Mammalian pro-apoptotic bok genes and their uses

Abstract

Nucleic acid compositions encoding a pro-apoptotic protein, Bok (Bcl-2-related ovarian killer) are identified. Bok has conserved Bcl-2 homology domains 1, 2 and 3 and a C-terminal transmembrane region present in other Bcl-2 related proteins, but lacks the BH4 domain found only in anti-apoptotic Bcl-2 proteins. Over-expression of Bok induces apoptosis. Cell killing induced by Bok is suppressed by co-expression with selective anti-apoptotic Bcl-2 proteins. Bok is highly expressed in the ovary, testis and uterus, particularly in granulosa cells, the cell type that undergoes apoptosis during follicle atresia. Identification of Bok as a new pro-apoptotic protein with wide tissue distribution and hetero-dimerization properties facilitates elucidation of apoptosis mechanisms in reproductive and other tissues, and provides a means for manipulating apoptosis.

What is claimed is:  
     
         1 . An isolated nucleic acid encoding a mammalian Bok protein.  
     
     
         2 . The isolated nucleic acid according to  claim 1 , wherein said Bok protein comprises the amino acid sequence as set forth in SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6 or SEQ ID NO:8.  
     
     
         3 . The isolated nucleic acid according to  claim 1 , wherein said Bok protein is a BH3 i  variant protein.  
     
     
         4 . An isolated nucleic acid comprising at least 18 contiguous nucleotides of the sequence of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5 or SEQ ID NO:7.  
     
     
         5 . An isolated nucleic acid that hybridizes under stringent conditions to the nucleic acid sequence of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5 or SEQ ID NO:7.  
     
     
         6 . An isolated nucleic acid encoding a BH3i variant of a pro-apoptotic Bok related protein.  
     
     
         7 . The isolated nucleic acid of  claim 6 , wherein said pro-apoptotic Bok related protein is Bak or Bax.  
     
     
         8 . An expression cassette comprising a transcriptional initiation region functional in an expression host and operably linked to a nucleic acid having a sequence of the isolated nucleic acid according to  claim 1 .  
     
     
         9 . A cell comprising an expression cassette according to  claim 8  as part of an extrachromosomal element or integrated into the genome of a host cell as a result of introduction of said expression cassette into said host cell, and the cellular progeny of said host cell.  
     
     
         10 . A method for producing pro-apoptotic protein, said method comprising: 
 growing a cell according to  claim 9 , whereby said protein is expressed; and    isolating said protein free of other proteins.    
     
     
         11 . A purified polypeptide composition comprising at least 50 weight % of the protein present as a Bok protein or a fragment thereof.  
     
     
         12 . A purified polypeptide composition comprising at least 50 weight % of the protein present as a BH3 i  variant of a pro-apoptotic Bok related protein.  
     
     
         13 . A monoclonal antibody binding specifically to a Bok protein.  
     
     
         14 . A non-human transgenic animal model for Bok gene function wherein said transgenic animal comprises an introduced alteration in a Bok gene.  
     
     
         15 . A method of inducing apoptosis in a susceptible cell population, the method comprising: 
 upregulating expression of Bok or a BH3 i  variant of a pro-apoptotic Bok related protein in said cell population; wherein apoptosis is induced.    
     
     
         16 . The method of  claim 15 , wherein said susceptible cell population comprises reproductive tissue.  
     
     
         17 . The method of  claim 15 , wherein said upregulating step comprises induction of expression of an endogenous Bok gene.  
     
     
         18 . The method of  claim 15 , wherein said upregulating step comprises introduction and expression of an exogenous Bok coding sequence.  
     
     
         19 . The method of  claim 15 , wherein said upregulating step comprises introduction and expression of an exogenous coding sequence for a BH3 i  variant of a pro-apoptotic Bok related protein.