IDENTIFICATION OF A NOVEL PROTEIN (VBP-1) BINDING TO THE VONHIPPEL-LINDAU (VHL) TUMOR-SUPPRESSOR GENE-PRODUCT

The tumor suppressor VHL gene product (pVHL), recently reported to bin d to elongins B and C, is thought to regulate transcription elongation . To establish whether the VHL gene may have other functions, we here searched for additional cellular protein(s) that might bind to pVHL us ing a two-hybrid system and identified seven independent clones, inclu ding elongin C, but not elongin B. Three clones (unknown, imidopeptida se, and unknown) presumably bind to the N-terminal nonconserved region , whereas the four other clones [elongin C, the HIV tat-binding protei n, the actin-binding protein Filamin (ABP280), and the HIBBJ46 (named VBP-1)] were found to bind to the wild-type pVHL but not to a C-termin al 156-amino acid deletion mutant, interestingly, the HIV tat-binding protein and Filamin could bind to C-terminal 26-amino acid deleted pVH L, but elongin C and VBP-1 failed to do so. Thus, elongin C and VBP-1 require the C-terminal end of pVHL for binding, It was also establishe d that epitope-tagged pVHL strongly forms complexes with VBP-1 in vivo using immunoprecipitation Western blotting analysis. VBP-1 was widely expressed in various cell lines tested, in which VHL mRNA can be dete cted, When the VBP-1 protein was solely expressed, it Located to the c ytoplasm and did not localize to the nucleus. However, when coexpresse d with VHL, it can translocate to the nucleus. These results indicate that VBP-1 can form a complex with VHL protein in vivo and hence VHL a ffects the intracellular localization of VBP-1 protein.