CDC25A phosphatase controls meiosis I progression in mouse oocytes

Abstract

CDK1 is a pivotal regulator of resumption of meiosis and meiotic maturation of oocytes. CDC25A/B/C are dual-specificity phosphatases and activate cyclin-dependent kinases (CDKs). Although CDC25C is not essential for either mitotic or meiotic cell cycle regulation, CDC25B is essential for CDK1 activation during resumption of meiosis. Cdc25a -/- mice are embryonic lethal and therefore a role for CDC25A in meiosis is unknown. We report that activation of CDK1 results in a maturation-associated decrease in the amount of CDC25A protein, but not Cdc25a mRNA, such that little CDC25A is present by metaphase I. In addition, expression of exogenous CDC25A overcomes cAMP-mediated maintenance of meiotic arrest. Microinjection of Gfp-Cdc25a and Gpf-Cdc25b mRNAs constructs reveals that CDC25A is exclusively localized to the nucleus prior to nuclear envelope breakdown (NEBD). In contrast, CDC25B localizes to cytoplasm in GV-intact oocytes and translocates to the nucleus shortly before NEBD. Over-expressing GFP-CDC25A, which compensates for the normal maturation-associated decrease in CDC25A, blocks meiotic maturation at MI. This MI block is characterized by defects in chromosome congression and spindle formation and a transient reduction in both CDK1 and MAPK activities. Lastly, RNAi-mediated reduction of CDC25A results in fewer oocytes resuming meiosis and reaching MII. These data demonstrate that CDC25A behaves differently during female meiosis than during mitosis, and moreover, that CDC25A has a function in resumption of meiosis, MI spindle formation and the MI-MII transition. Thus, both CDC25A and CDC25B are critical for meiotic maturation of oocytes.

Protein kinase a modulates Cdc25B activity during meiotic resumption of mouse oocytes

“…In support of this idea, a cell cycle-dependent transport of Cdc25B in and out of the nucleus, has been demonstrated in somatic cells (Girard et al, 1992;Garner-Hamrick and Fisher, 1998;Dalal et al, 1999;Davezac et al, 2000) and in mouse oocytes (Solc et al, 2008). Our immunocytochemical experiments revealed a restricted cytoplasmic localization of Cdc25B in G 2 -arrested oocytes and nuclear localization shortly before GVBD.…”

“…The results indicated that Cdc25A also play a role in meiotic resumption of mouse oocytes. The differences in GVBD-inducing ability of the two phosphatases, Cdc25A and Cdc25B, suggest that they have different substrates and nonredundant functions (Solc et al, 2008 (Yaffe et al, 2001;Obenauer et al, 2003). This program has been quite successful and some predictions have been experimentally verified (Gratton et al, 2001;Zhou et al, 2001;Gottlieb et al, 2002;Kim et al, 2002Kim et al, , 2005She et al, 2004;Park et al, 2006;Lemeer et al, 2008).…”

Checkpoint kinase 1 is essential for meiotic cell cycle regulation in mouse oocytes

“…The subcellular localization of Chk1 was controversial, with centrosomes, nuclear, cytoplasm and kinetochores being considered as possible sites. 25,[31][32][33] We find that Chk1 is localized in the germinal vesicle and to the spindle from pro-MI to MII stages, 6 -Chk1 mRNA and arrested in M2 medium containing 2.5 μM milrinone for 2 h before being collected for western blot. (B) Change of myc was calculated by gray-scale analysis using the software Quantity one (Bio-Rad).…”

DNA damage response during mouse oocyte maturation

“…Oocytes were microinjected as previously described 50 in M2 medium with 5 pl of 80 ng/ml H2b-mCherry, 51 125 ng/ml Egfp-Cenp-C, 51 150 ng/ml 2mEGFP-CENP-C 23 , 50 ng/ml Securin-Egfp 52 and 125 ng/ml Map4-Egfp 53 cRNAs. Oocytes were cultured for 2 h in MEM medium supplemented with milrinone or IBMX to allow protein expression after mRNAs microinjection.…”