Identification of the preferential ubiquitination site and ubiquitin-dependent degradation signal of Rpn4.
Lysine selection is a long-standing problem in protein ubiquitination catalyzed by the RING ubiquitin ligases. It is well known that many substrates carry multiple lysines that can be ubiquitinated. However, it has seldom been addressed whether one lysine is preferred for ubiquitin conjugation when all other lysines exist. Here we studied the mechanism underlying ubiquitin-dependent degradation of Rpn4, a transcription activator of the Saccharomyces cerevisiae proteasome genes. We found that the ubiquitin-dependent degradation of Rpn4 can be mediated by six different lysines. Interestingly, we showed through in vivo and in vitro assays that lysine 187 is selected for ubiquitination when all other lysines are available. To the best of our knowledge, this is the first demonstration of a preferential ubiquitination site chosen from a group of lysines susceptible for ubiquitination. We further demonstrated that lysine 187 and a proximal acidic domain constitute a portable degradation signal. The implications of our data are discussed.
Barbara Ann Karmanos Cancer Institute, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.
Donghong
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Ju
Youming
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D000595:Amino Acid Sequence; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000378:metabolism / Q000502:physiology*; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000737:chemistry; D047468:Immunoprecipitation; D008239:Lysine / Q000737:chemistry; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism / Q000502:physiology*; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism / Q000502:physiology*; D014158:Transcription, Genetic; D025801:Ubiquitin / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*
D004268:DNA-Binding Proteins; D000939:Epitopes; C119430:RPN4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D025801:Ubiquitin; D008239:Lysine