Identification of a novel protein (VBP-1) binding to the von Hippel-Lindau (VHL) tumor suppressor gene product.
The tumor suppressor VHL gene product (pVHL), recently reported to bind 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 using a two-hybrid system and identified seven independent clones, including elongin C, but not elongin B. Three clones (unknown, imidopeptidase, and unknown) presumably bind to the N-terminal nonconserved region, whereas the four other clones [elongin C, the HIV tat-binding protein, 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-terminal 156-amino acid deletion mutant. Interestingly, the HIV tat-binding protein and Filamin could bind to C-terminal 26-amino acid deleted pVHL, 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 established 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 detected. When the VBP-1 protein was solely expressed, it located to the cytoplasm and did not localize to the nucleus. However, when coexpressed 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 affects the intracellular localization of VBP-1 protein.
Tsuchiya
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Department of Experimental Pathology, Cancer Institute, Tokyo, Japan.
Iseda
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T
Hino
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D000595:Amino Acid Sequence; D000818:Animals; D030161:Avian Proteins; D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors; D015153:Blotting, Western; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D003598:Cytoskeletal Proteins; D016147:Genes, Tumor Suppressor; D006367:HeLa Cells; D006801:Humans; D006822:Hybrid Cells; D008025:Ligases; D018832:Molecular Chaperones; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D012334:RNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D014178:Translocation, Genetic; D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein
D030161:Avian Proteins; D050976:Basic-Leucine Zipper Transcription Factors; D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; D018832:Molecular Chaperones; D011506:Proteins; D012334:RNA, Neoplasm; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C108825:VBP1 protein, human; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D008025:Ligases; C497446:VHL protein, human