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Dephosphorylation of cyclin-dependent kinases by type 2C protein phosphatases
1Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, Connecticut 06520-8024, USA.
Abstract:
Activating phosphorylation of cyclin-dependent protein kinases (CDKs) is necessary for their kinase activity and cell cycle progression. This phosphorylation is carried out by the Cdk-activating kinase (CAK); in contrast, little is known about the corresponding protein phosphatase. We show that type 2C protein phosphatases (PP2Cs) are responsible for this dephosphorylation of Cdc28p, the major budding yeast CDK. Two yeast PP2Cs, Ptc2p and Ptc3p, display Cdc28p phosphatase activity in vitro and in vivo, and account for approximately 90% of Cdc28p phosphatase activity in yeast extracts. Overexpression of PTC2 or PTC3 results in synthetic lethality in strains temperature-sensitive for yeast CAK1, and disruptions of PTC2 and PTC3 suppress the growth defect of a cak1 mutant. Furthermore, PP2C-like enzymes are the predominant phosphatases toward human Cdk2 in HeLa cell extracts, indicating that the substrate specificity of PP2Cs toward CDKs is evolutionarily conserved.
Insights
Type 2C protein phosphatases (PP2Cs) dephosphorylate cyclin-dependent protein kinases (CDKs), regulating cell cycle progression. This phosphatase activity is conserved across yeast and human cells, impacting CDK regulation.
Area of Science:
- Cell Biology
- Biochemistry
- Molecular Biology
Background:
- Cyclin-dependent protein kinases (CDKs) require activating phosphorylation for cell cycle progression.
- Cdk-activating kinase (CAK) performs this phosphorylation, but the responsible phosphatase remains largely unknown.
Purpose of the Study:
- To identify the protein phosphatase responsible for dephosphorylating budding yeast CDKs.
- To investigate the role of PP2Cs in CDK regulation and conservation in human cells.
Main Methods:
- In vitro and in vivo assays to measure Cdc28p phosphatase activity.
- Genetic analysis using yeast strains with altered PTC2 and PTC3 gene expression.
- Analysis of human cell extracts for CDK phosphatase activity.
Main Results:
- Two yeast PP2Cs, Ptc2p and Ptc3p, were identified as the primary enzymes dephosphorylating Cdc28p.
- These phosphatases account for approximately 90% of Cdc28p phosphatase activity.
- PP2C-like enzymes also dephosphorylate human Cdk2, indicating conserved substrate specificity.
Conclusions:
- Type 2C protein phosphatases (PP2Cs) are the key phosphatases for budding yeast CDK Cdc28p.
- PP2C regulation of CDKs is evolutionarily conserved from yeast to humans.