Negative regulation of transcription factors by Srb10 cyclin-dependent kinase

[摘要] The ubiquitin-dependent proteolytic pathway plays an important role in a broadarray of cellular processes, inducting cell cycle control and transcription. Biochemicalanalysis of the ubiquitination of Sic1, the B-type cyclin-dependent kinase (CDK)inhibitor in budding yeast helped to define a ubiquitin ligase complex named SCF^cdc4 (forSkp1, Cdc53/cullin, F-box protein). We found that besides Sic1, the CDK inhibitor Far1and the replication initiation protein Cdc6 are also substrates of SCF^cdc4 in vitro. Acommon feature in the ubiquitination of the cell cycle SCF^cdc4 substrates is that they mustbe phosphorylated by the major cell cycle CDK, Cdc28. Gcn4, a transcription activatorinvolved in the general control of amino acid biosynthesis, is rapidly degraded in anSCF^cdc4-dependent manner in vivo. We have focused on this substrate to investigate thegenerality of the SCF^cdc4 pathway. Through biochemical fractionations, we found that theSrb10 CDK phosphorylates Gcn4 and thereby marks it for recognition by SCF^cdc4ubiquitin ligase. Srb10 is a physiological regulator of Gcn4 stability because bothphosphorylation and turnover of Gcn4 are diminished in srb10 mutants. Furthermore, wefound that at least two different CDKs, Pho85 and Srb10, conspire to promote the rapiddegradation of Gcn4 in vivo. The multistress response transcriptional regulator Msn2 isalso a substrate for Srb10 and is hyperphosphorylated in an Srb10-dependent mannerupon heat stress-induced translocation into the nucleus. Whereas Msn2 is cytoplasmic inresting wild type cells, its nuclear exclusion is partially compromised in srb10 mutantcells. Srb10 has been shown to repress a subset of genes in vivo, and has been proposedto inhibit transcription via phosphorylation of the C-terminal domain of RNA polymeraseII. Our results suggest a general theme that Srb10 represses the transcription of specificgenes by directly antagonizing the transcriptional activators.

[发布机构] University:California Institute of Technology;Department:Biology

[关键词] Biology