmid construct
 was introduced into the yeast reporter strain J692 along with a library of
 activation domain fusion plasmids prepared from human kidney cDNA
 (Clontech). Transformants were spread onto 20-150 mm plates of yeast
 minimal media lacking leucine, tryptophan, and histidine, and containing
 25 mM 3-amino-1,2,4-triazole (Gietz et al., 1995; Bartel and Fields,
 1995). After one week incubation at 30.degree. C., yeast colonies were
 assayed for expression of the lacZ reporter gene by beta-galactosidase
 filter assay (Breeden and Naysmyth, 1985). Colonies that both grew in the
 absence of histidine and were positive for production of
 beta-galactosidase were chosen for further characterization.
 The activation domain plasmid was purified from positive colonies by the
 smash-and-grab technique. These plasmids were introduced into E. coli DH
 10B (Gibco BRL) by electroporation and purified from E. coli by the
 alkaline lysis method. To test for the specificity of the interaction,
 specific activation domain plasmids were cotransformed into strain J692
 with plasmids encoding various DNA-binding domain fusion proteins,
 including fusions to C-terminal segments of MMAC1 and human lamin C.
 Transformants from these experiments were assayed for expression of the
 HIS3 and lcZ reporter genes. Positives that expressed reporter genes with
 MMAC1 constructs and not with lamin C constructs encode bona fide
 MMAC1-interacting proteins. These proteins were identified and
 characterized by sequence analysis of the insert of the appropriate
 activation domain plasmid.
 Five of the clones encoding bona fide MMAC1-interacting proteins were named
 PDZBN2B, PDZBN3A, PDZBN5B, PDZBN18D, and pdzk4. Comparison of the
 sequences of these clones suggested that they were all partial cDNAs
 derived from the same novel gene. A search of GenBank with these sequences
 revealed strong sequence similarity with a partial mouse cDNA sequence
 called 9ORF binding protein 1 (9BP-1)(GenBank Accession #AF000168).
 Several rounds of cDNA library screening were required to identify eDNA
 clones that could be assembled into the full length MMSC2 sequence. In the
 first round, a 509 base pair(bp) probe was developed from the 5' end of
 clone PDZBN2B using primers
 9BP-1 F1: AGACAGCAAAGATGACAGTAA (SEQ ID NO:5) and
 9BP-1 R4: CTTCCTCCTCTTTGTATGGG (SEQ ID NO:6).
 This probe was used to screen a human placental cDNA library and a human
 prostate cDNA library. Two of the informative clones obtained were
 p118a(placental) and pr63(prostate). A search of GenBank with this
 additional sequence yielded an additional human EST (GenBank Accesion
 #C75629). For the second round of cDNA library screening, a 202 bp probe
 was developed from the 5' end of this EST using primers
 9BP1 #2: GCTTTTGCCGAAATGGGTAGT (SEQ ID NO:7) and
 9BP1 #2: GATCGGTCTTTGTTCCCAGCA (SEQ ID NO:8).
 This probe was used to screen a human prostate cDNA library; two of the
 informative clones obtained were clone #10 and clone #3. For the third
 round of cDNA library screening, a 172 bp probe was developed from the 5'
 end of clone #3 using primers
 9BP-1 #5: TGTGAGCAAGTTTAGTGAG (SEQ ID NO:9) and
 9BP-1 #7: GGTGATTTTCCCCAAGTAA (SEQ ID NO: 10)
 and used to screen a human prostate cDNA library. One of the resulting
 clones, clone #6, yeiled the start codon and and part of the 5' UTR,
 including in-frame upstream stop codons. The nucleotide sequence for MMSC2
 is set forth in SEQ ID NO:2 with the amino acid sequence of the encoded
 protein set forth in SEQ ID NO:3.
 FIG. 1 shows a diagram of MMSC2 indicating the position of ORF and the
 positions of the 13 PDZ domains. FIG. 2 shows a diagram of the key clones
 used to assemble the full length MMSC2 sequence, the probes used to
 identify those clones, and the relative position of the partially
 sequenced mouse ortholog 9BP-1(Accession #AF000168). Within the 5' UTR,
 the in-frame upstream stop codon at nucleotide 42 demonstrates that the
 start codon at nucleotide 57 has been correctly indentified. The 13 PDZ
 domains correspond to the amino acids of MMSC2 as shown in Table 1.
 TABLE 1
 Sequence Correspondence of 13 PDZ Domains
 Domain Number Domain Name Amino Acid Span
 1 P15 136-222
 2 P14 256-335
 3 P13 376-461
 4 P10 555-632
 5 P12 699-785
 6 P11 1007-1090
 7 P9 1150-1241
 8 P8 1316-1398
 9 P7 1439-1529
 10 P1 1595-1677
 11 P2 1691-1772
 12 P3 1828-1913
 13 P4 1953-2037
 Example 2
 Identification of MMSC2-interacting Proteins by Two-hybrid Analysis
 DNA fragments encoding all or portions of MMSC2 are ligated to a two-hybrid
 DNA-binding domain vector such as pGBT.C such that the coding sequence of
 MMSC2 is in-frame with coding sequence for the Gal4p DNA-binding domain.
 These DNA fragments may encode specific PDZ domains of MMSC2 plus the 5 to
 10 amino acids N- and C-terminal of each specific PDZ. A plasmid that
 encodes a DNA-binding domain fusion to a fragment of MMSC2 PDZ is
 introduced into the yeast reporter strain (such as J692) along with a
 library of cDNAs fused to an activation domain. Transformants are spread
 onto 20-150 mm plates of selective media, such as yeast minimal media
 lacking leucine, tryptophan, and histidine, and containing 25 mM
 3-amino-1,2,4-triazole. After one week incubation at 30.degree. C., yeast
 colonies are assayed for expression of the lacZ reporter gene by
 beta-galactosidase filter assay. Colonies that both grow in the absence of
 histidine and are positive for production of beta-galactosidase are chosen
 for further characterization.
 The activation domain plasmid is purified from positive colonies by the
 smash-and-grab technique. These plasmids are introduced into E. coli
 (e.g., DH10B (Gibco BRL)) by electroporation and purified from E. coli by
 the alkaline lysis method. To test for the specificity of the interaction,
 specific activation domain plasmids are cotransformed into strain J692
 with plasmids encoding various DNA-binding domain fusion proteins,
 including fusions to segments of MMSC2 and human lamin C. Transformants
 from these experiments are assayed for expression of the HIS3 and lacZ
 reporter genes. Positives that express reporter genes with MMSC2
 constructs and not with larnin C constructs encode bona fide
 MMSC2-interacting proteins. These proteins are identified and
 characterized by sequence analysis of the insert of the appropriate
 activation domain plasmid.
 Example 3
 Characterization of the Binding Specificity of MMSC2 PDZ Domains by
 Two-Hybrid Analysis
 DNA fragments encoding specific PDZ domains of MMSC2 plus the 5 to 10 amino
 acids N- and C-terminal of each specific PDZ domain are generated by PCR
 amplification. These fragments are ligated to a two-hybrid DNA-binding
 domain vector such as pGBT.C such that the coding sequence of MMSC2 is
 in-frame with coding sequence for the Gal4p DNA-binding domain. An
 activation domain library is prepared that encodes an activation domain
 fused in-frame to random peptide sequences that end with a stop codon. An
 example of this type of library is the Clontech random peptide library. A
 plasmid that encodes a DNA-binding domain fusion to a specific MMSC2 PDZ
 domain is introduced into the yeast reporter strain (such as J692) along
 with a library of random peptides fused to an activation domain.
 Transformants are spread onto 20-150 mm plates of selective media, such as
 yeast minimal media lacking leucine, tryptophan, and histidine, and
 containing 25 mM 3-amino-1,2,4-triazole. After one week incubation at
 30.degree. C., yeast colonies are assayed for expression of the lacZ
 reporter gene by beta-galactosidase filter assay. Colonies that both grow
 in the absence of histidine and are positive for production of
 beta-galactosidase are chosen for sequence analysis. The insert of the
 activation domain construct is characterized by sequence analysis. The
 sequence of the peptide that binds to the MMSC2 PDZ domain is obtained by
 conceptual translation of the nucleotide sequence. Peptide sequences from
 multiple isolates are aligned to determine a consensus binding motif. This
 motif can be used to identify cellular proteins that bind MMSC2 and to
 develop small molecules that inhibit binding to these specific PDZ
 domains.
 Example 4
 In vitro Protein-Protein Interaction Assay
 cDNAs encoding each of the MMSC2 PDZ domains (amino acid residues
 identified in Table 1), and any desired control proteins, were generated
 by PCR and subcloned as glutathione S-transferase (GST) fusions in pGEX
 vectors (Pharmacia). After sequencing to confirm expression construct
 integrity, the resulting clones were expressed in E. coli and the desired
 fusion proteins isolated with glutathione-agarose and recovered with
 glutathione elution. These fusion proteins or control proteins were then
 adsorbed to different wells of a 96-well ELISA plate and remaining sites
 blocked with BSA. Synthetic commercially synthesized peptides encoding the
 desired PDZ-binding domain (i.e., the 16 C-terminal amino acids of MMAC1,
 or the C-terminal peptide sequences of interacting proteins identified by
 the approach of Example or the C-terminal peptide sequences identified by
 the approach of Example 3), or a control peptide, and biotinylated at the
 amino-terminus, were pre-bound to streptavidin-alkaline phosphatase in a
 4:1 molar ratio. The biotinylated peptide streptavidin-alkaline
 phosphatase complexes were then blocked with free biotin. These pre- bound
 peptide streptavidin-alkaline phosphatase complexes were then incubated
 with the immobilized PDZ domains in wash buffer containing PBS, BSA and
 triton-X100. Unbound material was removed with repeated washes. Bound
 peptide/streptavidin-alkaline phosphatase complex was then quantitated by
 a calorimetric phosphatase assay read on a 96-well plate reader. The
 following peptides were used in the initial study:
 SH3 binding peptide: biotin-SGSGILAPPVPPRNTR-COOH (SEQ ID NO:11)
 MMAC1.388-403: biotin-ENEPFDEDQHTQITKV-COOH (SEQ ID NO:12).
 The relative affinity of each of the 13 PDZ domains encoded by MMSC2 for an
 MMAC1 C-terminal PDZ peptide as measured by an ELISA assay is set forth in
 Table 2.
 TABLE 2
 PDZ Binding Assay
 Domain Number Domain Name Peptide A405
 1 P15 MMAC1 0.011
 SH3 0.007
 2 P14 MMAC1 0.001
 SH3 0.006
 3 P13 MMAC1 0.006
 SH3 0.008
 4 P10 MMAC1 0.024
 SH3 0.010
 5 P12 MMAC1 0.000
 SH3 0.005
 6 P11 MMAC1 0.016
 SH3 0.017
 7 P9 MMAC1 0.667
 SH3 0.006
 8 P8 MMAC1 0.006
 SH3 0.008
 9 P7 MMAC1 0.017
 SH3 0.009
 10 P1 MMAC1 0.174
 SH3 0.002
 11 P2 MMAC1 0.012
 SH3 0.009
 12 P3 MMAC1 0.003
 SH3 0.011
 13 P4 MMAC1 0.456
 SH3 0.006
 The GST-affinity pull down assay is a complementary in vitro method for
 investigating protein-protein interactions. PDZ domain-GST fusion proteins
 are incubated with synthetic biotinylated peptides in wash buffer (these
 peptides were described above). Streptavidin magnetic beads are then added
 to recover the biotinylated peptide, then unbound material removed by
 washing. The beads are then incubated with SDS/DTT loading buffer at
 100.degree. C. and bound protein detected by SDS/PAGE and coomasie blue
 staining.
 Example 5
 Mutation Screening of MMSC2
 Nested PCR amplifications were performed on cDNA from tumor cell lines.
 Total cell line RNAs were reverse transcribed with Superscript II (Life
 Technologies) and random hexamers. Using the outer primer pair from each
 amplicon (i.e. 9BP.1A and 9BP.1P or 9BP.2A and 9BP.2P), approximately 10
 ng of CDNA from each cell line was amplified for 26 cycles. Products were
 diluted 60 fold and then reamplified for 22-26 cycles using nested M13
 tailed primers (i.e. 9BP.1B and 9BP.21Q or 9BP.2B and 9BP.2Q). Typical
 primary amplification cycling conditions were an initial denaturation at
 95.degree. C. for 60s, followed by 26 cycles of 96.degree. C. (12s),
 58.degree. C. (15s) and 72.degree. C. (90s). Typical secondary
 amplification cycling conditions were an initial denaturation at
 95.degree. C. for 60s, followed by 22-26 cycles of 96.degree. C. (12s),
 58.degree. C. (15s) and 72.degree. C. (40s). The resulting RT-PCR products
 were sequenced with dye-primer chemistry on ABI 377 sequencers. Sequences
 were examined for the presence of variants using the program Sequencher.
 The primers used are set forth in Table 3. The sequence variants are set
 forth in Table 4.
 TABLE 3
 Table of Primers
 Primer SEQ ID Sequence
 9BP.1A 13 GCCACCGCGGGATTAAGTTTCT
 9BP.1P 14 TGTAGCCAGCAATGGTAATTCCT
 9BP.1B 15 GTTTTCCCAGTCACGACGGTTCCATTTTAATTGCTGTTAAT
 9BP.1Q 16 AGGAAACAGCTATGACCATGGGGATAATAAAAACGATTCATTT
 9BP.1C 17 GTTTTCCCAGTCACGACGTTGAATATGCCCACGTTCCTC
 9BP.1R 18 AGGAAACAGCTATGACCATTCTTTCAATCTTCCATCTCTATG
 9BP.1D 19 GTTTTCCCAGTCACGACGAACAGAGGAGAGCTGGGAATA
 9BP.1S 20 AGGAAACAGCTATGACCATCAAACCAGATCCATCATTCACC
 9BP.1E 21 GTTTTCCCAGTCACGACGGCACAATTTCAGCTCACTCTAA
 9BP.1T 22 AGGAAACAGCTATGACCATGGATGAGGAGAGGGTGATGC
 9BP.2A 23 TCTAGCAGGAATGAGCAGTGAG
 9BP.2P 24 GATCCTGATAATCTAAAATGCTAA
 9BP.2B 25 GTTTTCCCAGTCACGACGAAGTTGATGATTGCAAGAGGTG
 9BP.2Q 26 AGGAAACAGCTATGACCATGGTTTGTGCCATCTACTGCTAT
 9BP.2C 27 GTTTTCCCAGTCACGACGAAAGCAGTGCCGTTGAGCATG
 9BP.2R 28 AGGAAACAGCTATGACCATGCTGACAGTAATGGATACCCT
 9BP.2D 29 GTTTTCCCAGTCACGACGGATTTTTTATCTTCGACGAGAAA
 9BP.2S 30 AGGAAACAGCTATGACCATTTCCCCAAGTAAAGTTATGCCAT
 9BP.2E 31 GTTTTCCCAGTCACGACGTCCTGTTGGACACAGCGGGA
 9BP.2T 32 AGGAAACAGCTATGACCATCATGGCCAAAGGTGCTTGAA
 9BP.3A 33 CCACCCACCACCCAATCAGAAT
 9BP.3P 34 CATCTCGACTAATGGCACCTCC
 9BP.3B 35 GTTTTCCCAGTCACGACGGAGACAGAGGATCCAGTGCT
 9BP.3Q 36 AGGAAACAGCTATGACCATCCCTGACGGTGCTCCCTTCA
 9BP.3C 37 GTTTTCCCAGTCACGACGTTAACTTGGAAAACAGCAGTCT
 9BP.3R 38 AGGAAACAGCTATGACCATCATCACCACAAGAACTGCCATG
 9BP.3D 39 GTTTTCCCAGTCACGACTCTCCTGAAAATGACAGCAT
 9BP.3S 40 AGGAAACAGCTATGACCATTAAATGAGATTCAGTCCACACT
 9BP.3E 41 GTTTTCCCAGTCACGACGATAAATGACTACACACCTGCAA
 9BP.3T 42 AGGAAACAGCTATGACCATAACGATCATCCCCAAGCCATCT
 9BP.4A 43 CTGAGTACCTGCTTGAACAGAG
 9BP.4P 44 GACCATTGATCTCTAGAAGCTC
 9BP.4B 45 GTTTTCCCAGTCACGACGGGACTATTAATATAGCAAAAGGC
 9BP.4Q 46 AGGAAACAGCTATGACCATCAGTGCCATTACTCTTCCAGA
 9BP.4C 47 GTTTTCCCAGTCACGACGTACTTATGTGCCTGCAGAACA
 9BP.4R 48 AGGAAACAGCTATGACCATCATGTTTGATGAAAATGCCCC
 9BP.4D 49 GTTTTCCCAGTCACGACGATTGTTGGTGGACGAGGGATG
 9BP.4S 50 AGGAAACAGCTATGACCATCCATTTCGGCAAAGGCTGAAG
 9BP.4E 51 GTTTTCCCAGTCACGACGCAGAGTCAGAGCCAGAGAAGG
 9BP.4T 52 AGGAAACAGCTATGACCATAGAAGCTCATCTGCAATTTGC
 9BP.5A 53 CAGGCGAGCTGCATATGATTG
 9BP.5P 54 CCTCCTTTGACAATGTCTGACAC
 9BP.5B 55 GTTTTCCCAGTCACGACGGTGTCTTCATAGTGGGGATTGAT
 9BP.5Q 56 AGGAAACAGCTATGACCATGAAGCTCCAGATGTTGCACAT
 9BP.5C 57 GTTTTCCCAGTCACGACGAGAGCCAACTGTTACTACTTC
 9BP.5R 58 AGGAAACAGCTATGACCATTGAAGGAACAGCCTGGGAATC
 9BP.5D 59 GTTTTCCCAGTCACGACGTTAGCCTTCTGAAGACAGCAA
 9BP.5S 60 AGGAAACAGCTATGACCATCATGGATAATAATGGCACCCA
 9BP.5E 61 GTTTTCCCAGTCACGACGTTTCCAAAGGGGCGAACAGGGC
 9BP.5T 62 AGGAAACAGCTATGACCATCCAACAATACTTAATCCTAGGC
 9BP.6A 63 TGGAATTGACTTGAGAAAGGCCA
 9BP.6P 64 CCCCCTACAGTTTTGAAGACCC
 9BP.6B 65 GTTTTCCCAGTCACGACGAAGAGGAGGAAGTGTGTGACAC
 9BP.6Q 66 AGGAAACAGCTATGACCATGACAGGCTGCCTTCACTCACC
 9BP.6C 67 GTTTTCCCAGTCACGACGTCAAAGCTGGTCCATTCCATT
 9BP.6R 68 AGGAAACAGCTATGACCATGGATGTGCCACAGATGGTGAC
 9BP.6D 69 GTTTTCCCAGTCACGACGATGATGCACCCAACTGGAGTT
 9BP.6S 70 AGGAAACAGCTATGACCATGGCTGCCATATCCTCCAACTA
 9BP.6E 71 GTTTTCCCAGTCACGACGGGACCTCCTCAATGTAAGTCT
 9BP.6T 72 AGGAAACAGCTATGACCATATTGTCAGGACCAGTGCATTC
 TABLE 4
 Sequence Variants
 Cell line Type nt variant aa change note
 LNCAP.FGC prostatic G163A val-&gt;ile heterozygous
 OV-1063 ovarian G343T gly-&gt;trp non-het*
 UACC812 breast A1074G thr-&gt;thr heterozygous
 UACC8933 breast G5624A arg-&gt;lys non-het*
 HS776T pancreatic G5624A arg-&gt;lys non-het*
 *In mutation screening from cDNA, a non-heterozygous call for a sequence
 variant is consistent with non-expression of one allele, hemizygosity at
 that position, or homozygosity at that position.
 Example 6
 Generation of Polyclonal Antibody Against MMSC2
 Segments of MMSC2 coding sequence are expressed as fusion protein in E.
 coli. The overexpressed protein is purified by gel elution and used to
 immunize rabbits and mice using a procedure similar to the one described
 by Harlow and Lane, 1988. This procedure has been shown to generate Abs
 against various other proteins (for example, see Kraemer et al., 1993).
 Briefly, a stretch of MMSC2 coding sequence is cloned as a fusion protein
 in plasmid PET5A (Novagen, Inc., Madison, Wis.). After induction with
 IPTG, the overexpression of a fusion protein with the expected molecular
 weight is verified by SDS/PAGE. Fusion protein is purified from the gel by
 electroelution. Identification of the protein as the MMSC2 fusion product
 is verified by protein sequencing at the N-terminus. Next, the purified
 protein is used as immunogen in rabbits. Rabbits are immunized with 100
 .mu.g of the protein in complete Freund's adjuvant and boosted twice in 3
 week intervals, first with 100 .mu.g of immunogen in incomplete Freund's
 adjuvant followed by 100 .mu.g of immunogen in PBS. Antibody containing
 serum is collected two weeks thereafter. This procedure is repeated to
 generate antibodies against the mutant forms of the MMSC2 gene product.
 These antibodies, in conjunction with antibodies to wild type MMSC2, are
 used to detect the presence and the relative level of the mutant forms in
 various tissues and biological fluids.
 Example 7
 Generation of Polyclonal Antibody Against MMSC2-MMSC2 Interacting Protein
 Complex
 MMSC2 is capable of binding to certain proteins, e.g., MMAC1. A complex of
 the two proteins is prepared, e.g., by mixing purified preparations of
 each of the two proteins. If desired, the protein complex can be
 stabilized by cross-linking the proteins in the complex by methods known
 to those of skill in the art. The protein complex is used to immunize
 rabbits and mice using a procedure similar to the one described by Harlow
 and Lane, 1988. This procedure has been shown to generate Abs against
 various other proteins (for example, see Kraemer et al., 1993).
 Briefly, the purified protein complex is used as immunogen in rabbits.
 Rabbits are immunized with 100 pg of the protein in complete Freund's
 adjuvant and boosted twice in 3 week intervals, first with 100 .mu.g of
 immunogen in incomplete Freund's adjuvant followed by 100 .mu.g of
 immunogen in PBS. Antibody containing serum is collected two weeks
 thereafter.
 This procedure is repeated to generate antibodies against forms of the
 complex which comprise mutant MMSC2 or mutant MMSC2 interacting protein
 (e.g)., mutant MMAC1). These antibodies, in conjunction with antibodies to
 wild type MMSC2 or MMSC2 interacting protein (e.g., MMAC1), are used to
 detect the presence and the relative level of the mutant forms in various
 tissues and biological fluids.
 Example 8
 Generation of Monoclonal Antibodies Specific for MMSC2
 Monoclonal antibodies are generated according to the following protocol.
 Mice are immunized with immunogen comprising intact MMSC2 or MMSC2
 peptides (wild type or mutant) conjugated to keyhole limpet hemocyanin
 using glutaraldehyde or EDC as is well known.
 The immunogen is mixed with an adjuvant. Each mouse receives four
 injections of 10 to 100 .mu.g of immunogen and after the fourth injection
 blood samples are taken from the mice to determine if the serum contains
 antibody to the immunogen. Serum titer is determined by ELISA or RIA. Mice
 with sera indicating the presence of antibody to the immunogen are
 selected for hybridoma production.
 Spleens are removed from immune mice and a single cell suspension is
 prepared (see Harlow and Lanc, 1988). Cell fusions are performed
 essentially as described by Kohler and Milstein, 1975. Briefly, P3.65.3
 myeloma cells (American Type Culture Collection, Rockville, Md.) are fused
 with immune spleen cells using polyethylene glycol as described by Harlow
 and Lane, 1988. Cells are plated at a density of 2.times.10.sup.5
 cells/well in 96 well tissue culture plates. Individual wells are examined
 for growth and the supernatants of wells with growth are tested for the
 presence of MMSC2 specific antibodies by ELISA or RIA using wild type or
 mutant MMSC2 target protein. Cells in positive wells are expanded and
 subcloned to establish and confirm monoclonality.
 Clones with the desired specificities are expanded and grown as ascites in
 mice or in a hollow fiber system to produce sufficient quantities of
 antibody for characterization and assay development.
 Example 9
 Generation of Monoclonal Antibodies Specific for MMSC2-MMSC2 Interacting
 Protein Complex
 Monoclonal antibodies are generated according to the following protocol.
 Mice are immunized with immunogen comprising MMSC2-MMSC2 interacting
 protein complexes (wild type or mutant), such as MMAC1, conjugated to
 keyhole limpet hemocyanin using glutaraldehyde or EDC as is well known.
 The complexes may be stabilized by cross-linking.
 The immunogen is mixed with an adjuvant. Each mouse receives four
 injections of 10 to 100 .mu.g of immunogen and after the fourth injection
 blood samples are taken from the mice to determine if the serum contains
 antibody to the immunogen. Serum titer is determined by ELISA or RIA. Mice
 with sera indicating the presence of antibody to the immunogen are
 selected for hybridoma production.
 Spleens are removed from immune mice and a single cell suspension is
 prepared (see Harlow and Lane, 1988). Cell fusions are performed
 essentially as described by Kohler and Milstein, 1975. Briefly, P3.65.3
 myeloma cells (American Type Culture Collection, Rockville, Md.) are fused
 with immune spleen cells using polyethylene glycol as described by Harlow
 and Lane, 1988. Cells are plated at a density of 2.times.10.sup.5
 cells/well in 96 well tissue culture plates. Individual wells are examined
 for growth and the supernatants of wells with growth are tested for the
 presence of MMSC2-MMSC2 interacting protein complex specific antibodies by
 ELISA or RIA using wild type or mutant MMSC2-MMSC2 interacting protein
 complexes as target protein. Cells in positive wells are expanded and
 subcloned to establish and confirm monoclonality.
 Clones with the desired specificities are expanded and grown as ascites in
 mice or in a hollow fiber system to produce sufficient quantities of
 antibody for characterization and assay development. Antibodies are tested
 for binding to MMSC2 alone or to MMSC2 interacting protein alone to
 determine which are specific for the complex as opposed to binding to the
 individual proteins.
 Example 10
 Sandwich Assay for MMSC2
 Monoclonal antibody is attached to a solid surface such as a plate, tube,
 bead or particle. Preferably, the antibody is attached to the well surface
 of a 96-well ELISA plate. 100 .mu.L sample (e.g., serum, urine, tissue
 cytosol) containing the MMSC2 peptide/protein (wild-type or mutants) is
 added to the solid phase antibody. The sample is incubated for 2 hrs at
 room temperature. Next the sample fluid is decanted, and the solid phase
 is washed with buffer to remove unbound material. 100 .mu.L of a second
 monoclonal antibody (to a different determinant on the MMSC2
 peptide/protein) is added to the solid phase. This antibody is labeled
 with a detector molecule (e.g., .sup.125 I, enzyme, fluorophore, or a
 chromophore) and the solid phase with the second antibody is incubated for
 two hrs at room temperature. The second antibody is decanted and the solid
 phase is washed with buffer to remove unbound material.
 The amount of bound label, which is proportional to the amount of MMSC2
 peptide/protein present in the sample, is quantified. Separate assays are
 performed using monoclonal antibodies which are specific for the wild-type
 MMSC2 as well as monoclonal antibodies specific for each of the mutations
 identified in MMSC2.
 Example 11
 Sandwich Assay for MMAC1 Using MMSC2
 MMSC2 or PDZ domain 7 of MMSC2 is attached to a solid surface such as a
 plate, tube, bead or particle. Preferably, MMSC2 or its PDZ domain is
 attached to the well surface of a 96-well ELISA plate. 100 .mu.L sample
 (e.g., serum, urine, tissue cytosol) containing the MMAC1 peptide/protein
 (wild-type or mutants) is added to the solid phase MMSC2. The sample is
 incubated for 2 hrs at room temperature. Next the sample fluid is
 decanted, and the solid phase is washed with buffer to remove unbound
 material. 100 .mu.L of a monoclonal antibody to MMAC1 is added to the
 solid phase. The antibody is labeled with a detector molecule (e.g.,
 .sup.125 I, enzyme, fluorophore, or a chromophore) and the solid phase
 with the antibody is incubated for two hrs at room temperature. The
 antibody is decanted and the solid phase is washed with buffer to remove
 unbound material. The amount of bound label, which is proportional to the
 amount of wild-type MMAC1 present in the sample, is quantified.
 Example 12
 Drug Screening
 The invention is useful in screening for drugs which can overcome mutations
 in MMSC2 and also mutations in MMAC1. The knowledge that MMSC2 and MMAC1
 form a complex is useful in designing such assays. If a mutation is
 present in either MMSC2 or in MMAC1 which prevents the MMSC2-MMAC1 complex
 from forming, drugs may be screened which will overcome the mutation and
 allow the protein complex to form and to be active. Such screening assays
 can be, e.g., a yeast two hybrid assay which is dependent upon two
 proteins interacting. In such an assay, the presence of a mutant protein
 may show no activity or low activity in such an assay, while the presence
 of a useful drug will result in formation of a proper complex which
 results in activity in the assay.
 A simple binding assay which shows the binding, i.e., formation of a
 complex, can similarly be used as outlined above. Useful drugs will
 increase the formation of MMSC2-MMAC1 complexes. Antibodies may also be
 used to monitor the amount of complex present. Antibodies specific for the
 complex are especially useful. If the presence of a drug increases the
 amount of complex present then the drug is a good candidate for treating
 the cancer which is a result of the mutation in either the MMSC2 or the
 MMAC1.
 While the invention has been disclosed in this patent application by
 reference to the details of preferred embodiments of the invention, it is
 to be understood that the disclosure is intended in an illustrative rather
 than in a limiting sense, as it is contemplated that modifications will
 readily occur to those skilled in the art, within the spirit of the
 invention and the scope of the appended claims.
 LIST OF REFERENCES
 Anand, R. (1992). Techniques for the Analysis of Complex Genomes, (Academic
 Press).
 Anderson, et al. (1980). Proc. Natl. Acad. Sci. USA 77:5399-5403.
 Ausubel, F. M., et al. (1992). Current Protocols in Molecular Biology, (J.
 Wiley and Sons, N.Y.)
 Bartel, P. L. and Fields, S. (1995). Methods in Enzymology 254:241-263.
 Bartel, P. L. et al. (1993). "Using the 2-hybrid system to detect
 protein-protein interactions." In Cellular Interactions in Development: A
 Practical Approach, Oxford University Press, pp. 153-179.
 Beaucage and Carruthers (1981). Tetra. Letts. 22:1859-1862.
 Berglund, P. et al. (1993) Biotechnology 11:916-920
 Berkner, et al. (1988). BioTechniques 6:616-629.
 Berkner (1992). Curr. Top. Microbiol. Immunol. 158:39-61.
 Borman, S. (1996). Chemical & Engineering News, December 9 issue, pp.
 42-43.
 Brandyopadhyay and Temin (1984). Mol. Cell Biol. 4:749-754.
 Breakfield and Geller (1987). Mol. Neurobiol. 1:337-371.
 Breeden, L., and Naysmyth, K. (1985). Cold Spring Harbor Symp. Quant. Biol.
 50:643-650.
 Brenman, J. E. et al. (1996). Cell 84:757-767.
 Brinster, et al. (1981). Cell 27,:223-231.
 Buchschacher and Panganiban (1992). J. Virol. 66:2731-2739.
 Capecchi, M. R. (1989). Science 244:1288.
 Cariello (1988). Human Genetics 42:726.
 Chee, M., et al. (1996). Science 274:610-614.
 Chevray, P. M. & Nathans, D. N. (1992). Proc. Natl. Acad. Sci. USA
 89:5789-5793.
 Cho, K. O. et al. (1992). Neuron 9:929-942.
 Compton, J. (1991). "Nucleic acid sequence-based amplification." Nature
 350:91-92.
 Conner, B. J., et al. (1983). Proc. Natl. Acad. Sci. USA 80:278-282.
 Constantini and Lacy (1981). Nature 294:92-94.
 Cotten, M., et al. (1990). Proc. Natl. Acad. Sci. USA 87:4033-4037.
 Cotton, R. G., et al. (1988). Proc. Natl. Acad Sci. USA 85:4397-4401.
 Culver, K. (1996). Gene Therapy: A Primer for Physicians, 2nd Ed., Mary Ann
 Liebert.
 Curiel, et al. (1991a). Hum. Gene Ther. 3:147-154.
 Curiel, et al. (1991 b). Proc. Natl. Acad. Sci. USA 88:8850-8854.
 Deutscher, M. (1990). Meth. Enzymology 182 (Academic Press, San Diego,
 Calif.).
 Donehower, L. A., et al. (1992). Nature 356:215.
 Editorial (1996). Nature Genetics 14:367-370.
 Elghanian, R., et al. (1997). Science 277:1078-1081.
 Enhancers and Eukaryotic Gene Expression, Cold Spring Harbor Press, Cold
 Spring Harbor, N.Y. (1983).
 Erickson, J., et al. (1990). Science 249:527-533.
 Fahy, E., et al. (1991). PCR Methods Appl. 1:25-33.
 Feigner, et al. (1987). Proc. Natl. Acad. Sci. USA 84:7413-7417.
 Fields, S. & Song, O.-K. (1989). Nature 340:245-246.
 Fiers, et al. (1978). Nature 273:113.
 Fink, et al. (1992). Hum. Gene Ther. 3:11-19.
 Fink, D. J. et al. (1996). Ann. Rev. Neurosci. 19:265-287.
 Finkelstein, J., et al. (1990). Genomics 7:167-172.
 Fodor, S. P. A. (1997). DNA Sequencing. Massively Parallel Genomics.
 Science 277:393-395.
 Freese, et al. (1990). Biochem. Pharmacol. 40:2189-2199.
 Friedman, T. (1991). In Therapy for Genetic Diseases, T. Friedman, ed.,
 Oxford University Press, pp. 105-121.
 Furnari, F. B. et al. (1997). Proc. Natl. Acad. Sci. USA 94:12479-12484.
 Gietz, R. D., et al. (1995). Yeast 11:355-360.
 Glover, D. (1985). DNA Cloning, I and II (Oxford Press).
 Goding (1986). Monoclonal Antibodies: Principles and Practice, 2d ed.
 (Academic Press, N.Y.).
 Godowski, et al. (1988). Science 241:812-816.
 Gordon, et al. (1980). Proc. Natl. Acad. Sci. USA 77:7380-7384.
 Gorziglia and Kapikian (1992). J. Virol. 66:4407-4412.
 Graham and van der Eb (1973). Virology 52:456-467.
 Grompe, M. (1993). Nature Genetics 5:111-117.
 Grompe, M., et al. (1989). Proc. Natl. Acad. Sci. USA 86:5855-5892.
 Guthrie, G. and Fink, G. R. (1991). Guide to Yeast Genetics and Molecular
 Biology (Academic Press).
 Hacia, J. G., et al. (1996). Nature Genetics 14:441-447.
 Harlow and Lane (1988). Antibodies: A Laboratory Manual (Cold Spring Harbor
 Laboratory, Cold Spring Harbor, N.Y.).
 Harrison, S. C. (1996). Cell 86:341-343.
 Hasty, P., K., et al. (1991). Nature 350:243.
 Helseth, et al. (1990). J. Virol. 64:2416-2420.
 Hodgson, J. (1991). Bio/Technology 9:19-21.
 Huse, et al. (1989). Science 246:1275-1281.
 Innis, et al. (1990). PCR Protocols: A Guide to Methods and Applications
 (Academic Press, San Diego, Calif.).
 Jablonski, E., et al. (1986). Nucl. Acids Res. 14:6115-6128.
 Jakoby, W. B. and Pastan, I. H. (eds.) (1979). Cell Culture. Methods in
 Enzymology, volume 58 (Academic Press, Inc., Harcourt Brace Jovanovich
 (New York)).
 Johnson, et al. (1992). J. Virol 66:2952-2965.
 Johnson, et al. (1993). "Peptide Turn Mimetics" in Biotechnology and
 Pharmacy, Pezzuto et al., eds., Chapman and Hall, New York.
 Kaneda, et al. (1989). J. Biol. Chem. 264:12126-12129.
 Kanehisa (1984). Nucl. Acids Res. 12:203-213.
 Kavanaugh, W. M. et al. (1995). Science 268:1177-1179.
 Kennedy, M. B. (1995). Trends Biochem. Sci. 20:350.
 Kinszler, K. W., et al. (1991). Science 251:1366-1370.
 Kohler, G. and Milstein, C. (1975). Nature 256:495-497.
 Kong, D. et al. (1997). Nature Genetics 17:143-144.
 Kornau, H. C. et al. (1995). Science 269:1737-1740.
 Kraemer, F. B. et al. (1993). J. Lipid Res. 34:663-672.
 Kubo, T., et al. (1988). FEBS Letts. 241:119.
 Kyte, J. & Dolittle, R. F. (1982). J. Mol. Bio. 157:105-132.
 Landegren, et al. (1988). Science 242:229.
 Lee, J. E. et al. (1995). Science 268:836-844.
 Lemmon, M. A. et al. (1996). Cell 85:621-624.
 Li, D. M. & Sun, H. (1997). Cancer Res. 57:2124-2129.
 Li, J. et al. (1997). Science 275:1943-1947.
 Liaw, D. et al. (1997). Nature Genetics 16:64-67.
 Lim, et al. (1992). Circulation 83:2007-2011.
 Lipshutz, R. J., et al. (1995). Biotechniques 19:442-447.
 Lockhart, D. J., et al. (1996). Nature Biotechnology 14:1675-1680.
 Louis, D. N. & Gusella, J. F. (1995). Trends. Genet. 11:412-415.
 Madzak, et al. (1 992). J. Gen. Virol. 73:1533-1536.
 Maldonado, E., et al. (1996). Nature 381:86-89.
 Maniatis, T. et al. (1982). Molecular Cloning: A Laboratory Manual (Cold
 Spring Harbor Laboratory, Cold Spring Harbor, N.Y.).
 Mann and Baltimore (1985). J. Virol. 54:401-407.
 Margolskee (1992). Curr. Top. Aicrobiol. Immunol. 158:67-90.
 Marsh, D. J. et al. (1997). Nature Genetics 16:333-334.
 Martin, R., et al. (1990). BioTechniques 9:762-768.
 Matteucci, M. D. and Caruthers, M. H. (1981). J. Am. Chem. Soc. 103:3185.
 Matthews and Kricka (1988). Anal. Biochem. 169:1.
 Merrifield (1963). J. Am. Chem. Soc. 85:2149-2156.
 Metzger, et al. (1988). Nature 334:31-36.
 Miller (1992). Curr. Top. Microbiol. Immunol. 158:1-24.
 Miller, et al. (1985). Mol. Cell. Biol. 5:431 -437.
 Miller, et al. (1988). J. Virol. 62:4337-4345.
 Modrich, P. (1991). Ann. Rev. Genet. 25:229-253.
 Mombaerts, P., et al. (1992). Cell 68:869.
 Moss (1992). Curr. Top. Microbiol. Immunol. 158:25-38.
 Moss, B. (1996). Proc. Natl. Acad. Sci. USA 93:11341-11348.
 Muzyczka (1992). Curr. Top. Microbiol. Immunol. 158:97-123.
 Nabel, et al. (1990). Science 249:1285-1288.
 Nabel (1992). Hum. Gene Ther. 3:399-410.
 Naldini, L. et al. (1996). Science 272:263-267.
 Nelen, M. R. et al. (1997). Hum. Mol. Genet. 6:1383-1387.
 Newton, C. R., et al. (1989). Nucl. Acids Res. 17:2503-2516.
 Nguyen, Q., et al. (1992). BioTechniques 13:116-123.
 Novack, et al. (1986). Proc. Natl. Acad. Sci. USA 83:586.
 Ohi, et al. (1990). Gene 89:279-282.
 Olschwang, S. et al. (1998). Nature Genetics 18:12-13.
 Orita, M., et al. (1989). Proc. Natl. Acad. Sci. USA 86:2766-2770.
 Page, et al. (1990). J. Virol. 64:5370-5276.
 Pawson, T. (1994). Adv. Cancer Res. 64:87-110.
 Pawson, T. & Scott, J. D. (1997). Science 278:2075-2080.
 Pellicer, et al. (1980). Science 209:1414-1422.
 Petropoulos, et al. (1992). J. Virol. 66:3391-3397.
 Philpott, K. L., et al. (1992). Science 256:1448.
 Quantin, et al. (1992). Proc. Natl. Acad. Sci. USA 89:2581-2584.
 Rano and Kidd (1989). Nucl Acids Res. 17:8392.
 Remington's Pharmaceutical Sciences, 18th Ed. (1990, Mack Publishing Co.,
 Easton, Pa.).
 Rigby, P. W. J., et al. (1977). J. Mol. Biol. 113:237-251.
 Rosenfeld, et al. (1992). Cell 68:143-155.
 Russell, D. & Hirata, R. (1998). Nature Genetics 18:323-328.
 Sambrook, J., et al. (1989). Molecular Cloning: A Laboratory Manual, 2nd
 Ed. (Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.).
 Sande, S., and Privalsky, M. L. (1996). Molecular Endocrin. 10:813-825.
 Scharf(1986). Science 233:1076.
 Schiessinger, J. (1994). "SH2/SH3 signaling proteins." Curr. Opin. Genet.
 Dev. 4:25-30.
 Schneider, G. et al. (1998). Nature Genetics 18:180-183.
 Scopes, R. (1982). Protein Purification: Principles and Practice,
 (Springer-Verlag, N.Y.).
 Shaw, G. (1996). Bioessays 18:35-46.
 Sheffield, V. C., et al. (1989). Proc. Natl. Acad. Sci. USA 86:232-236.
 Sheffield, V. C., et al., (1991). Am. J. Hum. Genet. 49:699-706.
 Shenk, T. E., et al. (1975). Proc. Natl. Acad. Sci. USA 72:989-993.
 Shieh, B. & Zhu, M. (1996). Neuron 16:991-998.
 Shimada, et al. (1991). J. Clin. Invest. 88:1043-1047.
 Shinkai, Y., et al. (1992). Cell 68:855.
 Shoemaker, D. D., et al. (1996). Nature Genetics 14:450-456.
 Snouwaert, J. N., et al. (1992). Science 257:1083.
 Songyang, Z. et al. (1997). Science 275:73-77.
 Sorge, et al. (1984). Mol. Cell. Biol. 4:1730-1737.
 Spargo, C. A., et al. (1996). Mol. Cell. Probes 10:247-256.
 Steck, P. A. et al. (1997). Nature Genetics 15:356-362.
 Stewart, et al. (1992). Hum. Gene Ther. 3:267-275.
 Stratford-Perricaudet, et al. (1990). Hum. Gene Ther. 1:241-256.
 Tsunoda, S. et al. (1997). Nature 388:243-249.
 Valancius, V. and Smithies, O. (1991). Mol. Cell. Biol. 11:1402.
 van der Greer, P. & Pawson, T. (1995). Trends Biochem. Sci. 20:277-280.
 Wagner, et al. (1991). Proc. Natl. Acad. Sci. USA 88:4255-4259.
 Wagner, et al. (1990). Proc. Natl. Acad. Sci. USA 87:3410-3414.
 Walker, G. T., et al. (1992). Nucl. Acids Res. 20:1691-1696.
 Wang and Huang (1989). Biochemistry 28:9508-9514.
 Wartell, R. M., et al. (1990). Nucl. Acids Res. 18:2699-2705.
 Wells, J. A. (1991). Methods in Enzymol. 202:390-411.
 Wetmur and Davidson (1968). J. Mol. Biol. 31:349-370.
 White, M. B., et al. (1992). Genomics 12:301-306.
 White and Lalouel (1988). Ann. Rev. Genet. 22:259-279.
 Wilkinson, et al. (1992). Nucleic Acids Res. 20:2233-2239.
 Wolff, et al. (1990). Science 247:1465-1468.
 Wolff, et al. (1991). BioTechniques 11:474-485.
 Woods, D. F. & Bryant, P. J. (1991). Cell 66:451-464.
 Woods, D. F. & Bryant, P. J. (1993 ). Mech. Dev. 44:5-89.
 Wu, et al. (1989a). Genomics 4:560-569.
 Wu, et al. (1989 b). J. Biol. Chem. 264:16985-16987.
 Wu, et al. (1991). J. Biol. Chem. 266:14338-14342.
 Zenke, et al. (1990). Proc. Natl. Acad. Sci. USA 87:3655-3659.
 PATENTS AND PATENT APPLICATIONS
 Geysen, H., published PCT application WO 84/03564, published Sep. 13, 1984
 Hitzeman et al., EP 73,675A.
 EP 225,807A.
 EP 332,435A.
 EP425,731 A
 U.S. Pat. No. 3,817,837
 U.S. Pat. No. 3,850,752
 U.S. Pat. No. 3,939,350
 U.S. Pat. No. 3,996,345
 U.S. Pat. No. 4,275,149
 U.S. Pat. No. 4,277,437
 U.S. Pat. No. 4,366,241
 U.S. Pat. No. 4,376,110.
 U.S. Pat. No. 4,486,530.
 U.S. Pat. No. 4,554,101
 U.S. Pat. No. 4,683,195
 U.S. Pat. No. 4,683,202
 U.S. Pat. No. 4,816,567
 U.S. Pat. No. 4,868,105.
 U.S. Pat. No. 5,252,479.
 U.S. Pat. No. 5,270,184.
 U.S. Pat. No. 5,409,818.
 U.S. Pat. No. 5,455,166.
 U.S. Pat. No. 5,550,050
 U.S. Pat. No. 5,691,198
 WO 90/07936
 WO 92/19195
 WO 93/07282.
 WO 94/25503
 WO 95/01203
 WO 95/05452
 WO 96/02286
 WO 96/02646
 WO 96/40871
 WO 96/40959
 WO 97/02048
 WO 97/12635
 SEQUENCE LISTING
 &lt;100&gt; GENERAL INFORMATION:
 &lt;160&gt; NUMBER OF SEQ ID NOS: 72
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 1
 &lt;211&gt; LENGTH: 4
 &lt;212&gt; TYPE: PRT
 &lt;213&gt; ORGANISM: Artificial Sequence
 &lt;220&gt; FEATURE:
 &lt;223&gt; OTHER INFORMATION: Description of Artificial SequenceConsensus
 motif for interaction of PDZ domains.
 &lt;400&gt; SEQUENCE: 1
 Glu Xaa Xaa Xaa
 1
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 2
 &lt;211&gt; LENGTH: 7431
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;220&gt; FEATURE:
 &lt;221&gt; NAME/KEY: CDS
 &lt;222&gt; LOCATION: (57)..(6167)
 &lt;400&gt; SEQUENCE: 2
 gttccatttt aattgctgtt aatcatttca gagaagaaca ctgaactttg aaaaaa atg 59
 Met
 1
 ttg gaa gcc att gac aaa aat cgg gcc ctg cat gca gca gag cgc ttg 107
 Leu Glu Ala Ile Asp Lys Asn Arg Ala Leu His Ala Ala Glu Arg Leu
 5 10 15
 caa acc aag ctg cga gaa cgt ggg gat gta gca aat gaa gac aaa ctg 155
 Gln Thr Lys Leu Arg Glu Arg Gly Asp Val Ala Asn Glu Asp Lys Leu
 20 25 30
 agc ctt ctg aag tca gtc ctg cag agc cct ctc ttc agt cag att ctg 203
 Ser Leu Leu Lys Ser Val Leu Gln Ser Pro Leu Phe Ser Gln Ile Leu
 35 40 45
 agc ctt cag act tct gta cag cag ctg aaa gac cag gta aat att gca 251
 Ser Leu Gln Thr Ser Val Gln Gln Leu Lys Asp Gln Val Asn Ile Ala
 50 55 60 65
 act tca gca act tca aat att gaa tat gcc cac gtt cct cat ctc agc 299
 Thr Ser Ala Thr Ser Asn Ile Glu Tyr Ala His Val Pro His Leu Ser
 70 75 80
 cca gct gtg att cct act ctg caa aat gaa tcg ttt tta tta tcc cca 347
 Pro Ala Val Ile Pro Thr Leu Gln Asn Glu Ser Phe Leu Leu Ser Pro
 85 90 95
 aac aat ggg aat ctg gaa gca ctt aca gga cct ggt att cca cac att 395
 Asn Asn Gly Asn Leu Glu Ala Leu Thr Gly Pro Gly Ile Pro His Ile
 100 105 110
 aat ggg aaa cct gct tgt gat gaa ttt gat cag ctt atc aaa aat atg 443
 Asn Gly Lys Pro Ala Cys Asp Glu Phe Asp Gln Leu Ile Lys Asn Met
 115 120 125
 gcc cag ggt cgc cat gta gaa gtt ttt gag ctc ctc aaa cct cca tct 491
 Ala Gln Gly Arg His Val Glu Val Phe Glu Leu Leu Lys Pro Pro Ser
 130 135 140 145
 gga ggc ctt ggg ttt agt gtt gtg gga cta aga agt gaa aac aga gga 539
 Gly Gly Leu Gly Phe Ser Val Val Gly Leu Arg Ser Glu Asn Arg Gly
 150 155 160
 gag ctg gga ata ttt gtt caa gag ata caa gag ggc agt gtg gcc cat 587
 Glu Leu Gly Ile Phe Val Gln Glu Ile Gln Glu Gly Ser Val Ala His
 165 170 175
 aga gat gga aga ttg aaa gaa act gat caa att ctt gct atc aat gga 635
 Arg Asp Gly Arg Leu Lys Glu Thr Asp Gln Ile Leu Ala Ile Asn Gly
 180 185 190
 cag gct ctt gat cag aca att aca cat cag cag gct atc agc atc ctg 683
 Gln Ala Leu Asp Gln Thr Ile Thr His Gln Gln Ala Ile Ser Ile Leu
 195 200 205
 cag aaa gcc aaa gat act gtc cag cta gtt att gcc aga ggc tca ttg 731
 Gln Lys Ala Lys Asp Thr Val Gln Leu Val Ile Ala Arg Gly Ser Leu
 210 215 220 225
 cct cag ctt gtc agc ccc ata gtt tcc cgt tct cca tct gca gcc agc 779
 Pro Gln Leu Val Ser Pro Ile Val Ser Arg Ser Pro Ser Ala Ala Ser
 230 235 240
 aca att tca gct cac tct aat ccg gtt cac tgg caa cac atg gaa acg 827
 Thr Ile Ser Ala His Ser Asn Pro Val His Trp Gln His Met Glu Thr
 245 250 255
 att gaa ttg gtg aat gat gga tct ggt ttg gga ttt ggc atc ata gga 875
 Ile Glu Leu Val Asn Asp Gly Ser Gly Leu Gly Phe Gly Ile Ile Gly
 260 265 270
 gga aaa gca act ggt gtg ata gta aaa acc att ctg cct gga gga gta 923
 Gly Lys Ala Thr Gly Val Ile Val Lys Thr Ile Leu Pro Gly Gly Val
 275 280 285
 gct gat cag cat ggg cgt tta tgc agt gga gac cac att cta aag att 971
 Ala Asp Gln His Gly Arg Leu Cys Ser Gly Asp His Ile Leu Lys Ile
 290 295 300 305
 ggt gac aca gat cta gca gga atg agc agt gag caa gta gca caa gtc 1019
 Gly Asp Thr Asp Leu Ala Gly Met Ser Ser Glu Gln Val Ala Gln Val
 310 315 320
 ctt agg caa tgt gga aat aga gtt aag ttg atg att gca aga ggt gcc 1067
 Leu Arg Gln Cys Gly Asn Arg Val Lys Leu Met Ile Ala Arg Gly Ala
 325 330 335
 ata gaa gaa cgt aca gca ccc act gct ttg ggc atc acc ctc tcc tca 1115
 Ile Glu Glu Arg Thr Ala Pro Thr Ala Leu Gly Ile Thr Leu Ser Ser
 340 345 350
 tcc cca act tca aca cca gag ttg cgg gtt gat gct tct act cag aaa 1163
 Ser Pro Thr Ser Thr Pro Glu Leu Arg Val Asp Ala Ser Thr Gln Lys
 355 360 365
 ggt gaa gaa agt gag aca ttt gat gta gaa ctc act aaa aat gtc caa 1211
 Gly Glu Glu Ser Glu Thr Phe Asp Val Glu Leu Thr Lys Asn Val Gln
 370 375 380 385
 gga tta gga att acc att gct ggc tac att gga gat aaa aaa ttg gaa 1259
 Gly Leu Gly Ile Thr Ile Ala Gly Tyr Ile Gly Asp Lys Lys Leu Glu
 390 395 400
 cct tca gga atc ttt gta aag agc att aca aaa agc agt gcc gtt gag 1307
 Pro Ser Gly Ile Phe Val Lys Ser Ile Thr Lys Ser Ser Ala Val Glu
 405 410 415
 cat gat gga aga atc caa att gga gac caa att ata gca gta gat ggc 1355
 His Asp Gly Arg Ile Gln Ile Gly Asp Gln Ile Ile Ala Val Asp Gly
 420 425 430
 aca aac ctt cag ggt ttt act aat cag caa gca gta gag gta ttg cga 1403
 Thr Asn Leu Gln Gly Phe Thr Asn Gln Gln Ala Val Glu Val Leu Arg
 435 440 445
 cat aca gga caa act gtg ctc ctg aca cta atg agg aga gga atg aag 1451
 His Thr Gly Gln Thr Val Leu Leu Thr Leu Met Arg Arg Gly Met Lys
 450 455 460 465
 cag gaa gcc gag ctc atg tca agg gaa gac gtc aca aaa gat gca gat 1499
 Gln Glu Ala Glu Leu Met Ser Arg Glu Asp Val Thr Lys Asp Ala Asp
 470 475 480
 ttg tct cct gtt aat gcc agc ata atc aaa gaa aat tat gaa aaa gat 1547
 Leu Ser Pro Val Asn Ala Ser Ile Ile Lys Glu Asn Tyr Glu Lys Asp
 485 490 495
 gaa gat ttt tta tct tcg acg aga aac acc aac ata tta cca act gaa 1595
 Glu Asp Phe Leu Ser Ser Thr Arg Asn Thr Asn Ile Leu Pro Thr Glu
 500 505 510
 gaa gaa ggg tat cca tta ctg tca gct gag ata gaa gaa ata gaa gat 1643
 Glu Glu Gly Tyr Pro Leu Leu Ser Ala Glu Ile Glu Glu Ile Glu Asp
 515 520 525
 gca caa aaa caa gaa gct gct ctg ctg aca aaa tgg caa agg att atg 1691
 Ala Gln Lys Gln Glu Ala Ala Leu Leu Thr Lys Trp Gln Arg Ile Met
 530 535 540 545
 gga att aac tat gaa ata gtg gtg gcc cat gtg agc aag ttt agt gag 1739
 Gly Ile Asn Tyr Glu Ile Val Val Ala His Val Ser Lys Phe Ser Glu
 550 555 560
 aac agt gga ttg ggg ata agc ctg gaa gcg aca gtg gga cat cat ttt 1787
 Asn Ser Gly Leu Gly Ile Ser Leu Glu Ala Thr Val Gly His His Phe
 565 570 575
 atc cga tct gtt cta cca gag ggt cct gtt gga cac agc ggg aag ctc 1835
 Ile Arg Ser Val Leu Pro Glu Gly Pro Val Gly His Ser Gly Lys Leu
 580 585 590
 ttc agt gga gac gag cta ttg gaa gta aat ggc ata act tta ctt ggg 1883
 Phe Ser Gly Asp Glu Leu Leu Glu Val Asn Gly Ile Thr Leu Leu Gly
 595 600 605
 gaa aat cac caa gat gtg gtg aat atc tta aaa gaa ctg cct ata gaa 1931
 Glu Asn His Gln Asp Val Val Asn Ile Leu Lys Glu Leu Pro Ile Glu
 610 615 620 625
 gtg aca atg gtg tgc tgt cgt cga act gtg cca ccc acc acc caa tca 1979
 Val Thr Met Val Cys Cys Arg Arg Thr Val Pro Pro Thr Thr Gln Ser
 630 635 640
 gaa ttg gat agc ctg gac tta tgt gat att gag cta aca gaa aag cct 2027
 Glu Leu Asp Ser Leu Asp Leu Cys Asp Ile Glu Leu Thr Glu Lys Pro
 645 650 655
 cac gta gat cta ggt gag ttc atc ggg tca tca gag aca gag gat cca 2075
 His Val Asp Leu Gly Glu Phe Ile Gly Ser Ser Glu Thr Glu Asp Pro
 660 665 670
 gtg ctg gcg atg act gat gcg ggt cag agt aca gaa gag gtt caa gca 2123
 Val Leu Ala Met Thr Asp Ala Gly Gln Ser Thr Glu Glu Val Gln Ala
 675 680 685
 cct ttg gcc atg tgg gag gct ggc att cag cac ata gag ctg gag aaa 2171
 Pro Leu Ala Met Trp Glu Ala Gly Ile Gln His Ile Glu Leu Glu Lys
 690 695 700 705
 ggg agc aaa gga ctt ggt ttt agc att tta gat tat cag gat cca att 2219
 Gly Ser Lys Gly Leu Gly Phe Ser Ile Leu Asp Tyr Gln Asp Pro Ile
 710 715 720
 gat cca gca agc act gtg att ata att cgt tct ttg gtg cct ggc ggc 2267
 Asp Pro Ala Ser Thr Val Ile Ile Ile Arg Ser Leu Val Pro Gly Gly
 725 730 735
 att gct gaa aag gat gga cga ctt ctt cct ggt gac cga ctc atg ttt 2315
 Ile Ala Glu Lys Asp Gly Arg Leu Leu Pro Gly Asp Arg Leu Met Phe
 740 745 750
 gta aac gat gtt aac ttg gaa aac agc agt ctt gag gaa gct gta gaa 2363
 Val Asn Asp Val Asn Leu Glu Asn Ser Ser Leu Glu Glu Ala Val Glu
 755 760 765
 gca ctg aag gga gca ccg tca ggg act gtg aga ata gga gtt gct aag 2411
 Ala Leu Lys Gly Ala Pro Ser Gly Thr Val Arg Ile Gly Val Ala Lys
 770 775 780 785
 cct tta ccc ctt tca cca gaa gaa ggt tat gtt tct gct aag gag gat 2459
 Pro Leu Pro Leu Ser Pro Glu Glu Gly Tyr Val Ser Ala Lys Glu Asp
 790 795 800
 tcc ttt ctc tac cca cca cac tcc tgt gag gaa gca ggg ctg gct gac 2507
 Ser Phe Leu Tyr Pro Pro His Ser Cys Glu Glu Ala Gly Leu Ala Asp
 805 810 815
 aaa ccc ctc ttc agg gct gac ttg gct ctg gtg ggc aca aat gat gct 2555
 Lys Pro Leu Phe Arg Ala Asp Leu Ala Leu Val Gly Thr Asn Asp Ala
 820 825 830
 gac tta gta gat gaa tcc aca ttt gag tct cca tac tct cct gaa aat 2603
 Asp Leu Val Asp Glu Ser Thr Phe Glu Ser Pro Tyr Ser Pro Glu Asn
 835 840 845
 gac agc atc tac tct act caa gcc tct att tta tct ctt cat ggc agt 2651
 Asp Ser Ile Tyr Ser Thr Gln Ala Ser Ile Leu Ser Leu His Gly Ser
 850 855 860 865
 tct tgt ggt gat ggc ctg aac tat ggt tct tcc ctt cca tca tct cct 2699
 Ser Cys Gly Asp Gly Leu Asn Tyr Gly Ser Ser Leu Pro Ser Ser Pro
 870 875 880
 cct aag gat gtt att gaa aat tct tgt gat cca gta ctt gat ctg cat 2747
 Pro Lys Asp Val Ile Glu Asn Ser Cys Asp Pro Val Leu Asp Leu His
 885 890 895
 atg tct ctg gag gaa cta tat acc cag aat ctc ctg caa aga cag gat 2795
 Met Ser Leu Glu Glu Leu Tyr Thr Gln Asn Leu Leu Gln Arg Gln Asp
 900 905 910
 gag aat aca cct tcg gtg gac ata agt atg ggg cct gct tct ggc ttt 2843
 Glu Asn Thr Pro Ser Val Asp Ile Ser Met Gly Pro Ala Ser Gly Phe
 915 920 925
 act ata aat gac tac aca cct gca aat gct att gaa caa caa tat gaa 2891
 Thr Ile Asn Asp Tyr Thr Pro Ala Asn Ala Ile Glu Gln Gln Tyr Glu
 930 935 940 945
 tgt gaa aac aca ata gtg tgg act gaa tct cat tta cca agt gaa gtt 2939
 Cys Glu Asn Thr Ile Val Trp Thr Glu Ser His Leu Pro Ser Glu Val
 950 955 960
 ata tca agt gca gaa ctt cct tct gtg cta ccc gat tca gct gga aag 2987
 Ile Ser Ser Ala Glu Leu Pro Ser Val Leu Pro Asp Ser Ala Gly Lys
 965 970 975
 ggc tct gag tac ctg ctt gaa cag agc tcc ctg gcc tgt aat gct gag 3035
 Gly Ser Glu Tyr Leu Leu Glu Gln Ser Ser Leu Ala Cys Asn Ala Glu
 980 985 990
 tgt gtc atg ctt caa aat gta tct aaa gaa tct ttt gaa agg act att 3083
 Cys Val Met Leu Gln Asn Val Ser Lys Glu Ser Phe Glu Arg Thr Ile
 995 1000 1005
 aat ata gca aaa ggc aat tct agc cta gga atg aca gtt agt gct aat 3131
 Asn Ile Ala Lys Gly Asn Ser Ser Leu Gly Met Thr Val Ser Ala Asn
 1010 1015 1020 1025
 aaa gat ggc ttg ggg atg atc gtt cga agc att att cat gga ggt gcc 3179
 Lys Asp Gly Leu Gly Met Ile Val Arg Ser Ile Ile His Gly Gly Ala
 1030 1035 1040
 att agt cga gat ggc cgg att gcc att ggg gac tgc atc ttg tcc att 3227
 Ile Ser Arg Asp Gly Arg Ile Ala Ile Gly Asp Cys Ile Leu Ser Ile
 1045 1050 1055
 aat gaa gag tct acc atc agt gta acc aat gcc cag gca cga gct atg 3275
 Asn Glu Glu Ser Thr Ile Ser Val Thr Asn Ala Gln Ala Arg Ala Met
 1060 1065 1070
 ttg aga aga cat tct ctc att ggc cct gac ata aaa att act tat gtg 3323
 Leu Arg Arg His Ser Leu Ile Gly Pro Asp Ile Lys Ile Thr Tyr Val
 1075 1080 1085
 cct gca gaa cat ttg gaa gag ttc aaa ata agc ttg gga caa caa tct 3371
 Pro Ala Glu His Leu Glu Glu Phe Lys Ile Ser Leu Gly Gln Gln Ser
 1090 1095 1100 1105
 gga aga gta atg gca ctg gat att ttt tct tca tac act ggc aga gac 3419
 Gly Arg Val Met Ala Leu Asp Ile Phe Ser Ser Tyr Thr Gly Arg Asp
 1110 1115 1120
 att cca gaa tta cca gag cga gaa gag gga gag ggt gaa gaa agc gaa 3467
 Ile Pro Glu Leu Pro Glu Arg Glu Glu Gly Glu Gly Glu Glu Ser Glu
 1125 1130 1135
 ctt caa aac aca gca tat agc aat tgg aat cag ccc agg cgg gtg gaa 3515
 Leu Gln Asn Thr Ala Tyr Ser Asn Trp Asn Gln Pro Arg Arg Val Glu
 1140 1145 1150
 ctc tgg aga gaa cca agc aaa tcc tta ggc atc agc att gtt ggt gga 3563
 Leu Trp Arg Glu Pro Ser Lys Ser Leu Gly Ile Ser Ile Val Gly Gly
 1155 1160 1165
 cga ggg atg ggg agt cgg cta agc aat gga gaa gtg atg agg ggc att 3611
 Arg Gly Met Gly Ser Arg Leu Ser Asn Gly Glu Val Met Arg Gly Ile
 1170 1175 1180 1185
 ttc atc aaa cat gtt ctg gaa gat agt cca gct ggc aaa aat gga acc 3659
 Phe Ile Lys His Val Leu Glu Asp Ser Pro Ala Gly Lys Asn Gly Thr
 1190 1195 1200
 ttg aaa cct gga gat aga atc gta gag gtg gat gga atg gac ctc aga 3707
 Leu Lys Pro Gly Asp Arg Ile Val Glu Val Asp Gly Met Asp Leu Arg
 1205 1210 1215
 gat gca agc cat gaa caa gct gtg gaa gcc att cgg aaa gca ggc aac 3755
 Asp Ala Ser His Glu Gln Ala Val Glu Ala Ile Arg Lys Ala Gly Asn
 1220 1225 1230
 cct gta gtc ttt atg gta cag agc att ata aac aga cca agg gca ccc 3803
 Pro Val Val Phe Met Val Gln Ser Ile Ile Asn Arg Pro Arg Ala Pro
 1235 1240 1245
 agt cag tca gag tca gag cca gag aag gct cca ttg tgc agt gtg ccc 3851
 Ser Gln Ser Glu Ser Glu Pro Glu Lys Ala Pro Leu Cys Ser Val Pro
 1250 1255 1260 1265
 cca ccc cct cct tca gcc ttt gcc gaa atg ggt agt gat cac aca cag 3899
 Pro Pro Pro Pro Ser Ala Phe Ala Glu Met Gly Ser Asp His Thr Gln
 1270 1275 1280
 tca tct gca agc aaa atc tca caa gat gtg gac aaa gag gat gag ttt 3947
 Ser Ser Ala Ser Lys Ile Ser Gln Asp Val Asp Lys Glu Asp Glu Phe
 1285 1290 1295
 ggt tac agc tgg aaa aat atc aga gag cgt tat gga acc cta aca ggc 3995
 Gly Tyr Ser Trp Lys Asn Ile Arg Glu Arg Tyr Gly Thr Leu Thr Gly
 1300 1305 1310
 gag ctg cat atg att gaa ctg gag aaa ggt cat agt ggt ttg ggc cta 4043
 Glu Leu His Met Ile Glu Leu Glu Lys Gly His Ser Gly Leu Gly Leu
 1315 1320 1325
 agt ctt gct ggg aac aaa gac cga tcc agg atg agt gtc ttc ata gtg 4091
 Ser Leu Ala Gly Asn Lys Asp Arg Ser Arg Met Ser Val Phe Ile Val
 1330 1335 1340 1345
 ggg att gat cca aat gga gct gca gga aaa gat ggt cga ttg caa att 4139
 Gly Ile Asp Pro Asn Gly Ala Ala Gly Lys Asp Gly Arg Leu Gln Ile
 1350 1355 1360
 gca gat gag ctt cta gag atc aat ggt cag att tta tat gga aga agt 4187
 Ala Asp Glu Leu Leu Glu Ile Asn Gly Gln Ile Leu Tyr Gly Arg Ser
 1365 1370 1375
 cat cag aat gcc tca tca atc att aaa tgt gcc cct tct aaa gtg aaa 4235
 His Gln Asn Ala Ser Ser Ile Ile Lys Cys Ala Pro Ser Lys Val Lys
 1380 1385 1390
 ata att ttt atc aga aat aaa gat gca gtg aat cag atg gcc gta tgt 4283
 Ile Ile Phe Ile Arg Asn Lys Asp Ala Val Asn Gln Met Ala Val Cys
 1395 1400 1405
 cct gga aat gca gta gaa cct ttg cct tct aac tca gaa aat ctt caa 4331
 Pro Gly Asn Ala Val Glu Pro Leu Pro Ser Asn Ser Glu Asn Leu Gln
 1410 1415 1420 1425
 aat aag gag aca gag cca act gtt act act tct gat gca gct gtg gac 4379
 Asn Lys Glu Thr Glu Pro Thr Val Thr Thr Ser Asp Ala Ala Val Asp
 1430 1435 1440
 ctc agt tca ttt aaa aat gtg caa cat ctg gag ctt ccc aag gat cag 4427
 Leu Ser Ser Phe Lys Asn Val Gln His Leu Glu Leu Pro Lys Asp Gln
 1445 1450 1455
 ggg ggt ttg ggt att gct atc agc gaa gaa gat aca ctc agt gga gtc 4475
 Gly Gly Leu Gly Ile Ala Ile Ser Glu Glu Asp Thr Leu Ser Gly Val
 1460 1465 1470
 atc ata aag agc tta aca gag cat ggg gta gca gcc acg gat gga cga 4523
 Ile Ile Lys Ser Leu Thr Glu His Gly Val Ala Ala Thr Asp Gly Arg
 1475 1480 1485
 ctc aaa gtc gga gat cag ata ctg gct gta gat gat gaa att gtt gtt 4571
 Leu Lys Val Gly Asp Gln Ile Leu Ala Val Asp Asp Glu Ile Val Val
 1490 1495 1500 1505
 ggt tac cct att gaa aag ttt att agc ctt ctg aag aca gca aag atg 4619
 Gly Tyr Pro Ile Glu Lys Phe Ile Ser Leu Leu Lys Thr Ala Lys Met
 1510 1515 1520
 aca gta aaa ctt acc atc cat gct gag aat cca gat tcc cag gct gtt 4667
 Thr Val Lys Leu Thr Ile His Ala Glu Asn Pro Asp Ser Gln Ala Val
 1525 1530 1535
 cct tca gca gct ggt gca gcc agt gga gaa aaa aag aac agc tcc cag 4715
 Pro Ser Ala Ala Gly Ala Ala Ser Gly Glu Lys Lys Asn Ser Ser Gln
 1540 1545 1550
 tct ctg atg gtc cca cag tct ggc tcc cca gaa ccg gag tcc atc cga 4763
 Ser Leu Met Val Pro Gln Ser Gly Ser Pro Glu Pro Glu Ser Ile Arg
 1555 1560 1565
 aat aca agc aga tca tca aca cca gca att ttt gct tct gat cct gca 4811
 Asn Thr Ser Arg Ser Ser Thr Pro Ala Ile Phe Ala Ser Asp Pro Ala
 1570 1575 1580 1585
 acc tgc ccc att atc cct ggc tgc gaa aca acc atc gag att tcc aaa 4859
 Thr Cys Pro Ile Ile Pro Gly Cys Glu Thr Thr Ile Glu Ile Ser Lys
 1590 1595 1600
 ggg cga aca ggg ctg ggc ctg agc atc gtt ggg ggt tca gac acg ctg 4907
 Gly Arg Thr Gly Leu Gly Leu Ser Ile Val Gly Gly Ser Asp Thr Leu
 1605 1610 1615
 ctg ggt gcc att att atc cat gaa gtt tat gaa gaa gga gca gca tgt 4955
 Leu Gly Ala Ile Ile Ile His Glu Val Tyr Glu Glu Gly Ala Ala Cys
 1620 1625 1630
 aaa gat gga aga ctc tgg gct gga gat cag atc tta gag gtg aat gga 5003
 Lys Asp Gly Arg Leu Trp Ala Gly Asp Gln Ile Leu Glu Val Asn Gly
 1635 1640 1645
 att gac ttg aga aag gcc aca cat gat gaa gca atc aat gtc ctg aga 5051
 Ile Asp Leu Arg Lys Ala Thr His Asp Glu Ala Ile Asn Val Leu Arg
 1650 1655 1660 1665
 cag acg cca cag aga gtg cgc ctg aca ctc tac aga gat gag gcc cca 5099
 Gln Thr Pro Gln Arg Val Arg Leu Thr Leu Tyr Arg Asp Glu Ala Pro
 1670 1675 1680
 tac aaa gag gag gaa gtg tgt gac acc ctc act att gag ctg cag aag 5147
 Tyr Lys Glu Glu Glu Val Cys Asp Thr Leu Thr Ile Glu Leu Gln Lys
 1685 1690 1695
 aag ccg gga aaa ggc cta gga tta agt att gtt ggt aaa aga aac gat 5195
 Lys Pro Gly Lys Gly Leu Gly Leu Ser Ile Val Gly Lys Arg Asn Asp
 1700 1705 1710
 act gga gta ttt gtg tca gac att gtc aaa gga gga att gca gat gcc 5243
 Thr Gly Val Phe Val Ser Asp Ile Val Lys Gly Gly Ile Ala Asp Ala
 1715 1720 1725
 gat gga aga ctg atg cag gga gac cag ata tta atg gtg aat ggg gaa 5291
 Asp Gly Arg Leu Met Gln Gly Asp Gln Ile Leu Met Val Asn Gly Glu
 1730 1735 1740 1745
 gac gtt cgt aat gcc acc caa gaa gcg gtt gcc gct ttg cta aag tgt 5339
 Asp Val Arg Asn Ala Thr Gln Glu Ala Val Ala Ala Leu Leu Lys Cys
 1750 1755 1760
 tcc cta ggc aca gta acc ttg gaa gtt gga aga atc aaa gct ggt cca 5387
 Ser Leu Gly Thr Val Thr Leu Glu Val Gly Arg Ile Lys Ala Gly Pro
 1765 1770 1775
 ttc cat tca gag agg agg cca tct caa agc agc cag gtg agt gaa ggc 5435
 Phe His Ser Glu Arg Arg Pro Ser Gln Ser Ser Gln Val Ser Glu Gly
 1780 1785 1790
 agc ctg tca tct ttc act ttt cca ctc tct gga tcc agt aca tct gag 5483
 Ser Leu Ser Ser Phe Thr Phe Pro Leu Ser Gly Ser Ser Thr Ser Glu
 1795 1800 1805
 tca ctg gaa agt agc tca aag aag aat gca ttg gca tct gaa ata cag 5531
 Ser Leu Glu Ser Ser Ser Lys Lys Asn Ala Leu Ala Ser Glu Ile Gln
 1810 1815 1820 1825
 gga tta aga aca gtc gaa atg aaa aag ggc cct act gac tca ctg gga 5579
 Gly Leu Arg Thr Val Glu Met Lys Lys Gly Pro Thr Asp Ser Leu Gly
 1830 1835 1840
 atc agc atc gct gga gga gta ggc agc cca ctt ggt gat gtg cct ata 5627
 Ile Ser Ile Ala Gly Gly Val Gly Ser Pro Leu Gly Asp Val Pro Ile
 1845 1850 1855
 ttt att gca atg atg cac cca act gga gtt gca gca cag acc caa aaa 5675
 Phe Ile Ala Met Met His Pro Thr Gly Val Ala Ala Gln Thr Gln Lys
 1860 1865 1870
 ctc aga gtt ggg gat agg att gtc acc atc tgt ggc aca tcc act gag 5723
 Leu Arg Val Gly Asp Arg Ile Val Thr Ile Cys Gly Thr Ser Thr Glu
 1875 1880 1885
 ggc atg act cac acc caa gca gtt aac cta ctg aaa aat gca tct ggc 5771
 Gly Met Thr His Thr Gln Ala Val Asn Leu Leu Lys Asn Ala Ser Gly
 1890 1895 1900 1905
 tcc att gaa atg cag gtg gtt gct gga gga gac gtg agt gtg gtc aca 5819
 Ser Ile Glu Met Gln Val Val Ala Gly Gly Asp Val Ser Val Val Thr
 1910 1915 1920
 ggt cat cag cag gag cct gca agt tcc agt ctt tct ttc act ggg ctg 5867
 Gly His Gln Gln Glu Pro Ala Ser Ser Ser Leu Ser Phe Thr Gly Leu
 1925 1930 1935
 acg tca agc agt ata ttt cag gat gat tta gga cct cct caa tgt aag 5915
 Thr Ser Ser Ser Ile Phe Gln Asp Asp Leu Gly Pro Pro Gln Cys Lys
 1940 1945 1950
 tct att aca cta gag cga gga cca gat ggc tta ggc ttc agt ata gtt 5963
 Ser Ile Thr Leu Glu Arg Gly Pro Asp Gly Leu Gly Phe Ser Ile Val
 1955 1960 1965
 gga gga tat ggc agc cct cat gga gac tta ccc att tat gtt aaa aca 6011
 Gly Gly Tyr Gly Ser Pro His Gly Asp Leu Pro Ile Tyr Val Lys Thr
 1970 1975 1980 1985
 gtg ttt gca aag gga gca gcc tct gaa gac gga cgt ctg aaa agg ggc 6059
 Val Phe Ala Lys Gly Ala Ala Ser Glu Asp Gly Arg Leu Lys Arg Gly
 1990 1995 2000
 gat cag atc att gct gtc aat ggg cag agt cta gaa gga gtc acc cat 6107
 Asp Gln Ile Ile Ala Val Asn Gly Gln Ser Leu Glu Gly Val Thr His
 2005 2010 2015
 gaa gaa gct gtt gcc atc ctt aaa cgg aca aaa ggc act gtc act ttg 6155
 Glu Glu Ala Val Ala Ile Leu Lys Arg Thr Lys Gly Thr Val Thr Leu
 2020 2025 2030
 atg gtt ctc tct tgaattggct gccagaattg aaccaaccca acccctagct 6207
 Met Val Leu Ser
 2035
 cacctcctac tgtaaagaga atgcactggt cctgacaatt tttatgctgt gttcagccgg 6267
 gtcttcaaaa ctgtaggggg gaaataacac ttaagtttct ttttctcatc tagaaatgct 6327
 ttccttactg acaacctaac atcatttttc ttttcttctt gcattttgtg aacttaaaga 6387
 gaaggaatat ttgtgtaggt gaatctcgtt tttatttgtg gagatatcta atgttttgta 6447
 gtcacatggg caagaattat tacatgctaa gctggttagt ataaagaaag ataattctaa 6507
 agctaaccaa agaaaatggc ttcagtaaat taggatgaaa aatgaaaata taaaataaag 6567
 aagaaaatct cggggagttt aaaaaaaatg cctcaatttg gcaatctacc tcctctcccc 6627
 accccaaact aaaaaaagaa aaaaaggttt tctaatgaaa atctttaaaa atactgtcag 6687
 tattttaaaa ttttcaacag tattataaaa acattgcatc tccccacctc taatatgcat 6747
 atatattttt cctgctaaaa ttggtttcta caattgagta aatggcaaat acatgaagca 6807
 atgtccctaa attttataaa gaaattatat ttaatgcaca tttcaatttt cattcttatt 6867
 tttgaccttt tataaaatat tttcatgttg ctataagtaa atgatgatgc caccccatgt 6927
 tgactatggt ttttctagaa agcaactatg ctgctaacca tagaggaaca tagaagggtt 6987
 ccagaatctt tagtgctggt tttaacaacc gatgcaacat taaaaatgtg ttagtgtgct 7047
 gtgcaattgg ttttcaattc atattaatct taatgacaga gaacaatgtg ttactaatta 7107
 ttttggttgt atgccattag taaattgata gaaaaattaa ggggattaac ataacttcat 7167
 ttcattgcct tatattaaca tcttataata caatagttta agactaaggg aaacagatgg 7227
 agctgtttat tgagacaact ggtgaggaat tatcatgtgt tcattcccat tttagagcgt 7287
 gaaactccta cattagaata tataaagtca ctttaaatat ctatatttgt aacagaagta 7347
 gtgtacagat attttattac agcatttttg tgtaaatgca gaattaaagt gaataaataa 7407
 gaattttcag tggtgcaaaa aaaa 7431
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 3
 &lt;211&gt; LENGTH: 2037
 &lt;212&gt; TYPE: PRT
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 3
 Met Leu Glu Ala Ile Asp Lys Asn Arg Ala Leu His Ala Ala Glu Arg
 1 5 10 15
 Leu Gln Thr Lys Leu Arg Glu Arg Gly Asp Val Ala Asn Glu Asp Lys
 20 25 30
 Leu Ser Leu Leu Lys Ser Val Leu Gln Ser Pro Leu Phe Ser Gln Ile
 35 40 45
 Leu Ser Leu Gln Thr Ser Val Gln Gln Leu Lys Asp Gln Val Asn Ile
 50 55 60
 Ala Thr Ser Ala Thr Ser Asn Ile Glu Tyr Ala His Val Pro His Leu
 65 70 75 80
 Ser Pro Ala Val Ile Pro Thr Leu Gln Asn Glu Ser Phe Leu Leu Ser
 85 90 95
 Pro Asn Asn Gly Asn Leu Glu Ala Leu Thr Gly Pro Gly Ile Pro His
 100 105 110
 Ile Asn Gly Lys Pro Ala Cys Asp Glu Phe Asp Gln Leu Ile Lys Asn
 115 120 125
 Met Ala Gln Gly Arg His Val Glu Val Phe Glu Leu Leu Lys Pro Pro
 130 135 140
 Ser Gly Gly Leu Gly Phe Ser Val Val Gly Leu Arg Ser Glu Asn Arg
 145 150 155 160
 Gly Glu Leu Gly Ile Phe Val Gln Glu Ile Gln Glu Gly Ser Val Ala
 165 170 175
 His Arg Asp Gly Arg Leu Lys Glu Thr Asp Gln Ile Leu Ala Ile Asn
 180 185 190
 Gly Gln Ala Leu Asp Gln Thr Ile Thr His Gln Gln Ala Ile Ser Ile
 195 200 205
 Leu Gln Lys Ala Lys Asp Thr Val Gln Leu Val Ile Ala Arg Gly Ser
 210 215 220
 Leu Pro Gln Leu Val Ser Pro Ile Val Ser Arg Ser Pro Ser Ala Ala
 225 230 235 240
 Ser Thr Ile Ser Ala His Ser Asn Pro Val His Trp Gln His Met Glu
 245 250 255
 Thr Ile Glu Leu Val Asn Asp Gly Ser Gly Leu Gly Phe Gly Ile Ile
 260 265 270
 Gly Gly Lys Ala Thr Gly Val Ile Val Lys Thr Ile Leu Pro Gly Gly
 275 280 285
 Val Ala Asp Gln His Gly Arg Leu Cys Ser Gly Asp His Ile Leu Lys
 290 295 300
 Ile Gly Asp Thr Asp Leu Ala Gly Met Ser Ser Glu Gln Val Ala Gln
 305 310 315 320
 Val Leu Arg Gln Cys Gly Asn Arg Val Lys Leu Met Ile Ala Arg Gly
 325 330 335
 Ala Ile Glu Glu Arg Thr Ala Pro Thr Ala Leu Gly Ile Thr Leu Ser
 340 345 350
 Ser Ser Pro Thr Ser Thr Pro Glu Leu Arg Val Asp Ala Ser Thr Gln
 355 360 365
 Lys Gly Glu Glu Ser Glu Thr Phe Asp Val Glu Leu Thr Lys Asn Val
 370 375 380
 Gln Gly Leu Gly Ile Thr Ile Ala Gly Tyr Ile Gly Asp Lys Lys Leu
 385 390 395 400
 Glu Pro Ser Gly Ile Phe Val Lys Ser Ile Thr Lys Ser Ser Ala Val
 405 410 415
 Glu His Asp Gly Arg Ile Gln Ile Gly Asp Gln Ile Ile Ala Val Asp
 420 425 430
 Gly Thr Asn Leu Gln Gly Phe Thr Asn Gln Gln Ala Val Glu Val Leu
 435 440 445
 Arg His Thr Gly Gln Thr Val Leu Leu Thr Leu Met Arg Arg Gly Met
 450 455 460
 Lys Gln Glu Ala Glu Leu Met Ser Arg Glu Asp Val Thr Lys Asp Ala
 465 470 475 480
 Asp Leu Ser Pro Val Asn Ala Ser Ile Ile Lys Glu Asn Tyr Glu Lys
 485 490 495
 Asp Glu Asp Phe Leu Ser Ser Thr Arg Asn Thr Asn Ile Leu Pro Thr
 500 505 510
 Glu Glu Glu Gly Tyr Pro Leu Leu Ser Ala Glu Ile Glu Glu Ile Glu
 515 520 525
 Asp Ala Gln Lys Gln Glu Ala Ala Leu Leu Thr Lys Trp Gln Arg Ile
 530 535 540
 Met Gly Ile Asn Tyr Glu Ile Val Val Ala His Val Ser Lys Phe Ser
 545 550 555 560
 Glu Asn Ser Gly Leu Gly Ile Ser Leu Glu Ala Thr Val Gly His His
 565 570 575
 Phe Ile Arg Ser Val Leu Pro Glu Gly Pro Val Gly His Ser Gly Lys
 580 585 590
 Leu Phe Ser Gly Asp Glu Leu Leu Glu Val Asn Gly Ile Thr Leu Leu
 595 600 605
 Gly Glu Asn His Gln Asp Val Val Asn Ile Leu Lys Glu Leu Pro Ile
 610 615 620
 Glu Val Thr Met Val Cys Cys Arg Arg Thr Val Pro Pro Thr Thr Gln
 625 630 635 640
 Ser Glu Leu Asp Ser Leu Asp Leu Cys Asp Ile Glu Leu Thr Glu Lys
 645 650 655
 Pro His Val Asp Leu Gly Glu Phe Ile Gly Ser Ser Glu Thr Glu Asp
 660 665 670
 Pro Val Leu Ala Met Thr Asp Ala Gly Gln Ser Thr Glu Glu Val Gln
 675 680 685
 Ala Pro Leu Ala Met Trp Glu Ala Gly Ile Gln His Ile Glu Leu Glu
 690 695 700
 Lys Gly Ser Lys Gly Leu Gly Phe Ser Ile Leu Asp Tyr Gln Asp Pro
 705 710 715 720
 Ile Asp Pro Ala Ser Thr Val Ile Ile Ile Arg Ser Leu Val Pro Gly
 725 730 735
 Gly Ile Ala Glu Lys Asp Gly Arg Leu Leu Pro Gly Asp Arg Leu Met
 740 745 750
 Phe Val Asn Asp Val Asn Leu Glu Asn Ser Ser Leu Glu Glu Ala Val
 755 760 765
 Glu Ala Leu Lys Gly Ala Pro Ser Gly Thr Val Arg Ile Gly Val Ala
 770 775 780
 Lys Pro Leu Pro Leu Ser Pro Glu Glu Gly Tyr Val Ser Ala Lys Glu
 785 790 795 800
 Asp Ser Phe Leu Tyr Pro Pro His Ser Cys Glu Glu Ala Gly Leu Ala
 805 810 815
 Asp Lys Pro Leu Phe Arg Ala Asp Leu Ala Leu Val Gly Thr Asn Asp
 820 825 830
 Ala Asp Leu Val Asp Glu Ser Thr Phe Glu Ser Pro Tyr Ser Pro Glu
 835 840 845
 Asn Asp Ser Ile Tyr Ser Thr Gln Ala Ser Ile Leu Ser Leu His Gly
 850 855 860
 Ser Ser Cys Gly Asp Gly Leu Asn Tyr Gly Ser Ser Leu Pro Ser Ser
 865 870 875 880
 Pro Pro Lys Asp Val Ile Glu Asn Ser Cys Asp Pro Val Leu Asp Leu
 885 890 895
 His Met Ser Leu Glu Glu Leu Tyr Thr Gln Asn Leu Leu Gln Arg Gln
 900 905 910
 Asp Glu Asn Thr Pro Ser Val Asp Ile Ser Met Gly Pro Ala Ser Gly
 915 920 925
 Phe Thr Ile Asn Asp Tyr Thr Pro Ala Asn Ala Ile Glu Gln Gln Tyr
 930 935 940
 Glu Cys Glu Asn Thr Ile Val Trp Thr Glu Ser His Leu Pro Ser Glu
 945 950 955 960
 Val Ile Ser Ser Ala Glu Leu Pro Ser Val Leu Pro Asp Ser Ala Gly
 965 970 975
 Lys Gly Ser Glu Tyr Leu Leu Glu Gln Ser Ser Leu Ala Cys Asn Ala
 980 985 990
 Glu Cys Val Met Leu Gln Asn Val Ser Lys Glu Ser Phe Glu Arg Thr
 995 1000 1005
 Ile Asn Ile Ala Lys Gly Asn Ser Ser Leu Gly Met Thr Val Ser Ala
 1010 1015 1020
 Asn Lys Asp Gly Leu Gly Met Ile Val Arg Ser Ile Ile His Gly Gly
 1025 1030 1035 1040
 Ala Ile Ser Arg Asp Gly Arg Ile Ala Ile Gly Asp Cys Ile Leu Ser
 1045 1050 1055
 Ile Asn Glu Glu Ser Thr Ile Ser Val Thr Asn Ala Gln Ala Arg Ala
 1060 1065 1070
 Met Leu Arg Arg His Ser Leu Ile Gly Pro Asp Ile Lys Ile Thr Tyr
 1075 1080 1085
 Val Pro Ala Glu His Leu Glu Glu Phe Lys Ile Ser Leu Gly Gln Gln
 1090 1095 1100
 Ser Gly Arg Val Met Ala Leu Asp Ile Phe Ser Ser Tyr Thr Gly Arg
 1105 1110 1115 1120
 Asp Ile Pro Glu Leu Pro Glu Arg Glu Glu Gly Glu Gly Glu Glu Ser
 1125 1130 1135
 Glu Leu Gln Asn Thr Ala Tyr Ser Asn Trp Asn Gln Pro Arg Arg Val
 1140 1145 1150
 Glu Leu Trp Arg Glu Pro Ser Lys Ser Leu Gly Ile Ser Ile Val Gly
 1155 1160 1165
 Gly Arg Gly Met Gly Ser Arg Leu Ser Asn Gly Glu Val Met Arg Gly
 1170 1175 1180
 Ile Phe Ile Lys His Val Leu Glu Asp Ser Pro Ala Gly Lys Asn Gly
 1185 1190 1195 1200
 Thr Leu Lys Pro Gly Asp Arg Ile Val Glu Val Asp Gly Met Asp Leu
 1205 1210 1215
 Arg Asp Ala Ser His Glu Gln Ala Val Glu Ala Ile Arg Lys Ala Gly
 1220 1225 1230
 Asn Pro Val Val Phe Met Val Gln Ser Ile Ile Asn Arg Pro Arg Ala
 1235 1240 1245
 Pro Ser Gln Ser Glu Ser Glu Pro Glu Lys Ala Pro Leu Cys Ser Val
 1250 1255 1260
 Pro Pro Pro Pro Pro Ser Ala Phe Ala Glu Met Gly Ser Asp His Thr
 1265 1270 1275 1280
 Gln Ser Ser Ala Ser Lys Ile Ser Gln Asp Val Asp Lys Glu Asp Glu
 1285 1290 1295
 Phe Gly Tyr Ser Trp Lys Asn Ile Arg Glu Arg Tyr Gly Thr Leu Thr
 1300 1305 1310
 Gly Glu Leu His Met Ile Glu Leu Glu Lys Gly His Ser Gly Leu Gly
 1315 1320 1325
 Leu Ser Leu Ala Gly Asn Lys Asp Arg Ser Arg Met Ser Val Phe Ile
 1330 1335 1340
 Val Gly Ile Asp Pro Asn Gly Ala Ala Gly Lys Asp Gly Arg Leu Gln
 1345 1350 1355 1360
 Ile Ala Asp Glu Leu Leu Glu Ile Asn Gly Gln Ile Leu Tyr Gly Arg
 1365 1370 1375
 Ser His Gln Asn Ala Ser Ser Ile Ile Lys Cys Ala Pro Ser Lys Val
 1380 1385 1390
 Lys Ile Ile Phe Ile Arg Asn Lys Asp Ala Val Asn Gln Met Ala Val
 1395 1400 1405
 Cys Pro Gly Asn Ala Val Glu Pro Leu Pro Ser Asn Ser Glu Asn Leu
 1410 1415 1420
 Gln Asn Lys Glu Thr Glu Pro Thr Val Thr Thr Ser Asp Ala Ala Val
 1425 1430 1435 1440
 Asp Leu Ser Ser Phe Lys Asn Val Gln His Leu Glu Leu Pro Lys Asp
 1445 1450 1455
 Gln Gly Gly Leu Gly Ile Ala Ile Ser Glu Glu Asp Thr Leu Ser Gly
 1460 1465 1470
 Val Ile Ile Lys Ser Leu Thr Glu His Gly Val Ala Ala Thr Asp Gly
 1475 1480 1485
 Arg Leu Lys Val Gly Asp Gln Ile Leu Ala Val Asp Asp Glu Ile Val
 1490 1495 1500
 Val Gly Tyr Pro Ile Glu Lys Phe Ile Ser Leu Leu Lys Thr Ala Lys
 1505 1510 1515 1520
 Met Thr Val Lys Leu Thr Ile His Ala Glu Asn Pro Asp Ser Gln Ala
 1525 1530 1535
 Val Pro Ser Ala Ala Gly Ala Ala Ser Gly Glu Lys Lys Asn Ser Ser
 1540 1545 1550
 Gln Ser Leu Met Val Pro Gln Ser Gly Ser Pro Glu Pro Glu Ser Ile
 1555 1560 1565
 Arg Asn Thr Ser Arg Ser Ser Thr Pro Ala Ile Phe Ala Ser Asp Pro
 1570 1575 1580
 Ala Thr Cys Pro Ile Ile Pro Gly Cys Glu Thr Thr Ile Glu Ile Ser
 1585 1590 1595 1600
 Lys Gly Arg Thr Gly Leu Gly Leu Ser Ile Val Gly Gly Ser Asp Thr
 1605 1610 1615
 Leu Leu Gly Ala Ile Ile Ile His Glu Val Tyr Glu Glu Gly Ala Ala
 1620 1625 1630
 Cys Lys Asp Gly Arg Leu Trp Ala Gly Asp Gln Ile Leu Glu Val Asn
 1635 1640 1645
 Gly Ile Asp Leu Arg Lys Ala Thr His Asp Glu Ala Ile Asn Val Leu
 1650 1655 1660
 Arg Gln Thr Pro Gln Arg Val Arg Leu Thr Leu Tyr Arg Asp Glu Ala
 1665 1670 1675 1680
 Pro Tyr Lys Glu Glu Glu Val Cys Asp Thr Leu Thr Ile Glu Leu Gln
 1685 1690 1695
 Lys Lys Pro Gly Lys Gly Leu Gly Leu Ser Ile Val Gly Lys Arg Asn
 1700 1705 1710
 Asp Thr Gly Val Phe Val Ser Asp Ile Val Lys Gly Gly Ile Ala Asp
 1715 1720 1725
 Ala Asp Gly Arg Leu Met Gln Gly Asp Gln Ile Leu Met Val Asn Gly
 1730 1735 1740
 Glu Asp Val Arg Asn Ala Thr Gln Glu Ala Val Ala Ala Leu Leu Lys
 1745 1750 1755 1760
 Cys Ser Leu Gly Thr Val Thr Leu Glu Val Gly Arg Ile Lys Ala Gly
 1765 1770 1775
 Pro Phe His Ser Glu Arg Arg Pro Ser Gln Ser Ser Gln Val Ser Glu
 1780 1785 1790
 Gly Ser Leu Ser Ser Phe Thr Phe Pro Leu Ser Gly Ser Ser Thr Ser
 1795 1800 1805
 Glu Ser Leu Glu Ser Ser Ser Lys Lys Asn Ala Leu Ala Ser Glu Ile
 1810 1815 1820
 Gln Gly Leu Arg Thr Val Glu Met Lys Lys Gly Pro Thr Asp Ser Leu
 1825 1830 1835 1840
 Gly Ile Ser Ile Ala Gly Gly Val Gly Ser Pro Leu Gly Asp Val Pro
 1845 1850 1855
 Ile Phe Ile Ala Met Met His Pro Thr Gly Val Ala Ala Gln Thr Gln
 1860 1865 1870
 Lys Leu Arg Val Gly Asp Arg Ile Val Thr Ile Cys Gly Thr Ser Thr
 1875 1880 1885
 Glu Gly Met Thr His Thr Gln Ala Val Asn Leu Leu Lys Asn Ala Ser
 1890 1895 1900
 Gly Ser Ile Glu Met Gln Val Val Ala Gly Gly Asp Val Ser Val Val
 1905 1910 1915 1920
 Thr Gly His Gln Gln Glu Pro Ala Ser Ser Ser Leu Ser Phe Thr Gly
 1925 1930 1935
 Leu Thr Ser Ser Ser Ile Phe Gln Asp Asp Leu Gly Pro Pro Gln Cys
 1940 1945 1950
 Lys Ser Ile Thr Leu Glu Arg Gly Pro Asp Gly Leu Gly Phe Ser Ile
 1955 1960 1965
 Val Gly Gly Tyr Gly Ser Pro His Gly Asp Leu Pro Ile Tyr Val Lys
 1970 1975 1980
 Thr Val Phe Ala Lys Gly Ala Ala Ser Glu Asp Gly Arg Leu Lys Arg
 1985 1990 1995 2000
 Gly Asp Gln Ile Ile Ala Val Asn Gly Gln Ser Leu Glu Gly Val Thr
 2005 2010 2015
 His Glu Glu Ala Val Ala Ile Leu Lys Arg Thr Lys Gly Thr Val Thr
 2020 2025 2030
 Leu Met Val Leu Ser
 2035
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 4
 &lt;211&gt; LENGTH: 15
 &lt;212&gt; TYPE: PRT
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 4
 Asn Glu Pro Phe Asp Glu Asp Gln His Thr Gln Ile Thr Lys Val
 1 5 10 15
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 5
 &lt;211&gt; LENGTH: 21
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 5
 agacagcaaa gatgacagta a 21
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 6
 &lt;211&gt; LENGTH: 20
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 6
 cttcctcctc tttgtatggg 20
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 7
 &lt;211&gt; LENGTH: 21
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 7
 gcttttgccg aaatgggtag t 21
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 8
 &lt;211&gt; LENGTH: 21
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 8
 gatcggtctt tgttcccagc a 21
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 9
 &lt;211&gt; LENGTH: 19
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 9
 tgtgagcaag tttagtgag 19
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 10
 &lt;211&gt; LENGTH: 19
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 10
 ggtgattttc cccaagtaa 19
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 11
 &lt;211&gt; LENGTH: 16
 &lt;212&gt; TYPE: PRT
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 11
 Ser Gly Ser Gly Ile Leu Ala Pro Pro Val Pro Pro Arg Asn Thr Arg
 1 5 10 15
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 12
 &lt;211&gt; LENGTH: 16
 &lt;212&gt; TYPE: PRT
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 12
 Glu Asn Glu Pro Phe Asp Glu Asp Gln His Thr Gln Ile Thr Lys Val
 1 5 10 15
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 13
 &lt;211&gt; LENGTH: 22
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 13
 gccaccgcgg gattaagttt ct 22
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 14
 &lt;211&gt; LENGTH: 23
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 14
 tgtagccagc aatggtaatt cct 23
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 15
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 15
 gttttcccag tcacgacggt tccattttaa ttgctgttaa t 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 16
 &lt;211&gt; LENGTH: 43
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 16
 aggaaacagc tatgaccatg gggataataa aaacgattca ggg 43
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 17
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 17
 gttttcccag tcacgacgtt gaatatgccc acgttcctc 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 18
 &lt;211&gt; LENGTH: 42
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 18
 aggaaacagc tatgaccatt ctttcaatct tccatctcta tg 42
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 19
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 19
 gttttcccag tcacgacgtt gaatatgccc acgttcctc 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 20
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 20
 aggaaacagc tatgaccatc aaaccagatc catcattcac c 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 21
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 21
 gttttcccag tcacgacggc acaatttcag ctcactctaa 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 22
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 22
 aggaaacagc tatgaccatg gatgaggaga gggtgatgc 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 23
 &lt;211&gt; LENGTH: 22
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 23
 tctagcagga atgagcagtg ag 22
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 24
 &lt;211&gt; LENGTH: 24
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 24
 gatcctgata atctaaaatg ctaa 24
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 25
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 25
 gttttcccag tcacgacgaa gttgatgatt gcaagaggtg 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 26
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 26
 aggaaacagc tatgaccatg gtttgtgcca tctactgcta t 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 27
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 27
 gttttcccag tcacgacgaa acgagtgccg ttgagcatg 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 28
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 28
 aggaaacagc tatgaccatg ctgacagtaa tggataccct 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 29
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 29
 gttttcccag tcacgacgga ttttttatct tcgacgagaa a 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 30
 &lt;211&gt; LENGTH: 42
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 30
 aggaaacagc tatgaccatt tccccaagta aagttatgcc at 42
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 31
 &lt;211&gt; LENGTH: 38
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 31
 gttttcccag tcacgacgtc ctgttggaca cagcggga 38
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 32
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 32
 aggaaacagc tatgaccatc atggccaaag gtgcttgaa 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 33
 &lt;211&gt; LENGTH: 22
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 33
 ccacccacca cccaatcaga at 22
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 34
 &lt;211&gt; LENGTH: 22
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 34
 catctcgact aatggcacct cc 22
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 35
 &lt;211&gt; LENGTH: 38
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 35
 gttttcccag tcacgacgga gacagaggat ccagtgct 38
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 36
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 36
 aggaaacagc tatgaccatc cctgacggtg ctcccttca 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 37
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 37
 gttttcccag tcacgacgtt aacttggaaa acagcagtct 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 38
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 38
 aggaaacagc tatgaccatc atcaccacaa gaactgccat g 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 39
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 39
 gttttcccag tcacgacgac tctcctgaaa atgacagcat 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 40
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 40
 aggaaacagc tatgaccatt aaatgagatt cagtccacac t 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 41
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 41
 gttttcccag tcacgacgat aaatgactac acacctgcaa 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 42
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 42
 aggaaacagc tatgaccata acgatcatcc ccaagccatc t 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 43
 &lt;211&gt; LENGTH: 22
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 43
 ctgagtacct gcttgaacag ag 22
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 44
 &lt;211&gt; LENGTH: 22
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 44
 gaccattgat ctctagaagc tc 22
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 45
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 45
 gttttcccag tcacgacggg actattaata tagcaaaagg c 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 46
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 46
 aggaaacagc tatgaccatc agtgccatta ctcttccaga 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 47
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 47
 gttttcccag tcacgacgta cttatgtgcc tgcagaaca 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 48
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 48
 aggaaacagc tatgaccatc atgtttgatg aaaatgcccc 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 49
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 49
 gttttcccag tcacgacgat tgttggtgga cgagggatg 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 50
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 50
 aggaaacagc tatgaccatc catttcggca aaggctgaag 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 51
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 51
 gttttcccag tcacgacgca gagtcagagc cagagaagg 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 52
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 52
 aggaaacagc tatgaccata gaagctcagc tgcaatttgc 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 53
 &lt;211&gt; LENGTH: 21
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 53
 caggcgagct gcatatgatt g 21
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 54
 &lt;211&gt; LENGTH: 23
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 54
 cctcctttga caatgtctga cac 23
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 55
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 55
 gttttcccag tcacgacggt gtcttcatag tggggattga t 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 56
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 56
 aggaaacagc tatgaccatg aagctccaga tgttgcacat 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 57
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 57
 gttttcccag tcacgacgag agccaactgt tactacttc 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 58
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 58
 aggaaacagc tatgaccatt gaaggaacag cctgggaatc 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 59
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 59
 gttttcccag tcacgacgtt agccttctga agacagcaa 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 60
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 60
 aggaaacagc tatgaccatc atggataata atggcaccca 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 61
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 61
 gttttcccag tcacgacgtt tccaaagggc gaacagggc 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 62
 &lt;211&gt; LENGTH: 41
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 62
 aggaaacagc tatgaccatc caacaatact taatcctagg c 41
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 63
 &lt;211&gt; LENGTH: 23
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 63
 tggaattgac ttgagaaagg cca 23
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 64
 &lt;211&gt; LENGTH: 22
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 64
 ccccctacag ttttgaagac cc 22
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 65
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 65
 gttttcccag tcacgacgaa gaggaggaag tgtgtgacac 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 66
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 66
 aggaaacagc tatgaccatg acaggctgcc ttcactcacc 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 67
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 67
 gttttcccag tcacgacgtc aaagctggtc cattccatt 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 68
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 68
 aggaaacagc tatgaccatg gatgtgccac agatggtgac 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 69
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 69
 gttttcccag tcacgacgat gatgcaccca actggagtt 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 70
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 70
 aggaaacagc tatgaccatg gctgccatat cctccaacta 40
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 71
 &lt;211&gt; LENGTH: 39
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 71
 gttttcccag tcacgacggg acctcctcaa tgtaagtct 39
 &lt;200&gt; SEQUENCE CHARACTERISTICS:
 &lt;210&gt; SEQ ID NO 72
 &lt;211&gt; LENGTH: 40
 &lt;212&gt; TYPE: DNA
 &lt;213&gt; ORGANISM: Homo sapiens
 &lt;400&gt; SEQUENCE: 72
 aggaaacagc tatgaccata ttgtcaggac cagtgcattc 40