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PMID:19223768

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Citation

Pirino, G, Wescott, MP and Donovan, PJ (2009) Protein kinase A regulates resumption of meiosis by phosphorylation of Cdc25B in mammalian oocytes. Cell Cycle 8:665-70

Abstract

In mammalian oocytes, meiosis arrests at prophase I. Meiotic resumption requires activation of Maturation-Promoting Factor (MPF), comprised of a catalytic Cyclin-dependent kinase-1 (Cdk1) and a regulatory subunit cyclin B and results in germinal vesicle breakdown (GVBD). Cyclic AMP (cAMP)-mediated Protein Kinase A (PKA) activity sustains prophase arrest by inhibiting Cdk1. However, the link between PKA activity and MPF inhibition remains unclear. Cdc25 phosphatases can activate Cdks by removing inhibitory phosphates from Cdks. Thus one method for sustaining prophase arrest could be inhibition of the activity of the Cdc25 protein required for MPF activation. Indeed, studies in Xenopus identify Cdc25C as a target of PKA activity in meiosis. However, in mice, studies suggest that Cdc25B is the phosphatase essential for GVBD and, therefore, the likely target of PKA activity. To assess these questions, we targeted a potential PKA substrate, a highly conserved serine 321 residue of Cdc25B and evaluated the effect on oocyte maturation. A Cdc25B-Ser321Ala point mutant mRNA induces GVBD when injected into prophase-arrested oocytes more rapidly than wild type mRNA. Using fluorescently-tagged proteins we also determined that the mutant protein enters the nucleus more rapidly than its wildtype counterpart. These data suggest that phosphorylation of the Ser321 residue plays a key role in the negative regulation and localization of Cdc25B during prophase arrest. PKA also phosphorylates a wildtype Cdc25B protein but not a Ser321Ala mutant protein in vitro. Mutation of Ser321 in Cdc25B also affects its association with a sequestering protein, 14-3-3. Our studies suggest that Cdc25B is a direct target of PKA in prophase-arrested oocytes and that Cdc25B phosphorylation results in its inhibition and sequestration by the 14-3-3 protein.

Links

PubMed

Keywords

14-3-3 Proteins/metabolism; Animals; Cell Line; Cyclic AMP-Dependent Protein Kinases/genetics; Cyclic AMP-Dependent Protein Kinases/metabolism; Female; Meiosis/physiology; Mice; Mutagenesis, Site-Directed; Oocytes/physiology; Phosphorylation; Serine/metabolism; cdc25 Phosphatases/genetics; cdc25 Phosphatases/metabolism

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

MOUSE:KAPCA

involved_in

GO:0051726: regulation of cell cycle

ECO:0000314: direct assay evidence used in manual assertion

P

Seeded From UniProt

complete

MOUSE:KAPCA

involved_in

GO:0071158: positive regulation of cell cycle arrest

ECO:0000314: direct assay evidence used in manual assertion

P

Seeded From UniProt

complete


See also

References

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