Phosphorylation of Sic1p by G1 Cdk required for its degradation and entry into S phase.
G1 cyclin-dependent kinase (Cdk)-triggered degradation of the S-phase Cdk inhibitor Sic1p has been implicated in the transition from G1 to S phase in the cell cycle of budding yeast. A multidimensional electrospray mass spectrometry technique was used to map G1 Cdk phosphorylation sites in Sic1p both in vitro and in vivo. A Sic1p mutant lacking three Cdk phosphorylation sites did not serve as a substrate for Cdc34p-dependent ubiquitination in vitro, was stable in vivo, and blocked DNA replication. Moreover, purified phosphoSic1p was ubiquitinated in cyclin-depleted G1 extract, indicating that a primary function of G1 cyclins is to tag Sic1p for destruction. These data suggest a molecular model of how phosphorylation and proteolysis cooperate to bring about the G1/S transition in budding yeast.
Verma
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Division of Biology, Box 156-29, California Institute of Technology, Pasadena, CA 91125, USA.
Annan
R S
RS
Huddleston
M J
MJ
Carr
S A
SA
Reynard
G
G
Deshaies
R J
RJ
D000595:Amino Acid Sequence; D064173:Anaphase-Promoting Complex-Cyclosome; D056746:Cyclin G; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000378:metabolism*; D004261:DNA Replication; D004791:Enzyme Inhibitors / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism*; D016193:G1 Phase; D008025:Ligases / Q000378:metabolism; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010641:Phenotype; D010748:Phosphopeptides / Q000378:metabolism; D010766:Phosphorylation; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016196:S Phase; D029701:Saccharomyces cerevisiae Proteins; D043743:Ubiquitin-Protein Ligase Complexes; D044767:Ubiquitin-Protein Ligases; D014452:Ubiquitins / Q000378:metabolism; D015003:Yeasts / Q000166:cytology* / Q000378:metabolism
D056746:Cyclin G; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; D016213:Cyclins; D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D010748:Phosphopeptides; D011993:Recombinant Fusion Proteins; C086926:SIC1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014452:Ubiquitins; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D044767:Ubiquitin-Protein Ligases; D018844:Cyclin-Dependent Kinases; D008025:Ligases