Protein kinase A acts at multiple points to inhibit Xenopus oocyte maturation.

Abstract

In Xenopus oocytes, initiation of maturation is dependent on reduction of cyclic AMP-dependent protein kinase (PKA) activity and the synthesis of the mos proto-oncogene product. Mos is required during meiosis I for the activation of both maturation-promoting factor (MPF) and mitogen-activated protein kinase (MAPK).Here we show that injection of the catalytic subunit of PKA (PKAc) The events of maturation, including germinal vesicle breakdown (GVBD), chromosome condensation, and formation of the metaphase spindles during both meiosis I and II, occur upon activation of maturation-promoting factor (MPF), a complex consisting of p34cdc2 and cyclin B (see reference 55 for a review). Conversion of pre-MPF to active MPF requires phosphorylation on threonine 161 (56) and activation of the cdc25 phosphatase, which removes inhibitory phosphorylations at 15,30,37). cdc25 is itself regulated by net phosphorylation (22,25,28).MPF activation in meiosis I requires not only a reduction in PKA activity but also new protein synthesis. Following progesterone exposure, the product of the c-mos proto-oncogene is synthesized and it has been shown to account for the protein synthesis requirement for GVBD and MPF activation (49,60 MATERUILS AND METHODSOocyte injections. Xenopus laevis females were purchased from Xenopus I (Ann Arbor, Mich.). Oocytes were surgically removed and defolliculated by incubation in modified Barth solution [MBS; 88 mM NaCl, 1 mM KCI, 2.5 mM NaHCO3, 10 mM N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid 4419

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.…”

“…Using a cell-free system prepared from Xenopus oocytes, Van Renterghem et al (1994) reported that pretreatment of extracts with PKAc had no e ect on the ability of cmos to activate MAPK, suggesting that both the direct activation of MAPKK by c-mos and the activation of MAPK by MAPKK are insensitive to PKA. In contrast, Matten et al (1994), using microinjection into oocytes, reported that microinjected PKAc decreased activation of MAPK by microinjected cmos to 50% of the control level.…”

“…However, protein synthesis is de®cient in diluted oocyte extracts, thus the possible e ect of PKAc on synthesis of endogenous c-mos was not taken into account. In the second report by Matten et al (1994) the amount of microinjected PKAc was as high as six nanograms per oocyte, which corresponds to an intracellular concentration of about 200 mM. As emphasized by the authors themselves,`it is possible to envision a coordinated localization of PKAc and cmos during the maturation process that would modulate the e ects of their kinase activities'.…”

“…Furthermore, microinjection of PKAc has been shown to prevent progesterone from activating MAP kinase (Matten et al, 1994). However, con¯icting results were obtained when the e ect of PKA on c-mos-induced activation of MAPK was investigated.…”

“…As progesterone was also reported to trigger an initial drop of cAMP in Xenopus oocytes (Speaker and Butcher, 1977), this led to the view that progesterone acts through inactivation of protein kinase A, leading to diminished phosphorylation of an (unidenti®ed) inhibitory protein (reviewed in Maller, 1983) which would in its PKA-phosphorylated form suppress, directly or indirectly, c-mos translation (Sagata, 1997). Consistent with this view, hormonestimulated oocytes microinjected with the catalytic subunit of PKA (PKAc) or treated with known elevators of intracellular cAMP, do not synthesize cmos (Matten et al, 1994).…”