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Multiple protein kinases influence the redistribution of fission yeast Clp1/Cdc14 phosphatase upon genotoxic stress.

ABSTRACT: The Cdc14 phosphatase family antagonizes Cdk1 phosphorylation and is important for mitotic exit. To access their substrates, Cdc14 phosphatases are released from nucleolar sequestration during mitosis. Clp1/Flp1, the Schizosaccharomyces pombe Cdc14 orthologue, and Cdc14B, a mammalian orthologue, also exit the nucleolus during interphase upon DNA replication stress or damage, respectively, implicating Cdc14 phosphatases in the response to genotoxic insults. However, a mechanistic understanding of Cdc14 phosphatase nucleolar release under these conditions is incomplete. We show here that relocalization of Clp1 during genotoxic stress is governed by complex phosphoregulation. Specifically, the Rad3 checkpoint effector kinases Cds1 and/or Chk1, the cell wall integrity mitogen-activated protein kinase Pmk1, and the cell cycle kinase Cdk1 directly phosphorylate Clp1 to promote genotoxic stress-induced nucleoplasmic accumulation. However, Cds1 and/or Chk1 phosphorylate RxxS sites preferentially upon hydroxyurea treatment, whereas Pmk1 and Cdk1 preferentially phosphorylate Clp1 TP sites upon H(2)O(2) treatment. Abolishing both Clp1 RxxS and TP phosphosites eliminates any genotoxic stress-induced redistribution. Reciprocally, preventing dephosphorylation of Clp1 TP sites shifts the distribution of the enzyme to the nucleoplasm constitutively. This work advances our understanding of pathways influencing Clp1 localization and may provide insight into mechanisms controlling Cdc14B phosphatases in higher eukaryotes.

SUBMITTER: Broadus MR

PROVIDER: S-EPMC3469525 | biostudies-literature | 2012 Oct

REPOSITORIES: biostudies-literature

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Multiple protein kinases influence the redistribution of fission yeast Clp1/Cdc14 phosphatase upon genotoxic stress.

Broadus Matthew R MR Gould Kathleen L KL

Molecular biology of the cell 20120823 20

The Cdc14 phosphatase family antagonizes Cdk1 phosphorylation and is important for mitotic exit. To access their substrates, Cdc14 phosphatases are released from nucleolar sequestration during mitosis. Clp1/Flp1, the Schizosaccharomyces pombe Cdc14 orthologue, and Cdc14B, a mammalian orthologue, also exit the nucleolus during interphase upon DNA replication stress or damage, respectively, implicating Cdc14 phosphatases in the response to genotoxic insults. However, a mechanistic understanding of ...[more]

PMID: 22918952

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Project description:Background Gametes are generated through a specialized cell division called meiosis, in which ploidy is reduced by half because two consecutive rounds of chromosome segregation, meiosis I and meiosis II, occur without intervening DNA replication. This contrasts with the mitotic cell cycle where DNA replication and chromosome segregation alternate to maintain the same ploidy. At the end of mitosis, CDKs are inactivated. This low CDK state in late mitosis/G1 allows for critical preparatory events for DNA replication and centrosome/spindle pole body (SPB) duplication. However, their execution is inhibited until S phase, where further preparatory events are also prevented. This "licensing" ensures that both the chromosomes and the centrosomes/SPBs replicate exactly once per cell cycle, thereby maintaining constant ploidy. Crucially, between meiosis I and meiosis II, centrosomes/SPBs must be re-licensed, but DNA re-replication must be avoided. In budding yeast, the Cdc14 protein phosphatase triggers CDK down regulation to promote exit from mitosis. Cdc14 also regulates the meiosis I to meiosis II transition, though its mode of action has remained unclear. Methods Fluorescence and electron microscopy was combined with proteomics to probe SPB duplication in cells with inactive or hyperactive Cdc14. Results We demonstrate that Cdc14 ensures two successive nuclear divisions by re-licensing SPBs at the meiosis I to meiosis II transition. We show that Cdc14 is asymmetrically enriched on a single SPB during anaphase I and provide evidence that this enrichment promotes SPB re-duplication. Cells with impaired Cdc14 activity fail to promote extension of the SPB half-bridge, the initial step in morphogenesis of a new SPB. Conversely, cells with hyper-active Cdc14 duplicate SPBs, but fail to induce their separation. Conclusion Our findings implicate reversal of key CDK-dependent phosphorylations in the differential licensing of cyclical events at the meiosis I to meiosis I transition.

Dataset Information

Multiple protein kinases influence the redistribution of fission yeast Clp1/Cdc14 phosphatase upon genotoxic stress.

Publications

Multiple protein kinases influence the redistribution of fission yeast Clp1/Cdc14 phosphatase upon genotoxic stress.

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