title: The anaphase-promoting complex works together with the SCF complex for proteolysis of the S-phase cyclin Clb6 during the transition from G1 to S phase.
abstract: In Saccharomyces cerevisiae, the S-phase cyclin Clb6 is expressed shortly before the G1/S transition. It has been shown that in S phase the SCF(Cdc4) ubiquitin ligase controls Clb6 proteolysis, which requires cyclin-dependent kinases activity. A Clb6-3A mutant, bearing non-phosphorylatable mutations at S6A, T39A, and S147A, was observed to be hyperstabilized in S-phase but was unstable in mitosis. In this study, we found that the APC(Cdh1) form of the Anaphase-Promoting Complex (APC) was required for Clb6 proteolysis in both early and late G1. An in vitro ubiquitination assay confirmed that Clb6 is a substrate for APC(Cdh1). A KEN box and a destruction box in the Clb6N-terminus were identified. Mutations in the KEN box (mkb) and/or the destruction box (mdb) enhanced Clb6 stability in G1. Expression of Clb6mkd, bearing both mutations in the mkb and mdb, allowed cells to bypass the late G1 arrest caused by cdc4-1. This bypass phenotype was observed to depend upon CDK phosphorylation at residues S6, T39 and S147. Compared to Clb6, overexpression of Clb6ST, bearing all five mutations of S6A, T39A, S147A, mkb and mdb in combination, had a greater effect on promoting expression of Clb2 and S-phase entry, caused a greater G2 delay and a greater defect in cell division. Swe1 was also required for bud emergence when Clb6ST was overexpressed. Our observations suggest that both APC(Cdh1) and SCF(Cdc4)-dependent proteolysis of Clb6 at the G1/S border are crucial for multiple cell cycle regulated events including proper expression of Clb2, the G1/S and G2/M cell cycle transitions and for proper completion of cell division at mitotic exit.
authors.affiliation: Master Program in Microbiology and Immunology, Tzu-Chi University, Hualien, Taiwan.
authors.forename: Shiao-Yii
authors.initials: SY
authors.lastname: Wu
authors.affiliation: Department of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan.
authors.forename: Vivian Jen-Wei
authors.initials: VJ
authors.lastname: Kuan
authors.affiliation: Institute of Medical Sciences, Tzu-Chi University, Hualien, Taiwan.
authors.forename: Yao-Wei
authors.initials: YW
authors.lastname: Tzeng
authors.affiliation: Department of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan; Department of Colorectal Surgery, Chang Gung Memorial Hospital, Taoyuan, Taiwan. Electronic address: schuyler@mail.cgu.edu.tw.
authors.forename: Scott C
authors.initials: SC
authors.lastname: Schuyler
authors.affiliation: Institute of Biomedical Sciences, Mackay Medical College, New Taipei City, Taiwan. Electronic address: yljylk@mmc.edu.tw.
authors.forename: Yue-Li
authors.initials: YL
authors.lastname: Juang
mesh_terms: D064173:Anaphase-Promoting Complex-Cyclosome / Q000235:genetics; D064200:Cdh1 Proteins / Q000235:genetics*; D002453:Cell Cycle / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics*; D019926:Cyclin B / Q000096:biosynthesis / Q000235:genetics*; D044783:F-Box Proteins / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal; D008938:Mitosis; D009154:Mutation; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D059748:Proteolysis; D016196:S Phase / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000235:genetics*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics*
chemical_list: C080443:CDC4 protein, S cerevisiae; C577583:CDH1 protein, S cerevisiae; C093749:CLB2 protein, S cerevisiae; C095227:CLB6 protein, S cerevisiae; D064200:Cdh1 Proteins; D018797:Cell Cycle Proteins; D019926:Cyclin B; D044783:F-Box Proteins; D029701:Saccharomyces cerevisiae Proteins; D064173:Anaphase-Promoting Complex-Cyclosome; D044767:Ubiquitin-Protein Ligases; C099685:SWE1 protein, S cerevisiae; D011505:Protein-Tyrosine Kinases
keywords: Anaphase-promoting complex/cyclosome; Cdh1; Cyclin-dependent kinase; G1-S phase transition