Neutrokine-alpha and Neutrokine-alpha splice variant

Abstract

The present invention relates to a novel Neutrokine-a, and a splice variant thereof designated Neutrokine-aSV, polynucleotides and polypeptides which are members of the TNF family. In particular, isolated nucleic acid molecules are provided encoding the human Neutrokine-a and/or Neutrokine-aSV polypeptides, including soluble forms of the extracellular domain. Neutrokine-a and/or Neutrokine-aSV polypeptides are also provided as are vectors, host cells and recombinant methods for producing the same. The invention further relates to screening methods for identifying agonists and antagonists of Neutrokine-a and/or Neutrokine-aSV activity. Also provided are diagnostic methods for detecting immune system-related disorders and therapeutic methods for treating immune system-related disorders.

What is claimed is:  
     
         1 . An isolated nucleic acid molecule comprising a polynucleotide having a nucleotide sequence at least 95% identical to a sequence selected from the group consisting of: 
 (a) a nucleotide sequence encoding the Neutrokine-a polypeptide having the complete amino acid sequence in FIGS. 1A and 1B (SEQ ID NO:2);    (b) a nucleotide sequence encoding the Neutrokine-a polypeptide having the complete amino acid sequence encoded by the cDNA clone contained in the deposit having ATCC accession number 97768;    (c) a nucleotide sequence encoding the Neutrokine-a polypeptide extracellular domain;    (d) a nucleotide sequence encoding the Neutrokine-a polypeptide transmembrane domain;    (e) a nucleotide sequence encoding the Neutrokine-a polypeptide intracellular domain;    (f) a nucleotide sequence encoding a soluble Neutrokine-a polypeptide comprising the extracellular and intracellular domains but lacking the transmembrane domain; and    (g) a nucleotide sequence complementary to any of the nucleotide sequences in (a), (b), (c), (d), (e) or (f) above.    
     
     
         2 . The nucleic acid molecule of  claim 1  wherein said polynucleotide has the complete nucleotide sequence in FIGS. 1A and 1B (SEQ ID NO:1).  
     
     
         3 . The nucleic acid molecule of  claim 1  wherein said polynucleotide has the nucleotide sequence in FIGS. 1A and 1B (SEQ ID NO:1) encoding the Neutrokine-a polypeptide having the complete amino acid sequence in FIGS. 1A and 1B (SEQ ID NO:2).  
     
     
         4 . The nucleic acid molecule of  claim 1  wherein said polynucleotide has the nucleotide sequence encoding a soluble Neutrokine-a polypeptide comprising the extracellular domain shown in FIGS. 1A and 1B (SEQ ID NO:2).  
     
     
         5 . An isolated nucleic acid molecule comprising a polynucleotide having a nucleotide sequence at least 95% identical to a sequence selected from the group consisting of: 
 (a) a nucleotide sequence encoding a polypeptide having the amino acid sequence consisting of residues n-285 of SEQ ID NO:2, where n is an integer in the range of 2-190    (b) a nucleotide sequence encoding a polypeptide having the amino acid sequence consisting of residues 1-m of SEQ ID NO:2, where m is an integer in the range of 274-284;    (c) a nucleotide sequence encoding a polypeptide having the amino acid sequence consisting of residues n-m of SEQ ID NO:2, where n and m are integers as defined respectively in (a) and (b) above; and    (d) a nucleotide sequence encoding a polypeptide consisting of a portion of the complete Neutrokine-a amino acid sequence encoded by the cDNA clone contained in the deposit having ATCC accession number 97768, wherein said portion excludes from 1 to 190 amino acids from the amino terminus and from 1 to 11 amino acids from the C-terminus of said complete amino acid sequence.    
     
     
         6 . The nucleic acid molecule of  claim 1  wherein said polynucleotide has the complete nucleotide sequence of the cDNA clone contained in the deposit having ATCC accession number 97768.  
     
     
         7 . The nucleic acid molecule of  claim 1  wherein said polynucleotide has the nucleotide sequence encoding the Neutrokine-a polypeptide having the complete amino acid sequence encoded by the cDNA clone contained in the deposit having ATCC accession number 97768.  
     
     
         8 . The nucleic acid molecule of  claim 1  wherein said polynucleotide has the nucleotide sequence encoding a soluble Neutrokine-a polypeptide comprising the extracellular domain encoded by the cDNA clone contained in the deposit having ATCC accession number 97768.  
     
     
         9 . An isolated nucleic acid molecule comprising a polynucleotide which hybridizes under stringent hybridization conditions to a polynucleotide having a nucleotide sequence identical to a nucleotide sequence in (a), (b), (c), (d), (e) or (f) of  claim 1  wherein said polynucleotide which hybridizes does not hybridize under stringent hybridization conditions to a polynucleotide having a nucleotide sequence consisting of only A residues or of only T residues.  
     
     
         10 . An isolated nucleic acid molecule comprising a polynucleotide which encodes the amino acid sequence of an epitope-bearing portion of a Neutrokine-a polypeptide having an amino acid sequence in (a), (b), (c), (d), (e) or (f) of  claim 1 .  
     
     
         11 . The isolated nucleic acid molecule of  claim 10 , which encodes an epitope-bearing portion of a Neutrokine-a polypeptide selected from the group consisting of: a polypeptide comprising amino acid residues from about Phe-115 to about Leu-147 (SEQ ID NO:2); a polypeptide comprising amino acid residues from about Ile-150 to about Tyr-163 (SEQ ID NO:2); a polypeptide comprising amino acid residues from about Ser-171 to about Phe-194 (SEQ ID NO:2); a polypeptide comprising amino acid residues from about Glu-223 to about Tyr-247 (SEQ ID NO:2); and a polypeptide comprising amino acid residues from about Ser-271 to about Phe-278 (SEQ ID NO:2).  
     
     
         12 . A method for making a recombinant vector comprising inserting an isolated nucleic acid molecule of  claim 1  into a vector.  
     
     
         13 . A recombinant vector produced by the method of  claim 12 .  
     
     
         14 . A method of making a recombinant host cell comprising introducing the recombinant vector of  claim 13  into a host cell.  
     
     
         15 . A recombinant host cell produced by the method of  claim 14 .  
     
     
         16 . A recombinant method for producing a Neutrokine-a polypeptide, comprising culturing the recombinant host cell of  claim 15  under conditions such that said polypeptide is expressed and recovering said polypeptide.  
     
     
         17 . An isolated Neutrokine-a polypeptide comprising an amino acid sequence at least 95% identical to a sequence selected from the group consisting of: 
 (a) the amino acid sequence of the Neutrokine-a polypeptide having the complete amino acid sequence in FIGS. 1A and 1B (SEQ ID NO:2);    (b) the amino acid sequence of the Neutrokine-a polypeptide having the complete amino acid sequence encoded by the cDNA clone contained in the deposit having ATCC accession number 97768;    (c) the amino acid sequence of the Neutrokine-a polypeptide extracellular domain;    (d) the amino acid sequence of the Neutrokine-a polypeptide transmembrane domain;    (e) the amino acid sequence of the Neutrokine-a polypeptide intracellular domain;    (f) the amino acid sequence of a soluble Neutrokine-a polypeptide comprising the domain; and    (g) the amino acid sequence of an epitope-bearing portion of any one of the polypeptides of (a), (b), (c), (d), (e) or (f).    
     
     
         18 . An isolated polypeptide of  claim 17  comprising an epitope-bearing portion of the Neutrokine-a protein, wherein said portion is selected from the group consisting of: a polypeptide comprising amino acid residues from about Phe-115 to about Leu-147 (SEQ ID NO:2); a polypeptide comprising amino acid residues from about Ile-150 to about Tyr-163 (SEQ ID NO:2); a polypeptide comprising amino acid residues from about Ser-171 to about Phe-194 (SEQ ID NO:2); a polypeptide comprising amino acid residues from about Glu-223 to about Tyr-247 (SEQ ID NO:2); a polypeptide comprising amino acid residues from about Ser-271 to about Phe-278 (SEQ ID NO:2).  
     
     
         19 . An isolated antibody that binds specifically to a Neutrokine-a polypeptide of  claim 17 .  
     
     
         20 . A pharmaceutical composition comprising a polypeptide of  claim 17  and a pharmaceutically acceptable carrier.  
     
     
         21 . An isolated polynucleotide encoding a modified Neutrokine-a protein, wherein, except for at least one conservative amino acid substitution, said modified peptide has an amino acid sequence that is identical to a member selected from the group consisting of: 
 (a) amino acids 1 to 285 of SEQ ID NO:2;    (b) amino acids 2 to 285 of SEQ ID NO:2;    (c) amino acids 1 to 46 of SEQ ID NO:2;    (c) amino acids 47 to 72 of SEQ ID NO:2; and    (c) amino acids 73 to 286 of SEQ ID NO:2.    
     
     
         22 . A modified Neutrokine-a polypeptide molecule, wherein, except for at least one conservative amino acid substitution, said modified peptide has an amino acid sequence that is identical to a member selected from the group consisting of: 
 (a) amino acids 1 to 285 of SEQ ID NO:2;    (b) amino acids 2 to 285 of SEQ ID NO:2;    (c) amino acids 1 to 46 of SEQ ID NO:2;    (c) amino acids 47 to 72 of SEQ ID NO:2; and    (c) amino acids 73 to 286 of SEQ ID NO:2.    
     
     
         23 . An isolated nucleic acid molecule comprising a polynucleotide having a sequence at least 95% identical to a sequence selected selected from the group consisting of: 
 (a) the nucleotide sequence of SEQ ID NO:7;    (b) the nucleotide sequence of SEQ ID NO:8;    (c) the nucleotide sequence of SEQ ID NO:9;    (d) the nucleotide sequence of a portion of the sequence shown in FIGS. 1A and 1B (SEQ ID NO:1) wherein said portion comprises at least 30 contiguous nucleotides from nucleotide 1 to nucleotide 2442, excluding the sequence from nucleotide 1387 to 1421, the sequence from nucleotide 9 to 382, the sequence from nucleotide 1674 to 1996, the sequence from nucleotide 1401 to 1784, the sequence from nucleotide 900 to 1237, and any fragments located within these sequences; and    (e) a nucleotide sequence complementary to any of the nucleotide sequences in (a), (b), (c) or (d) above.    
     
     
         24 . An isolated nucleic acid molecule comprising a polynucleotide having a nucleotide sequence at least 95% identical to a sequence selected from the group consisting of: 
 (a) a nucleotide sequence encoding the Neutrokine-aSV polypeptide having the complete amino acid sequence in FIGS. 5A and 5B (SEQ ID NO:19);    (b) a nucleotide sequence encoding the Neutrokine-aSV polypeptide having the complete amino acid sequence encoded by the cDNA clone contained in the deposit having ATCC accession number 203518;    (c) a nucleotide sequence encoding the Neutrokine-aSV polypeptide extracellular domain;    (d) a nucleotide sequence encoding the Neutrokine-aSV polypeptide transmembrane domain;    (e) a nucleotide sequence encoding the Neutrokine-aSV polypeptide intracellular domain;    (f) a nucleotide sequence encoding a soluble Neutrokine-aSV polypeptide comprising the extracellular and intracellular domains but lacking the transmembrane domain; and    (g) a nucleotide sequence complementary to any of the nucleotide sequences in (a), (b), (c), (d), (e) or (f) above.