POLYNUCLEOTIDES ENCODING HUMAN AND MURINE ADHESION PROTEINS (BIgR)

Abstract: The present invention relates to isolated and purified polynucleotides encoding for a human adhesion protein or a murine adhesion protein.

Description: POLYNUCLEOTIDES ENCODING HUMAN AND MURINE ADHESION PROTEINS . BlgR. Technical Field of the Invention The present invention relates to molecular biology. More specifically, the present invention relates to polynucleotides which encode a human or a murine adhesion protein, polypeptides encoded by said polynucleotides and recombinant vectors expressing said polypeptides. Background of the Invention Cell adhesion is of prime importance for the formation and functional maintenance of multicellular organisms. Adhesion proteins can be classified as cell surface molecules that mediate intercellular bonds and/or participate in cell-substratum interactions. Their intracellular domains provide a functional link to the cytoskeleton and this appears to be important for efficient cell-cell adhesion to take place. They are expressed in characteristic spatiotemperal sequences. Different superfamilies have been described including immunoglobulin (hereinafter, "Ig"), cadherin, integrin, selectin (Aplin AE, Howe A, Alahari SK, Juliano RL, (1998) Pharmacol. Rev. 50:197-263). Adhesion proteins belonging to the immunoglobulin superfamily may operate in both a homotypic and/or heterotypic manner. The common building block is the Ig domain and the prototype is neural cell adhesion molecule (hereinafter, "NCAM") which possesses five Ig domains. This family participates in diverse biological functions including leukocyte-endothelial cell interactions, neural crest cell migration, neurite guidance and tumor invasion. It is well demonstrated that during inflammation members of the Ig superfamily interact with and participate in leukocyte adhesion, invasion and migration through the vessel wall (Gonzalez-Amaro R, Diaz-Gonzalez F, Sanchez-Madrid F, (1998) Drugs 56:977-88). Selectins are involved in the initial interactions (tethering/rolling) of leukocytes with activated endothelium, whereas integrins and Ig superfamily CAMs mediate the firm adhesion of these cells and their subsequent extravasation. Tight junctions (hereinafter, "TJ") and adherens junctions (hereinafter, "AJ") are specialized structures that occur between opposing cndothclial and epithelial cells. They form a scmipermeable intercellular diffusion barrier that is both dynamic and regulated. Obviously these structures must be disrupted, or reorganized, in order to facilitate leukocyte passage from the circulation. The platelet endothelial cell adhesion molecule, PECAM-1 , a member of the Ig superfamily of adhesion proteins, localizes to the lateral membranes between endothelial cells (Zocchi MR, Ferrero E, Leone BE, Rovere P, Bianchi E, Toninelli E, Pardi R, (1996) Eur. J. Immunol. 26:759-67). However, it is not associated with the TJ and AJ structures (Ayalon O, Sabanai H, Lampugnani MG, Dejana Ε, Geiger B, (1994) J. Cell. Bio. 126(l):247-58). The crucial role of PΕCAM-1 in paracellular migration of leukocytes to extravascular sites has been established (Muller WA, Weigl SA, Deng X, Phillips DM, (1993) J. Exp. Med. 178:449-60). Another cell-cell adhesion molecule of the Ig superfamily, nectin (hereinafter, "PRR"), is recruited to the AJ through interaction with a PDZ-domain in the novel protein afadin (Takahashi K, Nakanishi H, Miyahara M, Mandai K, Satoh K, Satoh A, Nishioka H, Aoki J, Nomoto A, Mizoguchi A, Takai Y, (1999) J. Cell. Biol. 145:539-49). Tight junctions are crucial structures for maintenance of the blood-brain (hereinafter, "BBB") and blood-retinal (hereinafter, "BRB") barriers. In some instances it may be desirable to selectively disrupt TJs. For example, disruption of the BBB may provide a method for transvascular delivery of therapeutic agents to the brain (Muldoon LL, Pagel MA, Kroll RA, Roman-Goldstein S, Jones RS, Neuwelt ΕA, (1999) Am. J. Neuroradiol. 20:217- 22). On the other hand, breakdown of the BBB is part of the pathology of multiple sclerosis and stroke. In this instance it would be desirable to prevent reorganization of the TJs. In 1998, a novel mouse junctional adhesion molecule (hereinafter, "JAM") was cloned and identified as an additional transmembrane protein component of the tight junction (Martin-Padura I, Lostaglio S, Schneemann M, Williams L, Romano M, Fruscella P, Panzeri C, Stoppacciaro A, Ruco L, Villa A, Simmons D, Dejana Ε, (1998) J. Cell. Biol. 142(1): 117- 27). JAM possesses two Ig domains; a single transmembrane and a short intracellular domain. Thus it belongs to the Ig superfamily of adhesion molecules and evidence suggests that it also influences the paracellular transmigration of immune cells. Whether its extracellular domain engages in heterotypic interactions remains to be elucidated. The nucleotide sequence of mouse JAM was utilized to identify novel putative adhesion proteins belonging to the Ig superfamily. Summary of the Invention The present invention relates to an isolated and purified human adhesion polynucleotide which encodes a human adhesion polypeptide or fragment thereof. Moreover, the present invention further relates to an isolated and purified polynucleotide having the nucleotide sequence of SEQ ID NO: 1. The human adhesion polypeptide has the amino acid sequence shown in SEQ ID NO:2. The present invention relates to an isolated and purified murine adhesion polynucleotide which encodes a murine adhesion polypeptide or fragment thereof. Moreover, the present invention further relates to an isolated and purified polynucleotide having the nucleotide sequence of SEQ ID NO:3. The murine adhesion polypeptide has the amino acid sequence shown in SEQ ID NO:4. The present invention also relates to a recombinant vector. This vector contains a polynucleotide having the nucleotide sequence of SEQ ID NO: l or SEQ ID NO:3, which encodes for either a human or murine adhesion protein. The polynucleotide is operatively linked to a promoter that controls expression of the nucleotide sequence and a termination segment. The present invention also relates to a host cell containing the recombinant vector. The host cell can be a bacterial cell, an animal cell or a plant cell. The present invention also relates to transgenic mammals carrying a null mutation of the polynucleotide or animals overexpressing the polynucleotide described herein. The present invention also relates to a transgenic mammal comprising a null mutation of the nucleotide sequence of SEQ ID NO:l or SEQ ID NO:3. Additionally, the present invention also relates to a transgenic mammal overexpressing the nucleotide sequence of SEQ ID NO: 1 or SEQ ID NO:3. Finally, the present invention relates to an antibody that binds to one or both of the hereinbefore described polypeptides. Brief Description of the Figures Fig. 1A shows the alignment of a homologous Expressed Sequence Tag (hereinafter referred to as "EST") obtained from the databases accessed through the home page of the National Center for Biotechnology Information at www.ncbi.nlm.nih.gov. with the open reading frame of a murine junctional adhesion protein (hereinafter referred to as "mouse JAM"). Identity is shown on the amino acid level. Fig IB shows the alignment of overlapping ESTs that encode various sections of a human brain immunoglobulin superfamily receptor (hereinafter referred to as "human BlgR") including the stop codon. Fig. 1 C summarizes the Rapid Amplification of cDNA Ends (hereinafter referred to as "RACE") procedure employed to obtain the full open reading frame of human BlgR. The longest clones identified from each reaction are aligned. Fig. 2 shows the alignment of the sequences from RACE reactions 1 and 2 with GenBank Accession AF062733. RACE reaction 2 isolated a clone with a 34 amino acid deletion compared to AF062733. Fig. 3 shows the full cDNA and amino acid sequence of the human BlgR. The predicted signal sequence and transmembrane domain are underlined. N-linked glycosylation sites are highlighted as are cysteine residues which form disulfide bonds within the immunoglobulin-like folds of the extracellular domain ♦, position and sequence (below) of the 34 amino acid insert. Fig. 4 shows the alignment of the intracellular domains of human BlgR with glycophorin C and drosophila Ncurcxin. Residues that arc conserved between at least two of the sequences are highlighted. Fig. 5 shows the complete cDNA and protein sequence of murine adhesion protein (hereinafter referred to as "mouse BlgR"). Conserved cysteine residues arc highlighted. Fig. 6 shows the alignment of mouse (upper) and human (lower) BlgR amino acid sequences. Fig. 7 shows transcripts that were identified on a multiple tissue Northern blot probed under high stringency with a [α- 32 P]dCTP BlgR probe. Arrows highlight human BlgR transcripts. Fig. 8 shows that transcripts were identified on a normalized human brain multiple tissue Northern blot probed under high stringency with a [α- 32 P]dCTP BlgR (A) or β-actin (B). Arrows highlight human BlgR transcripts. Fig. 9 is a Western blot of Cos cells, control and expressing BlgR, probed with a rabbit anti-BIgR antibody. Fig. 10 defines the subcellular localization of BlgR when expressed in CHO cells. Immunofluoresence was performed with the rabbit anti-BIgR antibody and viewed using a NoranTM Confocal laser-scanning microscope (Noran Instruments, Middleton, WI). Fig. 11 shows screening for BlgR counter-receptors on various leukocyte cell lines. Calcein loaded cells were added to BIgR-Fc captured in 96 well plates. Binding was performed in TBS (n=6) or TBS+lMn (n=6); Wells were washed, retained cells lysed and fluorescence quantitated with a fluorimeter at excitation 485/emission 530 nm. Data from a representative experiment. Average ±SEM. Detailed Description of the Invention 1. The Present Invention The present invention relates to isolated and purified polynucleotide sequences which encode for a human adhesion protein (referred to herein as "human BigR") and a murine adhesion protein (referred to herein as "mouse BlgR"). In another embodiment, the present invention relates to polypeptides for human BlgR and mouse BlgR. In yet another embodiment, the present invention relates to recombinant vectors which, upon expression, produce human BlgR or mouse BlgR. The present invention also relates to host cells transformed with these recombinant vectors. II. Sequence Listing The present application also contains a sequence listing that contains 10 sequences. The sequence listing contains nucleotide sequences and amino acid sequences. For the nucleotide sequences, the base pairs are represented by the following base codes: Symbol Meaning A A; adenine C C; cytosine G G; guanine T T; thymine u U; uracil M A or C R A or G W A or T/U s C or G Svmbol Meaning Y C or T/U K G or T/U V A or C or G; not T/U H A or C or T U; not G D A or G or T U; not C B C or G or T U; not A N (A or C or G or T/U) The amino acids shown in the application are in the L-form and are represented by the following amino acid-three letter abbreviations: Abbreviation Amino acid name Ala L-Alanine Arg L-Arginine Asn L-Asparagine Asp L-Aspartic Acid Asx L-Aspartic Acid or Asparaginc Cys L-Cysteine Glu L-Glutamic Acid Gin L-Glutamine Glx L-Glutamine or Glutamic Acid Gly L-Glycine His L-Histidine He L-Isoleucine Leu L-Leucine Lys L-Lysine Met L-Methionine Phe L-Phenylalanine Pro L-Proline Ser L-Serine Thr L-Threonine Tip L-Tryptophan Tyr L-Tyrosine Val L-Valine Xaa L-Unknown or other . Polvnucleotides In one aspect, the present invention provides an isolated and purified polynucleotide which encodes human BlgR. In another aspect, the present invention provides an isolated and purified polynucleotide which encodes mouse BlgR. These polynucleotides can be DNA molecules, such as gene sequences, cDNAs or synthetic DNAs. The DNA molecules can be double-stranded or single-stranded, and if single stranded may be the coding strand. In addition, the polynucleotides can be RNA molecules such as mRNAs. The present invention also provides non-coding strands (antisense) which are complementary to the coding sequences as well as RNA sequences identical to or complementary to those coding sequences. One of ordinary skill in the art will readily appreciate that corresponding RNA sequences contain uracil (U) in place of thymidine (T). In one embodiment, the polynucleotide of the present invention is an isolated and purified cDNA molecule that contains the coding sequence of human BlgR. An exemplary cDNA molecule is shown as SEQ ID NO: 1. In a second embodiment, the polynucleotide of the present invention is an isolated and purified cDNA molecule that contains the coding sequence of mouse BlgR. An exemplary cDNA molecule is shown as SEQ ID NO:3. As is well known in the art, because of the degeneracy of the genetic code, there arc numerous other DNA and RNA molecules that can code for the same polypeptides as those encoded by SEQ ID NO: 1 or SEQ ID NO:3 or portions or fragments thereof. The present invention also contemplates homologous polynucleotides having at least 70% homology to the sequence shown in SEQ ID NO: l or SEQ ID NO:3, preferably at least 80% homology, and most preferably at least 90% homology. The term "homology", as used herein, refers to a degree of complementarity. There may be partial homology or complete homology (i.e., identity). A partially complementary sequence is one that at least partially inhibits an identical sequence from hybridizing to a target nucleic acid; it is referred to using the functional term "substantially homologous." The inhibition of hybridization of the completely complementary sequence to the target sequence may be examined using a hybridization assay (Southern or Northern blot, solution hybridization and the like) under conditions of low stringency. A substantially homologous sequence or probe will compete for and inhibit the binding (i.e., the hybridization) of a completely homologous sequence or probe to the target sequence under conditions of low stringency. This is not to say that conditions of low stringency are such that non-specific binding is permitted; low stringency conditions require that the binding of two sequences to one another be a specific (i.e., selective) interaction. The absence of non-specific binding may be tested by the use of a second target sequence which lacks even a partial degree of complementarity (e.g., less than about 30% identity); in the absence of non-specific binding, the probe will not hybridize to the second non-complementary target sequence. Moreover, the present invention also contemplates naturally occurring allelic variations and mutations of the cDNA sequence set forth above so long as those variations and mutations code, on expression, for the human or murine adhesion protein of the present invention. The present invention also encompasses splice variations of the human and murine BlgR polynucleotides. SUBSTTTUTE SHEET (RULE 26) The polynucleotide of the present invention can be used in marker-aided selection using techniques which arc well-known in the art. Marker-aided selection does not require the complete sequence of the gene. Instead, partial sequences can be used as hybridization probes or as the basis for oligonucleotide primers to amplify by PCR or other methods to identify nucleotides specific for human or murine adhesion proteins in other mammals. IV. Polypeptides The present invention also provides for polypeptides which encode for human BlgR and mouse BlgR. The amino acid sequence for human BlgR protein is provided in SEQ ID NO:2 and contains 398 amino acid residues. The amino acid sequence for the mouse BlgR adhesion protein is provided in SEQ ID NO:4 and contains 396 amino acid residues. The present invention also contemplates amino acid residue sequences that are substantially duplicative of the sequences set forth herein such that those sequences demonstrate like biological activity to the disclosed sequences. Such contemplated sequences include those sequences characterized by a minimal change in amino acid residue sequence or type (e.g., conservatively substimted sequences) which insubstantial change does not alter the basic nature and biological activity of the polypeptides. It is well known in the art that modifications and changes can be made in the structure of a polypeptide without substantially altering the biological function of that peptide. For example, certain amino acids can be substituted for other amino acids in a given polypeptide without any appreciable loss of function. In making such changes, substitutions of like amino acid residues can be made on the basis of relative similarity of side-chain substitutents, for example, their size, charge, hydrophobicity, hydrophilicity, and the like. As detailed in United States Patent No. 4,554,101, incorporated herein by reference, the following hydrophilicity values have been assigned to amino acid residues: Arg (+3.0); Lys (+3.0); Asp (+3.0); Glu (+3.0); Ser (+0.3); Asn (+0.2); Gin (+0.2); Gly (0); Pro (-0.5); Thr (-0.4); Ala (-0.5); His (-0.5); Cys (-1.0); Met (-1.3); Val (-1.5); Leu (-1.8); He (-1.8); Tyr (-2.3); Phe (-2.5); and Trp (-3.4). It is understood that an amino acid residue can be 9 SUBSTTTUTE SHEET (RULE 26) substituted for another having a similar hydrophilicity value (e.g., within a value of plus or minus 2.0) and still obtain a biologically equivalent polypeptide. In a similar manner, substitutions can be made on the basis of similarity in hydropathic index. Each amino acid residue has been assigned a hydropathic index on the basis of its hydrophobicity and charge characteristics. Those hydropathic index values are: He (+4.5); Val (+4.2); Leu (+3.8); Phe (+2.8); Cys (+2.5); Met (+1.9); Ala (+1.8); Gly (-0.4); Thr (-0.7); Ser (-0.8); Tφ (-0.9); Tyr (-1.3); Pro (-1.6); His (-3.2); Glu (-3.5); Gin (-3.5); Asp (-3.5); Asn (-3.5); Lys (-3.9); and Arg (-4.5). In making a substitution based on the hydropathic index, a value of within plus or minus 2.0 is preferred. V. Recombinant Vectors The present invention also relates to recombinant vectors which contain the polynucleotide of the present invention, host cells which are genetically engineered with recombinant vectors of the present invention and the production of the polypeptide of the present invention by recombinant techniques. The polynucleotide of the present invention can be employed for producing polypeptides using recombinant techniques which are well known in the art. For example, the polynucleotide may be included in any one of a variety of expression vectors for expressing a polypeptide. Such vectors include chromosomal, nonchromosomal and synthetic DNA sequences, e.g., derivatives of SV40, bacterial plasmids, phage DNA, baculovirus, yeast plasmids, vectors derived from combinations of plasmids and phage DNA, viral DNA such as vaccinia, adenovirus, fowl pox virus, and pseudorabies. One of the most popular vectors for obtaining genetic elements is from the well known cloning vector pBR322 (available from the American Type Culture Collection, Manassas, Virginia as ATCC Accession Number 37017). The pBR322 "backbone" sections can be combined with an appropriate promoter and the structural sequence to be expressed. However, any other vector may be used as long as it is replicable and viable in the host. The polynucleotide sequence of the present invention may be inserted into one of the hereinbefore mentioned recombinant vectors, in a forward or reverse orientation. A variety of procedures, which arc well known in the art may be used to achieve this. In general, the polynucleotide is inserted into an appropriate restriction endonucleasc site(s). When inserted into an appropriate expression vector, the polynucleotide of the present invention is operatively linked to an appropriate expression control sequence(s), such as a promoter, to direct mRNA synthesis. As used herein, the term "operatively linked" includes reference to a functional linkage between a promoter and a second sequence, wherein the promoter sequence initiates and mediates transcription of the DNA sequence corresponding to the second sequence. Generally, operably linked means that the nucleotide sequences being linked are contiguous and, where necessary to join two protein coding regions, contiguous and in the same reading frame. The heterologous structural sequence can encode a fusion protein including either an N-terminal or C-terminal identification peptide imparting desired characteristics, such as stablization or simplified purification of expressed recombinant product. Promoter regions can be selected from any desired gene using chloramphenicol transferase (CAT) vectors or other vectors with selectable markers. Such promoters can be derived from operons encoding glycolytic enzymes such as 3-phosphoglycerate kinase (PGK), α-factor, acid phosphatase, or heat shock proteins. Examples of bacterial promoters which can be used include, but are not limited to, lad, lacZ, T3, T7, gpt, lambda P R , P L and tφ. Eukaryotic promoters include CMV immediate early, HSV thymidine kinase, early and late SV40, LTRs from retrovirus, and mouse metallothionein-I. Examples of other promoters that can be used include the polyhedrin promoter of baculovirus. Typically, recombinant expression vectors contain an origin of replication to ensure maintenance of the vector. They preferably contain one or more selectable marker genes to provide a phenotypic trait for selection of transformed host cells. Examples of selectable marker genes which can be used include, but are not limited to, dihydrofolate reductase, 11 SUBSTTTUTE SHEET (RULE 26) neomycin or blasticidin resistance for eukaryotic cell culture, tetracycline or ampicillin resistance for E. coli. and the TRP1 gene for S. cerevisiae. The expression vector may also contain a ribosome binding site for translation initiation and a transcription termination segment. The vector may also include appropriate sequences for amplifying expression. Appropriate cloning and expression vectors for use with prokaryotic and eukaryotic hosts are described in Sambrook, et al., Molecular Cloning: A Laboratory Manual, Second Edition, Cold Spring Harbor, N.Y., (1989), which is herein incoφorated by reference. Large numbers of suitable vectors and promoters are commercially available and can be used in the present invention. Examples of vectors which can be used include, but are not limited to: Bacterial: pQE70, pQE60, pQE-9 (Qiagen), pBS, pDIO, phagescript, psiX174, pBluescript SK, pSKS, pNH8A, pkrHlόa, pNH18A, pNH46A (Stratagene), ptrc99a, PKK223-3, pKK233-3, pDR540, pRIT5 (Pharmacia), pGEM (Promega). Eukaryotic: pWLNEO, pSV2CAT, p)G44, pXTl, pSG (Stratagene), pSVK3, pBPV, pMSG, pSVL (Pharmacia), pcDNA3, pcDNA6 (InVitrogen). In another embodiment, the present invention relates to host cells containing the hereinbefore described recombinant vectors. The vector (such as a cloning or expression vector ) containing the hereinbefore described polynucleotide, may be employed to transform, transduce or transfect an appropriate host to permit the host to express the protein. Appropriate hosts which can be used in the present invention, include, but are not limited to prokaryotic cells such as E. coli, Streptomyces, Bacillus subtilis, Salmonella typhimurium, as well as various species within the general Pseudomonas, Streptomyces, and Staphylococcu . Lower eukaryotic cells such as yeast and insect cells such as Drosophila S2 and Spodoptera Sf9 and Sf21. Introduction of the recombinant construct into the host cell can be effected by calcium phosphate, DEAE-Dextran or liposome mediated transfection, or electroporation (see, Davis, L., Dibner, M., Battey, L. Basic Methods in Molecular Biology, (1986), herein incoφorated by reference). Various higher eukaryotic cells such as mammalian cell culture systems can also be employed to express recombinant protein. Examples of mammalian expression systems include the COS-7 lines of monkey kidney fibroblasts, described by Gluzman, Cell, 23: 175 ( 1981 ), and other cell lines capable of expressing a compatible vector, for example, the C127, 3T3, CHO, HeLa and BHK cell lines. Mammalian expression vectors will contain an origin of replication, a suitable promoter and enhancer, and any necessary ribosomc binding sites, polyadenylation site, splice donor and acceptor sites, transcriptional termination sequences, and 5' flanking nontranscribed sequences. DNA sequences derived from the SV40 splice, and polyadenylation sites may be used to provide the required nontranscribed genetic elements. The engineered host cells can be cultured in conventional nutrient media modified as appropriate for activating promoters, selecting transformants or amplifying genes encoding for the human junctional adhesion protein of the present invention. The culture conditions, such as temperature, pH and the like, are those previously used with the host cell selected for expression and can be determined experimentally, using techniques which are well known in the art. Transcription of the polynucleotide encoding the polypeptides of the present invention by higher eukaryotes can be increased by inserting an enhancer sequence into the vector. Enhancers are cis-acting elements of DNA which are about from 10 to about 300 base pairs in length, which act on a promoter to increase its transcription. Examples of suitable enhancers which can be used in the present invention include the SV40 enhancer on the late side of the replication origin base pairs 100 to 270, a cytomegalovirus early promoter enhancer, the polyoma enhancer on the late side of the replication origin, and adenovirus enhancers. Following transformation of a suitable host strain and growth of the host strain to an appropriate cell density, the selected promoter is induced by appropriate means (such as temperamre shift or chemical induction) and cells are cultured for an additional period. Cells are typically harvested by centrifugation, disrupted by physical or chemical means, and the resulting crude extract retained for further purification. Microbial cells employed in expression of proteins can be disrupted by any convenient method, including freeze -thaw 13 SUBSTTTUTE SHEET (RULE 26) cycling, sonication, mechanical disruption, or use of cell lysing agents. Such methods are well-known to those skilled in the art. The polypeptides of the present invention can be recovered and purified from recombinant cell cultures, the cell mass or otherwise according to methods of protein chemistry which are known in the art. For example, ammonium sulfate or ethanol precipitation, acid extraction, and various forms of chromatography e.g. anion / cation exchange, phosphocellulose, hydrophobic interaction, affinity chromatography including immunoaffinity, lectin and hydroxylapatite chromatography. Other methods may include dialysis, ultrafiltration, gelfiltration, SDS-PAGE and isoelectric focusing. Protein refolding steps can be used, as necessary, in completing configuration of the mature protein. Finally, high performance liquid chromatography (hereinafter, "HPLC") on normal or reverse systems or the like, can be employed for final purification steps. BlgR may be used to inactivate the endogenous gene by homologous recombination and thereby create a BlgR deficient cell, tissue or animal. Such cells, tissue or animals may then be used to define specific in vivo processes normally dependent upon BlgR. The cDNA sequence can be used to prepare stable cell lines expressing either wt BlgR or BlgR mutated at pertinent positions to determine which part of the molecule is responsible for function. Stable or transient cell lines can be created with BlgR possessing a tag at either the 5' or 3' end, e.g. V5 or HA epitope, to enable monitoring of BlgR function/modification/cellular interactions. The extracellular sequence of BlgR can be used to make recombinant protein fused to the Fc region of human or mouse IgG. This protein can be used: a) To screen for a BlgR ligand. If BlgR is an adhesion protein, interactions with, but not limited to, leucocytes/neutrophils will be analyzed. Briefly, BIgR-Fc fusion can be captured on ELISA plates. Cultured cells, control or stimulated with inflammatory cytokines, can be labeled with calcien dye, incubated with the immobilized BIgR-Fc, washed and fluorescence monitored. Alternatively, BlgR can be coupled to a solid support and then used to prepare a column for purification of solubilized proteins derived from various cells/tissues. Peptide sequencing could then be used to identify the ligand. Another approach would be to bind the BlgR-Fc to cell lysatcs and perform cross-linking with DSS. b) Upon identification of a BlgR ligand, the BIgR-Fc can be used to screen for a small molecule inhibitor of BlgR heterotypic or homotypic interactions. c) As a tool to neutralize BlgR function, either heterotypic or homotypic interactions. d) If BlgR displays homotypic/heterotypic interactions, the fusion can be used to bind to either BlgR or its ligand to initiate a cellular response. Recombinant protein derived from the extracellular domain can be used to analyze homotypic interactions. Such protein would not possess a Fc Tag. Single immunoglobulin- like domains can be made to determine which one is responsible for homotypic interactions. The interactions of the separate domains with each other or with a recombinant form possessing all three Ig-like domains may be assessed by various means. Examples are cross- linking with DSS, analytical ultracentrifugation or sizing columns. The BlgR sequence can be used to identify antisense oligonucleotides for inhibition of BlgR function in cell systems. Further, degenerate oligonucleotides may be designed to aid in the identification of additional members of this family by the polymerase chain reaction. Alternatively, low stringency hybridization of cDNA libraries may be performed with BlgR sequence to identify closely related sequences. The intracellular domain of BlgR can be used to "fish" for novel interacting partners in the yeast two-hybrid system. Additionally, recombinant purified BlgR, and cell lines expressing recombinant BlgR, can be used to screen for small molecule inhibitors of BlgR. Apart from low expression in the placenta, BlgR is located exclusively in the brain. BlgR may be located in glial, neuronal or endothelial cells. If expressed in brain endothelial cells then it may play a role in the blood brain barrier. If BlgR is an adhesion protein, it is believed that it might play roles in, but not be limited to, neuronal plasticity, growth cone guidance and neurite outgrowth. It might possibly be involved in stroke and multiple sclerosis. VI. Antibodies The polypeptides of the present invention, fragments thereof, or cells expressing said polypeptides can be used as an immunogen to produce antibodies. These antibodies can be, for example, polyclonal or monoclonal antibodies. The present invention also includes chimeric, single chain, and humanized antibodies, as well as Fab fragments, or the product of a Fab expression library. Antibodies generated against the polypeptides of the present invention can be obtained by administering the polypeptides to an animal, preferably a nonhuman. Even a sequence encoding only a fragment of a polypeptide of the present invention can be used to generate antibodies binding to the whole native polypeptide. Such antibodies can then be used to isolate the polypeptide from tissue expressing that polypeptide. For preparation of monoclonal antibodies, any technique which provides antibodies produced by continuous cell line cultures can be used. Examples include the hybridoma technique (described by Kohler and Milstein, 1975, Nature, 256:495-497, herein incoφorated by reference), the trioma technique, the human B-cell hybridoma technique (described by Kozbor et al., 1983, Immunology Today 4:72, herein incoφorated by reference), and the EBV-hybridoma technique to produce human monoclonal antibodies (described by Cole, et al., 1985, in Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc., pp-77-96, herein incoφorated by reference). 16 SUBSTTTUTE SHEET (RULE 26) For preparation of polyclonal antibodies any technique known in the art which provides such antibodies can be used. Techniques for the production of single chain antibodies, such as those described in U.S. Patent 4,946,778, herein incoφoratcd by reference, can be adapted to produce single chain antibodies to immunogenic polypeptides of the present invention. In yet another embodiment, the present invention relates to a method for identifying human or murine cells expressing endogenous human or murine adhesion proteins. The method involves identifying cells which express proteins having the same function as the human or murine adhesion proteins described herein and then characterizing the requisite cells producing said proteins using techniques known in the art. Antibodies, prepared pursuant to the techniques described herein, can be used to screen for BlgR and to identify cells (diseased or normal) in a subject (human or animal) which express endogenous BlgR. The antibodies of the present invention can be used to: a) Probe subcellular localization/expression of endogenous BlgR in cells (diseased or normal) in a subject (human or animal). b) Immunoprecipitate endogenous BlgR protein from cells or recombinant BlgR from transfected cells, to determine whether it is modified by glycosylation, phosphorylation, etc. Co-precipitation of proteins that associate with BlgR can also be identified in this manner. c) As a tool to neutralize BlgR function, either heterotypic or homotypic interactions. The anti-BIgR antibody may be administered in vivo in various animal models in order to perturb BlgR function. Alternatively, proof of concept studies may be conducted in vitro. d) Map functional epitopes of the BlgR molecule. By way of example, and not of limitation, examples of the present invention shall 17 SUBSTTTUTE SHEET (RULE 26) now be given. EXAMPLE I: CLONING OF HUMAN BlgR cDNA AND MAPPING OF THE BlgR GENE. The polynucleotide sequences shown in SEQ ID NO: 1 and SEQ ID NO:3 were cloned using a combination of electronic and conventional cloning techniques. The electronic techniques used involved utilizing the Expressed Sequence Tag (EST) databases accessed through the home page of the National Center for Biotechnology Information (NCBI) at www.ncbi.nlm.nih.gov. As a template for electronic cloning the cDNA sequence of a novel mouse Junctional Adhesion Protein (JAM) published by Martin-Padura I, Lostaglio S, Schneemann M, Williams L, Romano M, Fruscella P, Panzeri C, Stoppacciaro A, Ruco L, Villa A, Simmons D, Dejana E, J. Cell. Biol. (1998) 142(1): 117-27, herein incoφorated by reference, was used. The mouse JAM cDNA sequence is also available on GenBank (Accession No. U89915). The advanced Basic Local Alignment Search Tool (BLAST 2.0) was used to identify ESTs displaying homology with mouse JAM. Electronic Cloning The complete mouse JAM peptide sequence (Ace. No. U89915) was searched for homology with human EST sequences using the tblastn program which compares a protein query sequence against a nucleotide sequence database dynamically translated in all reading frames. The complete mouse JAM protein sequence is 300 amino acids in length. The initiation codon begins at base pair (hereinafter "bp") 71 and the stop codon at 971 (see Fig. 1A). Some 116 amino acids encoded within EST R88252 showed 38% similarity and 28% identity over a 129 amino acid stretch of mouse JAM (see Fig 1A). A subsequent search of the SWISS-PROT protein sequence database revealed that this EST displayed similarities to other known adhesion proteins. Based on this, and the conservation of cysteine residues important in the formation of the immunoglobulin-like fold, EST R88252 was analyzed further. Throughout the assembly of the virtual sequence, the translation was continually monitored in all reading frames in order to identify the putative codons for initiation and 18 SUBSTTTUTE SHEET (RULE 25) termination of the virtual protein. Where available, examination of overlapping ESTs was conducted to verify sequence/errors. The 5' 150 bp of R88252 was blasted through the nr human dbEST (a non-redundant sub-category of the EST database containing only human ESTs) using the blastn program. EST T08949 showed 100% identity over a 109 bp overlap and gave the most additional 5'- sequence (see Fig. IB). A blastn analysis of T08949 through the nr human EST database was unsuccessful in isolating additional bases at the 5' end. Thus, a putative initiation codon could not be identified for this protein by this method. The final 120 bp of R88252 was blasted through the nr human dbEST using the blastn program. HI 4720 showed 97% identity over 229 base pairs and gave 147 additional bases at the 3' end (see Fig. IB). The 3' 106 bp of HI 4720 was blasted through the nr human dbEST using the blastn program. The EST R15338 showed 95% identity over 170 base pairs and lengthened the novel sequence by 367 bases (see Fig. IB). Within this sequence a putative stop codon was identified. Thus, further searching of the database was terminated. Conventional Cloning In order to obtain further 5' sequence for this cDNA, a RACE technique was performed. Human fetal brain mRNA was reverse transcribed with AMV reverse transcriptase (Clontech Palo Alto, CA.) at 42°C. Amplification reactions were performed using the Marathon cDNA amplification kit (Clontech) and either of two antisense primers (see below). The following program utilized: 1 cycle at 94°C, 30sec; 5 cycles at 94°C, 5sec and 72°C, 4min; 5 cycles at 94°C, 5sec and 70°C, 4min; 25 cycles at 94°C, 5sec and 68°C, 4min. Products were ligated into the pCR-Blunt II-TOPO vector (Invitrogen) and sequenced using an ABI sequencer (Seqwright, TX). The primer used for the first RACE reaction was directed at a site 176 bp into the EST T08949 (5'- CCCAGAAGACTGACAGTTTAGGGTGGCTG-3') (SEQ ID NO:5). A 19 SUBSTTTUTE SHEET (RULE 26) product from this reaction allowed an additional 272 bp of BlgR to be isolated (see Fig. 1C). The primer used for the second RACE reaction was directed at a site 86 bp into the EST T08949 (5'-GCGCAGTGACGAGGGACTTGGCAGTTC-3') (SEQ ID NO:6). The longest product sequenced gave an extension of 232 bp from the 5' end of T08949 bp (see Fig. 1C). Most interestingly, an alignment of the sequences from each RACE reaction showed that at some 204 nucleotides upstream of the 5' end of T08949, the sequences diverged (see Fig. 3). Neither product possessed a putative consensus initiation ATG with upstream stop codon. Coincident with this result, further searching of the databases showed that a recent release of GenBank (March 4 th 1999) contained a cDNA sequence (Accession No. AF062733) the alignment of which showed that the product of RACE reaction 1 fell 41 amino acids short of the full-open reading frame clone (see Fig. 2). Further, a comparison of the sequence obtained from RACE reaction 2 demonstrated that a splice variant of AF062733 with a 34 amino acid deletion (see Fig. 2) had been isolated. Construction of a Full-Length BlgR In order to construct full-length human BlgR, a sense oligonucleotide directed against Accession No. AF062733 in the 5 '-untranslated region was designed, some 45 bp upstream of the ATG. This oligonucleotide 5 '-TTCAGGCTCGCCAGCGCCCAG-3' (SEQ ID NO:7) was coupled with an antisense primer 5'-CTAGATGAAATATTCCTTCTTGTCGTC-3' (SEQ ID NO: 8) that incoφorated the TAG stop codon. Human fetal brain mRNA was reverse transcribed and amplified using the following program: 1 cycle, 95°C for 7 min; 35 cycles, 95°C for 20 s, 60°C for 20 s, 72°C for 20 s; 1 cycle, 72°C for 5 min. Products were ligated into an E. coli vector and sequenced using an ABI sequencer (Seqwright, TX). Sequence Features The polymerase chain reaction allowed the identification of two different full-length isoforms of BlgR. Consistent with the RACE reaction, clones were isolated with an 20 SUBSTTTUTE SHEET (RULE 26) additional 34 amino acids just following the signal sequence, and clones in which these residues were spliced out. In Figure 3, the complete coding region of the shorter splice variant that possesses 398 amino acids is displayed. Below this, the sequence corresponding to the 34 amino acid (SEQ ID NO: 1 1 ) insert is shown. Sequencing proved these residues to be identical to those of GenBank accession AF062733. BlgR features a putative signal sequence (underlined), three immunoglobulin-like folds, a single transmembrane domain (underlined) and a short intracellular domain (see Fig. 3). Thus, the protein belongs to the immunoglobulin superfamily. The cysteine residues predicted to form disulfide bonds within each immunoglobulin-like domain are highlighted. The 1 st and 2 nd cysteine are located in the first, the 3 rd and 4 th in the second, and the 5 th and 6 th in the third immunoglobulin-like fold respectively. Highlighted are two consensus N-linked glycosylation sites (NxS/T) at amino acid 25 and 353 (although the first falls within the predicted signal sequence). Differences between BlgR polypeptide and GenBank Accession No. AF062733 The BlgR polypeptide shown in SEQ ID NO: 2 is different from AF062733 at two places. First, the polypeptide of the present invention is a smaller isoform because it has 34 amino acids which are deleted (see Figs. 2, 3). Second, a single glutamic acid deletion occurs at position 104 in SEQ ID NO:2, regardless of whether the 34 amino acids are retained or spliced out. Thus SEQ ID NO:2 reads, from amino acid 100, ALAD_EGEY while AF062733 reads ALADEEGEY in this same region. Proteins Homologous to BlgR Extracellular and Intracellular Domains BlgR is not a member of the family of junctional adhesion molecules. Nevertheless, a blast analysis demonstrates that its various immunoglobulin-like domains show similarity to other novel putative adhesion molecules. Most interestingly, the intracellular domain of BlgR showed 66% similarity and 53% identity to glycophorin C, a protein required for anchoring protein 4.1 in red blood cells. Further, this homology is also displayed with the Drosophila transmembrane protein Neurexin IV, which is required for localization of the drosophila 4.1 protein, coracle, to 21 SUBSTTTUTE SHEET (RULE 26) septate junctions (see Fig. 4). Chromosomal Mapping Segments of chromosomal sequence, identical to BlgR cDNA, were retrieved from the public non-redundant database. Results required minor manual modification due to dual designation of isolated bases at the end of some exon boundaries. The correct designation was based on 5' and 3' splice-site consensus sequences. Using the public database, 10 exons for BlgR (see Table 1 below) were identified. The exon/intron structure was determined from Ace No. AL035403 which was derived from human chromosome lq21.2-22. All intron/exon boundaries conform to the GT/AG rule. Most pertinent, the structure of the BlgR gene shows that exclusion of exon 2 during splicing would result in the 34 amino acid deletion described here. Table 1. Intron/Exon Boundaries of human BlgR 3' splice Exon No. Exon (bp) 5'splice Intron (bp) nnnnnn/(N) 1 >224 AGGACG/gtgagt 17,917 ccccag/GCTACT 2 103 GTCAAG/gtgaga 2,063 ctgcag/ACAGCC 3 141 AGAGAG/gtagta 51 1 ccccag/CCCTTC 4 153 TGCTAG/gtgaga 691 atggag/GAATTC 5 138 TCCACG/gtgagt 308 ctctag/GAGAAC 6 171 TTTTAT/gtatgt 2,323 ccacag/ACACAC 7 91 TCCAGT/gtaaga 435 cgacag/CCCCCA 8 170 TTAATG/gtaagc 2,679 ttccag/ACCCCA 9 126 ACAAAG/gtcaga 926 acacag GAACCT 10 >1,208 NNNNNN/(n) EXAMPLE II: CLONING OF MOUSE BlgR A search of the databases (see Example 1) did not result in identification of any mouse ESTs encoding BlgR. Using the primers described in Example 1 to clone full length human BlgR, a product from mouse brain mRNA (Clontech) was amplified. Sequencing of three independent PCR reactions showed that the isoform with a 34 amino acid deletion in the extracellular domain (see Fig. 5) was isolated. Fig. 6 shows that at the amino acid level the mouse and human sequences display 95% identity and 96% similarity. A notable 22 SUBSTTTUTE SHEET (RULE 26) difference is the deletion of a serine and leucine, in the signal sequence. Further, mouse BlgR does not possess the additional glutamic acid residue in the ALAD_EGEY sequence in the extracellular domain. EXAMPLE III: EXPRESSION PATTERN OF BlgR Tissue expression of BlgR was examined on a normalized human Multiple Tissue Northern blot (Clontech) with an [α- 32 P]dCTP labeled probe derived from the extracellular domain. The results show that BlgR is expressed as two transcripts of approximately 2.6 and 3.8 kb (see Fig. 7). The blot was probed at high stringency and thus these two species likely represent alternatively spliced products. Fig.7 shows that human BlgR is abundantly expressed in the brain with extremely low levels in the placenta. This has been independently confirmed in this latter tissue using PCR (see Table 2, below). To further localize BlgR, the expression in various regions of the brain (see Fig. 8) was examined. Clearly, the cerebellum expresses this gene to a higher level than other areas although expression is noted throughout. In the cerebellum an additional minor transcript of approximately 6.5 kb is apparent. The mRNA is barely detectable in the medulla and spinal cord. In all cases the 3.8 kb transcript is expressed to a higher level than the 2.6 kb species. The EST database confirms that BlgR is a brain specific transcript that is expressed in both infant and adult brain (see Table 2, below). All EST sequence data was derived from brain cDNA libraries. Table 2 Expression Characteristics of Human BlgR EXAMPLE IV: EXPRESSION OF EXTRACELLULAR DOMAIN IN INSECT CELLS AND POLYCLONAL ANTIBODY GENERATION. Oligonucleotides were designed to amplify the extracellular domain of human BlgR from the full-length clone. Sense 5'- CCGGATATCGCG 77GGGOCCCC A -3'(SEQ ID NO:9) and antisense 5 '-GATGGTACCGTGGTAGGTGCTGGAGG A- ' (SEQ ID NO: 10) oligonucleotides with EcoRV and Kpnl restriction sites (underlined) respectively, were used to enable subcloning into a Sacl/Kpnl restricted pFastBacl (Life Technologies, GIBCO BRL, Grand Island, NY ) vector containing the constant region of mouse IgG-2a (Fc; Cunningham SA, Tran TM, Arrate MP, Brock TA, (1999) J. Biol. Chem. 274: 18421-7). This vector drives protein expression from the polyhedrin promotor. Secreted recombinant BIgR-Fc was purified from the media of infected Sf21 cells using Hi-Trap protein A columns (Amersham Pharmacia Biotech., Piscataway, NJ). The Fc- fusion was removed following thrombin digestion. A female New Zealand White rabbit (12- weeks old, Myrtle's Rabbitry, Thompson Station, TN) was immunized with 500μg of BlgR emulsified with an equal volume of Freund's Adjuvant. Six weeks thereafter, the animal was boosted with another 500μg BlgR emulsified with Incomplete Freund's Adjuvant. Serum was collected 14 days following the boost. To test the specificity of the antibody, the BlgR cDNA was subcloned into the pcDNA6 vector and lOμg transfected into Cos cells using FuGENE 6 (Roche). Three days following transfection the cells were lyzed in Tris buffered saline (pH 7.5) / 1% Triton X-100 with the inclusion of Protease Inhibitor Cocktail Set III (Calbiochem, La Jolla, CA). Figure 9 shows that the BlgR polyclonal serum detects a protein species of 46kDa in transfected Cos cells. The antibody does not cross-react with mock-transfected Cos cells. EXAMPLE V: EXPRESSION OF THE FULL LENGTH CLONE IN MAMMALIAN CELLS AND SUBCELLULAR LOCALIZATION. The full-length clone of BlgR, in the pcDNA6 vector, was modified at its C-terminus by PCR mutagenesis to incoφorate a V5-epitope Tag for additional detection puφoses. 24 SUBSTTTUTE SHEET (RULE 26) Some lOμg of pcDNA6-BIgR was transfected into CHO cells using FuGENE 6 (Roche). Three days following transfection, cells were split and maintained in 10 ug/ml Blasticidin for clone selection. CHO-K1, control or expressing BlgR, grown on glass slides to confluence, were fixed with 1% paraformaldehyde and stained with 1 : 10 times dilution of either preimmunc or anti- BIgR rabbit polyclonal serum. GAR-FITC at 1 : 100X was used as secondary. Fluorescence was viewed using a NoranTM Confocal laser-scanning microscope (Noran Instruments, Middleton, WI) equipped with argon laser and appropriate optics and filter module for FITC detection. Digital images were obtained at x400 optical magnification. Figure 10 shows that BlgR partitions to sites of cell-cell contact in addition to distributing to the plasma membrane of CHO cells. This pattern of localization provides evidence towards BlgR operating through homotypic interactions. As previously mentioned, the intercellular distribution is a characteristic of the JAM family and thus this feature is maintained. EXAMPLE VI: ADHESION OF BigR-Fc TO LEUKOCYTE CELL LINES. In vitro adhesion assays were performed in 96 well plates essentially as described in Todderud, G., J. Leukoc. Biol, 52:85 (1992), herein incoφorated by reference. Briefly, 50 μl of goat anti-mouse IgG2a was coated at 5 μg/ml in PBS and used to capture 4.8 pmoles of BIgR-Fc or mIgG2a (control). Various leukocyte cell lines, i.e., T lymphocytes, HSB, TK-1; B-iymphocytes, RAMOS; monocytic cells, HL60 and the erythroleukemic, K562 lines were labeled with calcein (Molecular Probes Inc., Eugene, OR) at 50 g/ml for 25 minutes at 37 °C with 250,000 cells/well in binding buffer that consisted of Tris buffered saline with and without the addition of ImM MnCl 2 . Wells were washed 3X, lysed with 50 mM Tris (pH 7.5), 5 mM EDTA, 1% NP40, and fluorescence read in a Cytofluor with excitation at 485/20 nm and emission at 530/25 nm. Specific binding was calculated as fluorescence with BIgR- Fc minus fluorescence with mIgG2a. 25 SUBSTTTUTE SHEET (RULE 26) Fig. 1 1 shows that BIgR-Fc is able to capture the RAMOS B-lymphocyte cell line but not the T-lymphocyte, monocyte or crythroleukemic cells tested. The interaction of BlgR with these cells was dependent upon the addition of manganese in the buffer. These results support the hypothesis that BlgR is an adhesion protein and may operate by engaging with an integrin counter-receptor. BlgR displays similarities to proteins of known function in both its intracellular and extracellular domain. However, BlgR aligns most closely throughout its length with IgSF4 (Ace. No. NP_055148) a putative adhesion protein of unknown function and ubiquitous expression localized on chromosome 1 lq23.2 (Gomyo et al., Genomics 62: 139-146, 1999). Another member of this family would appear to be Ace. No. AAC32740: a hypothetical gene identified on chromosome 19ql3.2. The percent identity for IGSF4 and AAC32740 is 38.5% and 35% respectively. Thus these three proteins likely form a new family. The function of BlgR can be extrapolated from its brain specific expression and its homologies to other proteins of known function in conjunction with its capacity to partition to inter-cellular membranes (see Figures 4, 7, 8, 10). BlgR shows 45% similarity with the poliovirus receptor (PVR) from β-sheet F in the first Ig fold through to the end of the transmembrane domain. The cellular function of PVR is unknown. However, the binding site for poliovirus appears to be contained within Ig-fold 1 , and thus it is not likely that BlgR shares this binding capacity. Other members of the PVR family are the poliovirus-related receptors (PRR), otherwise known as nectin-1, nectin-2 and nectin-3. Nectin-1 and nectin-2 serve as α-heφesvirus entry mediators. The nectin family are calcium independent homophilic adhesion molecules. They display ct-f-homo-dimerization in addition to trans- homointeraction and tra«_.-heterointeraction. For nectin-2, Ig-fold 1 is crucial for adhesion of nectin-2 between cells, but not for cis homo-dimerization. Nectin-1 and nectin-2 show 43% and 41% similarity with BlgR over amino acid overlaps of 237 and 231 : this region corresponds mainly to Ig-folds 2 and 3. Nectin-3 shows 43% similarity over a 280 amino acid overlap encompassing all three Ig folds. Most interestingly, the nectin family localize to adherens junctions. Thus, the inventors believe that BlgR is capable of homotypic 26 SUBSTTTUTE SHEET (RULE 26) interactions, particularly in trans as evidenced by the cellular expression of BlgR in CHO cells. Further, the inventors believe that heterotypic interactions for BlgR cither between closely related homologues or other adhesive cell surface proteins occurred (see Figure 1 1). BlgR may even serve as a receptor for viral entry. In the brain, Ig cell adhesion molecules play roles in growth cone guidance, neurite outgrowth and synaptogenesis. BlgR may participate in any of these processes. As discussed in Example 1, the short intracellular domain of BlgR shares high homology with glycophorin C, drosophila neurexin IV and Caspr2. Homology is highest between the novel putative adhesion protein IgSF4 (see Table 3, below). These proteins all conserve a juxtamembrane binding site for members of the protein 4.1 family and all possess a PDZ binding motif at the extreme C-terminus (with the exception of Caspr). A direct interaction between caspr and protein 4.1 in the brain has been demonstrated. By analogy, a complex between BlgR intracellular domain and protein 4.1 is predicted. Most recently, brain specific expression of novel members of the protein 4.1 family has been documented. It has been suggested that band 4.1 in the brain is involved in the formation and maintenance of the synapse as a membrane skeletal component at presynaptic terminals in the cerebellum. It is conceivable that BlgR may be a component of such a complex. Further, mice lacking brain 4.1 have deficits in movement, coordination, balance and learning. Thus BlgR may impinge on these pathways. Band 4.1 belongs to a growing superfamily of proteins that possess a FERM domain (NF2/ERM/4.1). A conserved property of the domain is its capacity to bind to the membrane-proximal region of the C-terminal cytoplasmic tail of proteins with a single transmembrane segment. It functions to connect cell surface transmembrane proteins to cytoskeletal molecules. BlgR may link to any of these superfamily members. Table 3 Proteins containing PDZ domains are predominantly localized to the plasma membrane and are recruited to specialized sites of cell-cell contact. PDZ domains are found in diverse membrane-associated proteins including members of the membrane-associated guanylate kinase (MAGUK) family, several protein phosphatases and kinases, neuronal nitric oxide synthase, and several dystrophin-associated proteins. BlgR may interact with PDZ domains of proteins in any of these categories. Members of the MAGUK protein family use multiple domains to cluster ion channels, receptors, adhesion molecules and cytosolic signaling proteins at synapses and cellular junctions. They play a fundamental organization role at both the pre- and postsynaptic plasma membranes. Thus, it is possible that BlgR is localized to these structures and functions, either directly or indirectly, in the efficient transmission of signals from the presynaptic terminal. In support of a PDZ domain interaction for BlgR is the established interactions of Neurexin IV with Discs Lost (DLT), mutations in which lead to aberrant localization of Neurexin IV and concomitant loss of epithelial cell polarity. Another example is provided for by glycophorin C interactions with p55 in the ternary complex with protein 4.1. Whilst neurexins are found in presynaptic sites and mediate interactions between neurons, Caspr2 localizes to juxtaparanodal regions of myelinated nerves and is thought to mediate neuron-glia interactions. Therefore, it is highly likely that BlgR is recruited to these sites and participates in the axo-glial intercellular junction. Once again, the intercellular localization of BlgR when expressed in CHO cells supports this possibility. It has been 28 SUBSTTTUTE SHEET (RULE 26) proposed that the axo-glial junctions function in the establishment and maintenance of axolemmal protein domains of the nodal and paranodal regions. In fact, Caspr2 is found to associate with K+ channels in this region, an interaction dependent upon it C-tcrminal PDZ binding domain motif. It may be hypothesized that BlgR plays a similar role in channel clustering. Finally, if BlgR were found to be transcribed in endothelial cells, the inventors believe that it would play a role in maintenance of the blood brain barrier, similar to the function of drosophila neurexin IV in septate junctions. It may also participate in inflammatory reactions of the brain such as occur during stroke and multiple sclerosis. The present invention is illustrated by way of the foregoing description and examples. The foregoing description is intended as a non-limiting illustration, since many variations will become apparent to those skilled in the art in view thereof. It is intended that all such variations within the scope and spirit of the appended claims be embraced thereby. Changes can be made to the composition, operation and arrangement of the method of the present invention described herein without departing from the concept and scope of the invention as defined in the following claims.

Claims: WHAT IS CLAIMED IS: 1. An isolated and purified polynucleotide comprising SEQ ID NO: 1. 2. An isolated and purified polynucleotide comprising SEQ ID NO:3. 3. An isolated and purified polypeptide comprising an amino acid sequence of SEQ ID NO:2 or fragment thereof. 4. An isolated and purified polypeptide comprising an amino acid sequence of SEQ ID NO: 4 or fragment thereof. 5. A recombinant vector comprising a polynucleotide having a nucleotide sequence of SEQ ID NO: l or SEQ ID NO:3 or a fragment thereof, said polynucleotide being operatively linked to a promoter that controls expression of said polynucleotide sequence and a termination segment. 6. The vector of claim 5 wherein the promoter is a LTR, SV40, E. coli, lac, tφ, or phage lambda P L promoter. 7. A host cell comprising the recombinant vector of claim 5. 8. The host cell of claim 7 wherein the host cell is a bacterial cell, an animal cell or a plant cell. 9. A transgenic mammal comprising a null mutation of the nucleotide sequence of SEQ ID NO: 1 or SEQ ID NO:3. 10. A transgenic mammal overexpressing the nucleotide sequence of SEQ ID NO: 1 or SEQ ID NO:3. 30 SUBSTTTUTE SHEET (RULE 26) 1. An antibody binding to the polypeptide of claims 3 or 4. 31 SUBSTTTUTE SHEET (RULE 26)

Inventors: BARROS MARIA PIA TRINDAD ARRAT, CUNNINGHAM SONIA A, TRAN TUAN MINH, BARROS, MARIA, PIA, TRINDAD, ARRATE, CUNNINGHAM, SONIA, A., TRAN, TUAN, MINH
Applicants: TEXAS BIOTECHNOLOGY CORP, TEXAS BIOTECHNOLOGY CORPORATION
Application year: 2000
Main classification: C07K14
IPC codes: A01K67, C07K14, C07K16, C12N1, C12N5, C12N15