Abstract
In S. cerevisiae, the G1/S transition requires Cdc4p, Cdc34p, Cdc53p, Skp1p, and the Cln/Cdc28p cyclin-dependent kinase (Cdk). These proteins are thought to promote the proteolytic inactivation of the S-phase Cdk inhibitor Sic1p. We show here that Cdc4p, Cdc53p, and Skp1p assemble into a ubiquitin ligase complex named SCFCdc4p. When mixed together, SCFCdc4p subunits, E1 enzyme, the E2 enzyme Cdc34p, and ubiquitin are sufficient to reconstitute ubiquitination of Cdk-phosphorylated Sic1p. Phosphorylated Sic1p substrate is specifically targeted for ubiquitination by binding to a Cdc4p/Skp1p subcomplex. Taken together, these data illuminate the molecular basis for the G1/S transition in budding yeast and suggest a general mechanism for phosphorylation-targeted ubiquitination in eukaryotes.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / isolation & purification
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Cell Cycle Proteins / metabolism*
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Cullin Proteins*
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Cyclin-Dependent Kinase Inhibitor Proteins
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Enzyme Inhibitors / isolation & purification
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Enzyme Inhibitors / metabolism*
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F-Box Proteins*
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Fungal Proteins / isolation & purification
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Fungal Proteins / metabolism*
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Insecta
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Phosphorylation
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Proto-Oncogene Proteins c-myc / genetics
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Proto-Oncogene Proteins c-myc / metabolism
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Recombinant Proteins / metabolism
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S-Phase Kinase-Associated Proteins
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Saccharomyces cerevisiae
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Saccharomyces cerevisiae Proteins*
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Ubiquitin-Protein Ligases*
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Ubiquitins / metabolism*
Substances
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CDC4 protein, S cerevisiae
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Cdc53 protein, S cerevisiae
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Cell Cycle Proteins
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Cullin Proteins
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Cyclin-Dependent Kinase Inhibitor Proteins
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Enzyme Inhibitors
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F-Box Proteins
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Fungal Proteins
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Proto-Oncogene Proteins c-myc
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Recombinant Proteins
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S-Phase Kinase-Associated Proteins
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SIC1 protein, S cerevisiae
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Saccharomyces cerevisiae Proteins
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Ubiquitins
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Ubiquitin-Protein Ligases