Microinjection of Cdc25 protein phosphatase into Xenopus prophase oocyte activates MPF and arrests meiosis at metaphase I

Abstract

Microinjection of bacterially expressed human cdc25A protein into Xenopus prophase oocytes provokes the activation of p34cdc2 kinase and the tyrosine dephosphorylation of p34cdc2 in the presence or absence of protein synthesis. The level of p34cdc2 kinase activity then drops in parallel with the degradation of cyclin B2 and finally increases again to stabilize at a high level. Cdc25 microinjection induces the assembly of a metaphase I spindle which is abnormally located in the deep cytoplasm. Moreover, oocytes arrest at the metaphase I stage and do not reach metaphase II even 10 h after cdc25 microinjection. The extended metaphase I period observed in cdc25-injected oocytes results from an equilibrium between degradation of cyclins and synthesis of new cyclins. This is in contrast with progesterone-stimulated oocytes where cyclin degradation is turned off when oocytes enter metaphase II. During metaphase I, the reactivation of MPF activity can be disrupted in two different ways: 1) cycloheximide, an inhibitor of protein synthesis, by preventing the synthesis of new cyclins, provokes the disappearance of MPF kinase activity and the reformation of a nucleus; 2) when the cAMP level is increased during the metaphase I period in cdc25-injected oocytes, MPF kinase activity drops following a rephosphorylation of tyrosine 15 of p34cdc2, while the cyclin turn-over remains unaffected. Moreover, increasing the cAMP level in prophase oocytes totally prevents the action of cdc25. Our results indicate that in Xenopus oocytes, the PKA pathway negatively regulates the activation of MPF and the activity of p34cdc2/cyclin B complex through tyrosine phosphorylation of p34cdc2 during metaphase I.

Inactivation of protein kinase A is not required for c-mos translation during meiotic maturation of Xenopus oocytes

“…Recently, it was proposed that PKA may have pleiotropic e ects and inhibit oocyte maturation at multiple points (Matten et al, 1994;Rime et al, 1994). One of these e ects was proposed to be the inhibition of endogenous c-mos synthesis, another e ect the suppression of cdc25 activation.…”

Potential role of protein tyrosine phosphatase nonreceptor type 13 in the control of oocyte meiotic maturation

“…It is well established that the dual-specific phosphatase Cdc25 is involved in oocyte maturation by controlling the phosphorylation state of one of the components of the maturation promoting factor (MPF), Cdc2 (Gautier et al, 1991;Kumagai and Dunphy, 1991;Lincoln et al, 2002;Rime et al, 1994). Therefore, one possibility is that PTPN13 substitutes for Cdc25 by dephosphorylating Tyr15 on Cdc2.…”

“…The distal components are well characterized (Karaiskou et al, 2001;Nebreda and Ferby, 2000;Schmitt and Nebreda, 2002b). Extensive data are available on the control of MPF activity and the most immediate partners, including Cdc25 (Gautier et al, 1991;Kumagai and Dunphy, 1991;Rime et al, 1994) and Myt1 (Palmer et al, 1998). Activation of MAP kinase cascade (Gotoh et al, 1995;Nebreda and Hunt, 1993) and de novo synthesis of Ringo/Speedy (Ferby et al, 1999;Lenormand et al, 1999) are additional components involved in the activation of MPF and resumption of meiosis.…”

Phosphorylation Mediated Regulation of Cdc25 Activity, Localization and Stability

“…[40,45] Injection of recombinant Xenopus Cdc25A or Cdc25C into oocytes induces their maturation, the first direct indication that this protein was a positive regulator of MPF. [40,45,46] Xenopus Cdc25C also complements the cdc25-22 ts mutation in fission yeast. [45] Four Cdc25 homologues are present in Caenorhabditis [47], whereas Drosophila contains two, string and twine.…”