Identification of elongin C sequences required for interaction with the von Hippel-Lindau tumor suppressor protein.
Elongin C is a 112-amino acid protein that is found in mammalian cells as a positive regulatory subunit of heterotrimeric RNA polymerase II elongation factor Elongin (SIII) and as a component of a multiprotein complex containing the von Hippel-Lindau (VHL) tumor suppressor protein. As a subunit of the Elongin complex, Elongin C interacts directly with the transcriptionally active Elongin A subunit and potently induces its elongation activity; in addition, Elongin C interacts with the ubiquitin-like Elongin B subunit, which regulates the interaction of Elongin C with Elongin A. As a component of the VHL complex, Elongin C interacts directly with both Elongin B and the VHL protein. Binding of the VHL protein to Elongin C was found to prevent Elongin C from interacting with and activating Elongin A in vitro, leading to the proposal that one function of the VHL protein may be to regulate RNA polymerase II elongation by negatively regulating the Elongin complex. In this report, we identify Elongin C sequences required for its interaction with the VHL protein. We previously demonstrated that the ability of Elongin C to bind and activate Elongin A is sensitive to mutations in the C-terminal half of Elongin C, as well as to mutations in an N-terminal Elongin C region needed for formation of the Elongin BC complex. Here we show that interaction of Elongin C with the VHL tumor suppressor protein depends strongly on sequences in the C terminus of Elongin C but is independent of the N-terminal Elongin C region required for binding to Elongin B and for binding and activation of Elongin A. Taken together, our results are consistent with the proposal that the VHL protein negatively regulates Elongin C activation of the Elongin complex by sterically blocking the interaction of C-terminal Elongin C sequences with Elongin A. In addition, our finding that only a subset of Elongin C sequences required for its interaction with Elongin A are critical for binding to VHL may offer the opportunity to develop reagents that selectively interfere with Elongin and VHL function.
Takagi
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Program in Molecular and Cell Biology, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma 73104, USA.
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D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D017173:Caenorhabditis elegans; D029742:Caenorhabditis elegans Proteins; D000075065:Elongin; D016147:Genes, Tumor Suppressor; D006801:Humans; D007700:Kinetics; D008025:Ligases; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D016415:Sequence Alignment; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D016324:Sequence Tagged Sites; D014157:Transcription Factors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein
D029742:Caenorhabditis elegans Proteins; C000616397:ELOA protein, human; C000616396:ELOB protein, human; C000616395:ELOC protein, human; C000616404:Eloa protein, rat; C000616402:Elob protein, rat; C000616400:Eloc protein, rat; D000075065:Elongin; D011506:Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C000616405:elb-1 protein, C elegans; C000616412:tceb-3 protein, C elegans; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D008025:Ligases; C497446:VHL protein, human