RESUMEN
Xenopus oocytes are arrested in prophase of the first meiotic division. In response to progesterone, they re-enter meiosis and arrest again in metaphase of the second meiotic division. This process, called meiotic maturation, is under the control of the Cyclin B-Cdc2 complex, M phase promoting factor (MPF). Injection of a constitutively active Xenopus H-Ras protein activates MPF, suggesting that Ras proteins could be implicated in the progesterone transduction pathway. The aim of this study was (1) to elucidate the pathway triggered by H-Ras leading to MPF activation in Xenopus oocytes and (2) to investigate whether endogenous H-Ras is involved in the physiological process of meiotic maturation. We generated three constitutively active double mutants, each of them recruiting a single effector in mammalian cells, mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (PI3K) or RalGDS. Our results show that the activation of a PI3K-related enzyme is crucial for H-Ras-induced MPF activation, whereas the recruitment of either MAPK or RalGDS is not. However, although the H-Ras/PI3K pathway is functional in Xenopus oocytes, it is not the physiological transducer of progesterone responsible for meiotic resumption.
Asunto(s)
Genes ras , Oocitos/fisiología , Xenopus laevis/fisiología , Animales , División Celular , Femenino , Mutagénesis , Oocitos/citología , ARN Mensajero/genética , Xenopus laevis/genéticaRESUMEN
Since almost two decades, it is known that progesterone is responsible of the release of the prophase I arrest of amphibian oocytes and leads to the activation of the universal MPF, through a puzzling transduction pathway. It involves negative regulation of the cAMP-dependent protein kinase (PKA) and synthesis of new proteins, among them the c-Mos protooncogene product. The implication of the Mos/mitogenic activated protein kinase (MAP kinase) pathway in Cdc2 activation has been extensively studied and is now at the centre of a controversial debate. In this paper, we discuss the current progress and our recent results on the molecular mechanisms allowing progesterone to activate MPF and propose a model to partly resolve the long-standing inconsistencies concerning the role of Mos/MAP kinase during this process.
Asunto(s)
Proteína Quinasa CDC2/metabolismo , Oocitos/fisiología , Progesterona/metabolismo , Proteínas de Xenopus , Animales , Proteínas de Ciclo Celular/fisiología , Ciclinas/fisiología , Activación Enzimática , Genes mos/fisiología , Humanos , Factor Promotor de Maduración/fisiología , Meiosis/fisiología , Proteínas Quinasas Activadas por Mitógenos/fisiología , Proteínas de Unión al GTP Monoméricas/fisiología , Transducción de Señal , XenopusRESUMEN
Xenopus oocytes arrested in prophase I resume meiotic division in response to progesterone and arrest at metaphase II. Entry into meiosis I depends on the activation of Cdc2 kinase [M-phase promoting factor (MPF)]. To better understand the role of Cdc2, MPF activity was specifically inhibited by injection of the CDK inhibitor, Cip1. When Cip1 is injected at germinal vesicle breakdown (GVBD) time, Cdc25 and Plx1 are both dephosphorylated and Cdc2 is rephosphorylated on tyrosine. The autoamplification loop characterizing MPF is therefore not only required for MPF generation before GVBD, but also for its stability during the GVBD period. The ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C), responsible for cyclin degradation, is also under the control of Cdc2; therefore, Cdc2 activity itself induces its own inactivation through cyclin degradation, allowing the exit from the first meiotic division. In contrast, cyclin accumulation, responsible for Cdc2 activity increase allowing entry into metaphase II, is independent of Cdc2. The c-Mos/mitogen-activated protein kinase (MAPK) pathway remains active when Cdc2 activity is inhibited at GVBD time. This pathway could be responsible for the sustained cyclin neosynthesis. In contrast, during the metaphase II block, the c-Mos/MAPK pathway depends on Cdc2. Therefore, the metaphase II block depends on a dynamic interplay between MPF and CSF, the c-Mos/MAPK pathway stabilizing cyclin B, whereas in turn, MPF prevents c-Mos degradation.
Asunto(s)
Proteína Quinasa CDC2/fisiología , Metafase , Proteínas Quinasas Activadas por Mitógenos/fisiología , Oocitos/fisiología , Proteínas Proto-Oncogénicas c-mos/fisiología , Proteínas de Xenopus , Animales , Proteínas de Ciclo Celular , Ciclina B/metabolismo , Ciclina B1 , Inhibidor p21 de las Quinasas Dependientes de la Ciclina , Ciclinas/farmacología , Femenino , Meiosis , Fosforilación , Proteínas Serina-Treonina Quinasas/fisiología , Xenopus , Fosfatasas cdc25/fisiologíaRESUMEN
Xenopus prophase oocytes reenter meiotic division in response to progesterone. The signaling pathway leading to Cdc2 activation depends on neosynthesized proteins and a decrease in PKA activity. We demonstrate that Eg2 protein, a Xenopus member of the Aurora/Ipl1 family of protein kinases, accumulates in response to progesterone and is degraded after parthenogenetic activation. The polyadenylation and cap ribose methylation of Eg2 mRNA are not needed for the protein accumulation. Eg2 protein accumulation is induced by progesterone through a decrease in PKA activity, upstream of Cdc2 activation. Eg2 kinase activity is undetectable in prophase and is raised in parallel with Cdc2 activation. In contrast to Eg2 protein accumulation, Eg2 kinase activation is under Cdc2 control. Furthermore, by using an anti-sense strategy, we show that Eg2 accumulation is not required in the transduction pathway leading to Cdc2 activation. Altogether, our results strongly suggest that Eg2 is not necessary for Cdc2 activation, though it could participate in the organization of the meiotic spindles, in agreement with the well-conserved roles of the members of the Aurora family, from yeast to man.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Oocitos/enzimología , Progesterona/fisiología , Proteínas Quinasas/metabolismo , Animales , Aurora Quinasas , Proteína Quinasa CDC2/metabolismo , Proteína Quinasa CDC2/fisiología , Diferenciación Celular , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Activación Enzimática , Femenino , Meiosis , Oocitos/metabolismo , Oocitos/fisiología , Partenogénesis , Fosforilación , Poli A/metabolismo , Proteínas Serina-Treonina Quinasas , Caperuzas de ARN/metabolismo , ARN Mensajero/metabolismo , Ribosa/metabolismo , Proteínas de Xenopus , Xenopus laevisRESUMEN
Progesterone-induced meiotic maturation of Xenopus oocytes requires the synthesis of new proteins, such as Mos and cyclin B. Synthesis of Mos is thought to be necessary and sufficient for meiotic maturation; however, it has recently been proposed that newly synthesized proteins binding to p34(cdc2) could be involved in a signaling pathway that triggers the activation of maturation-promoting factor. We focused our attention on cyclin B proteins because they are synthesized in response to progesterone, they bind to p34(cdc2), and their microinjection into resting oocytes induces meiotic maturation. We investigated cyclin B accumulation in response to progesterone in the absence of maturation-promoting factor-induced feedback. We report here that the cdk inhibitor p21(cip1), when microinjected into immature Xenopus oocytes, blocks germinal vesicle breakdown induced by progesterone, by maturation-promoting factor transfer, or by injection of okadaic acid. After microinjection of p21(cip1), progesterone fails to induce the activation of MAPK or p34(cdc2), and Mos does not accumulate. In contrast, the level of cyclin B1 increases normally in a manner dependent on down-regulation of cAMP-dependent protein kinase but independent of cap-ribose methylation of mRNA.
Asunto(s)
Ciclina B/metabolismo , Ciclinas/metabolismo , Quinasas Quinasa Quinasa PAM/metabolismo , Oocitos/metabolismo , Progesterona/farmacología , Animales , Proteína Quinasa CDC2/metabolismo , Ciclina B1 , Inhibidor p21 de las Quinasas Dependientes de la Ciclina , Ciclinas/farmacología , Femenino , Factor Promotor de Maduración/metabolismo , Microinyecciones , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Mitosis , Ácido Ocadaico/farmacología , Oocitos/efectos de los fármacos , Unión Proteica , Proteínas Quinasas/metabolismo , Proteínas Recombinantes/metabolismo , Transducción de Señal , XenopusRESUMEN
The activation of Cdc2 kinase induces the entry into M-phase of all eukaryotic cells. We have developed a cell-free system prepared from prophase-arrested Xenopus oocytes to analyze the mechanism initiating the all-or-none activation of Cdc2 kinase. Inhibition of phosphatase 2A, the major okadaic acid-sensitive Ser/Thr phosphatase, in these extracts, provokes Cdc2 kinase amplification and concomitant hyperphosphorylation of Cdc25 phosphatase, with a lag of about 1 h. Polo-like kinase (Plx1 kinase) is activated slightly after Cdc2. All these events are totally inhibited by the cdk inhibitor p21(Cip1), demonstrating that Plx1 kinase activation depends on Cdc2 kinase activity. Addition of a threshold level of recombinant Cdc25 induces a linear activation of Cdc2 and Plx1 kinases and a partial phosphorylation of Cdc25. We propose that the Cdc2 positive feedback loop involves two successive phosphorylation steps of Cdc25 phosphatase: the first one is catalyzed by Cdc2 kinase and/or Plx1 kinase but it does not modify Cdc25 enzymatic activity, the second one requires a new kinase counteracted by phosphatase 2A. Furthermore we demonstrate that, under our conditions, Cdc2 amplification and MAP kinase activation are two independent events.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Factor Promotor de Maduración/biosíntesis , Oocitos/enzimología , Fosfoproteínas Fosfatasas/metabolismo , Animales , Proteína Quinasa CDC2/metabolismo , Proteínas Quinasas Dependientes de Calcio-Calmodulina/metabolismo , Citosol/enzimología , Activación Enzimática/efectos de los fármacos , Retroalimentación/efectos de los fármacos , Factor Promotor de Maduración/metabolismo , Ácido Ocadaico/farmacología , Oocitos/citología , Profase , Proteína Fosfatasa 2 , Proteínas Tirosina Fosfatasas/farmacología , Xenopus , Fosfatasas cdc25RESUMEN
Xenopus cyclin D2 mRNA is a member of the class of maternal RNAs. It is rare and stable during early embryonic development. To investigate the potential role of cyclin D2 during early embryonic cell cycles, cyclin D2 was injected into one blastomere of a two-cell embryo. This injection induced a cell cycle arrest in the injected blastomere. To analyze more precisely the mechanism of this arrest, we took advantage of cycling egg extracts that recapitulate major events of the cell cycle when supplemented with demembranated sperm heads. When Xenopus cyclin D2 is added to egg extracts, the first round of DNA replication occurs as in control extracts. However, Xenopus cyclin D2 blocks subsequent rounds of DNA replication and the oscillations of histone H1 kinase activity associated with cdc2 kinase, indicating that the cell cycle is arrested after the first S-phase. The block induced by Xenopus cyclin D2 is not due to a lack of the mitotic cyclin B2 that accumulates normally. Radiolabeled Xenopus cyclin D2 enters nuclei after completion of the first S-phase and remains stable over the entire period of the arrest. These features suggest that Xenopus cyclin D2 could play an original role during early development, controlling the G2-phase and/or the G2/M transition.
Asunto(s)
Ciclo Celular , Fase de Segmentación del Huevo/citología , Ciclinas/fisiología , Inhibidores de Crecimiento , Proteínas de Xenopus , Xenopus laevis/embriología , Animales , Compartimento Celular , Núcleo Celular/metabolismo , Ciclina B/fisiología , Ciclina E/metabolismo , Fase SRESUMEN
The function of the Xenopus c-mos proto-oncogene product (Mos(xe)) has been investigated during oocyte maturation. Experiments with a new antibody able to immunoblot Mos(xe) demonstrated the time course of MAP kinase (MAP K) activation in oocytes paralleled Mos(xe) accumulation, and in activated eggs the deactivation of MAP K paralleled the degradation of Mos(xe). Ablation of Mos synthesis by microinjection of antisense oligodeoxynucleotides abolished activation of MAP K by progesterone, but microinjection of GST-Mos fully restored both MAP K activation and germinal vesicle breakdown (GVBD). The Mos(xe) level at metaphase of Meiosis I (MI) was 2 - 3-fold less than that at metaphase of Meiosis II (MII), but MAP K activation was maximal at metaphase in both MI and MII. In the transition between MI and MII, both cyclin B and Mos(xe) levels rapidly declined in the presence of cycloheximide and injection of exogenous GST-Mos(xe) did not prevent degradation of either protein, although MAP K was activated. Microinjection of GST-Mos(xe) into oocytes was able to activate MAP K before GVBD and H1 kinase activation, and microinjection of constitutively-activated thiophosphorylated MAP K induced de novo synthesis of Mos(xe) before H1 kinase activation, suggesting the existence of a positive feedback loop between MAP K and Mos(xe) accumulation.
Asunto(s)
Proteínas Oncogénicas v-mos/fisiología , Oocitos/fisiología , Animales , Proteínas Quinasas Dependientes de Calcio-Calmodulina/metabolismo , Activación Enzimática , Femenino , Genes mos , Meiosis/fisiología , Metafase/fisiología , Proteínas Oncogénicas v-mos/genética , Proteínas Oncogénicas v-mos/metabolismo , Oocitos/citología , Oocitos/enzimología , Xenopus laevisRESUMEN
Mitogen-activated protein kinase (MAPK) is one of the protein kinases activated during meiotic maturation of Xenopus laevis oocytes. The c-Mosxe protein kinase, which has been shown to be sufficient to promote germinal vesicle breakdown (GVBD) in meiosis I, can directly activate MAP kinase kinase in vitro and leads to the activation of MAPK in vivo. Recently we have shown that constitutively activated MAPK induces metaphase arrest when injected into one blastomere of a two-cell embryo. This arrest mimics the natural arrest of vertebrate unfertilized eggs in second meiotic metaphase due to cytostatic factor and c-Mosxe activity. We show here that microinjection of constitutively activated thiophosphorylated MAPK into resting oocytes is able to activate maturation-promoting factor (MPF) and promote GVBD. These results strongly support the hypothesis that MAPK plays an important role in the pathway that links c-Mosxe to the activation of MPF.
Asunto(s)
Proteínas Quinasas Dependientes de Calcio-Calmodulina/farmacología , Meiosis/efectos de los fármacos , Oocitos/citología , Animales , Oocitos/efectos de los fármacos , Oogénesis , XenopusRESUMEN
p34cdc2 protein is found in prophase, metaphase and activated Xenopus oocytes at a similar level whereas its kinase activity oscillates within meiosis. Using an anti-PSTAIRE antibody that recognizes Xenopus p34cdc2, it was demonstrated that the major part of p34cdc2 was associated with microtubules isolated in vitro from Xenopus oocytes. Conversely, tubulin was recovered in association with p34cdc2 in p13-Sepharose pellets. The abundance of the fraction of p34cdc2 which was associated with microtubules did not oscillate during the meiotic maturation and the activation process. By contrast, the histone H1 kinase activity of p34cdc2 estimated in microtubular oocyte pellets was much higher in metaphase than in prophase oocytes. Cyclin B, which is associated in vivo with p34cdc2 in prophase and metaphase oocytes, was also present in the microtubular fractions. However, cyclin was not necessary for the binding of p34cdc2 to microtubules since p34cdc2 from activated eggs, where cyclin was missing, still copurified with microtubules. Purified MAP2, but not tubulin, was able to bind to p34cdc2, demonstrating that the association between p34cdc2 and microtubules was mediated by microtubule-associated proteins. During the meiotic maturation of Xenopus oocytes, several protein kinases were activated, among them MAP kinase. MAP kinase also associated with microtubules. It was demonstrated that both p34cdc2 kinase and MAP kinase purified from Xenopus oocytes were able to phosphorylate in vitro rat brain MAP2. However both protein kinases phosphorylated different domains of MAP2, suggesting that they might regulate microtubules in different ways.
Asunto(s)
Proteína Quinasa CDC2/metabolismo , Proteínas Quinasas Dependientes de Calcio-Calmodulina/metabolismo , Meiosis , Microtúbulos/enzimología , Oocitos/ultraestructura , Animales , Sitios de Unión , Química Encefálica , Precipitación Química , Ciclinas/fisiología , Femenino , Microesferas , Proteínas Asociadas a Microtúbulos/metabolismo , Oocitos/fisiología , Fosforilación , Ratas , Tubulina (Proteína)/metabolismo , Xenopus laevisRESUMEN
The natural arrest of vertebrate unfertilized eggs in second meiotic metaphase results from the activity of cytostatic factor (CSF). The product of the c-mos(xe) proto-oncogene is thought to be a component of CSF and can induce metaphase arrest when injected into blastomeres of two-cell embryos. The c-Mos(xe) protein can directly activate the mitogen-activated protein kinase kinase (MAP kinase kinase) in vitro, leading to activation of MAP kinase. MAP kinase and c-Mos(xe) are active in unfertilized eggs and are rapidly inactivated after fertilization. Microinjection of thiophosphorylated MAP kinase into one blastomere of a two-cell embryo induced metaphase arrest similar to that induced by c-Mos(xe). However, only arrest with c-Mos(xe) was associated with activation of endogenous MAP kinase. These results indicate that active MAP kinase is a component of CSF in Xenopus and suggest that the CSF activity of c-Mos(xe) is mediated by MAP kinase.
Asunto(s)
Blastómeros/citología , Metafase , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-mos/metabolismo , Secuencia de Aminoácidos , Animales , Blastómeros/metabolismo , Activación Enzimática , Proteína Quinasa 1 Activada por Mitógenos , Quinasas de Proteína Quinasa Activadas por Mitógenos , Modelos Biológicos , Datos de Secuencia Molecular , Fosforilación , Proteínas Quinasas/metabolismo , Xenopus laevisRESUMEN
Mitogen-activated protein kinase (MAP kinase) is a serine/threonine kinase whose enzymatic activity is thought to play a crucial role in mitogenic signal transduction and also in the progesterone-induced meiotic maturation of Xenopus oocytes. We have purified MAP kinase from Xenopus oocytes and have shown that the protein is present in metaphase II oocytes under two different forms: an inactive 41-kD protein able to autoactivate and to autophosphorylate in vitro, and an active 42-kD kinase resolved into two tyrosine phosphorylated isoforms on 2D gels. During meiotic maturation, MAP kinase becomes tyrosine phosphorylated and activated following the activation of the M-phase promoting factor (MPF), a complex between the p34cdc2 kinase and cyclin B. In vivo, MAP kinase activity displays a different stability in metaphase I and in metaphase II: protein synthesis is required to maintain MAP kinase activity in metaphase I but not in metaphase II oocytes. Injection of either MPF or cyclin B into prophase oocytes promotes tyrosine phosphorylation of MAP kinase, indicating that its activation is a downstream event of MPF activation. In contrast, injection of okadaic acid, which induces in vivo MPF activation, promotes only a very weak tyrosine phosphorylation of MAP kinase, suggesting that effectors other than MPF are required for the MAP kinase activation. Moreover, in the absence of protein synthesis, cyclin B and MPF are unable to promote in vivo activation of MAP kinase, indicating that this activation requires the synthesis of new protein(s).
Asunto(s)
Oocitos/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Animales , Activación Enzimática , Éteres Cíclicos/farmacología , Femenino , Técnicas In Vitro , Factor Promotor de Maduración/farmacología , Proteína Quinasa 1 Activada por Mitógenos , Ácido Ocadaico , Oocitos/efectos de los fármacos , Oocitos/crecimiento & desarrollo , Fosforilación , Biosíntesis de Proteínas , Proteínas Serina-Treonina Quinasas/aislamiento & purificación , Proteínas Tirosina Quinasas/aislamiento & purificación , Tirosina/metabolismo , Xenopus laevisRESUMEN
Microinjection of a bacterially expressed stable delta 90 sea urchin cyclin B into Xenopus prophase oocytes, in absence or presence of cycloheximide, provokes the activation of histone H1 kinase and the tyrosine dephosphorylation of p34cdc2. Unexpectedly, when prophase oocytes are submitted to a treatment known to elevate the intracellular cAMP level (3-isobutyl-1-methylxanthine and cholera toxin), delta 90 cyclin has no effect and the oocytes remain blocked in prophase. This inhibition is reverted by the microinjection of the inhibitor of cAMP-dependent protein kinase. When delta 90 cyclin is microinjected into oocytes depleted of endogenous cyclins (cycloheximide-treated metaphase I) and in the presence of a high intracellular concentration of cAMP, p34cdc2 kinase is tyrosine rephosphorylated. Altogether, our results indicate that in Xenopus oocyte, cAMP-dependent protein kinase (A-kinase) controls the formation of the cyclin B/p34cdc2 complex which remains inactive and tyrosine phosphorylated.
Asunto(s)
Proteína Quinasa CDC2/metabolismo , Ciclinas/farmacología , Oocitos/metabolismo , Proteínas Quinasas/fisiología , Animales , Ciclinas/antagonistas & inhibidores , Cicloheximida/farmacología , Activación Enzimática/efectos de los fármacos , Femenino , Microinyecciones , Fosforilación/efectos de los fármacos , Profase , Inhibidores de Proteínas Quinasas , Tirosina/metabolismo , XenopusRESUMEN
The tyrosine phosphorylation/dephosphorylation of p34cdc2 was estimated by immunoblotting with antiphosphotyrosine antibody during meiotic maturation of Xenopus oocytes. At the time of germinal vesicle breakdown (GVBD), p34cdc2 is tyrosine dephosphorylated whereas a p42 protein, which might correspond to a MAP2 kinase, becomes tyrosine phosphorylated. No modification in the level of tyrosine phosphorylation of either proteins was noticed during the whole maturation process from GVBD until metaphase II. When added to prophase oocytes, 6-DMAP (6-dimethyl-aminopurine) blocks GVBC, M-phase-promoting factor (MPF) activation and H1-histone, kinase activation induced by either progesterone, MPF transfer or okadaic acid microinjection. In each case, the tyrosine dephosphorylation reaction of p34cdc2 is inhibited. In meiosis I oocytes (just after the initiation of GVBD), 6-DMAP provokes the rephosphorylation of p34cdc2 on tyrosine residue(s), inactivation of MPF and H1-histone kinase and re-entry of the cell into an interphase-like state. These processes are reversible by simply removing the agent. In contrast to the observations in prophase oocytes, okadaic acid is able to reverse the inhibitory effect of 6-DMAP in meiosis I oocytes on MPF and H1-histone kinase activities and to initiate dephosphorylation of p34cdc2 on tyrosyl residue(s) even in the presence of 6-DMAP. Altogether, our results show that 6-DMAP and okadaic acid antagonistically control in vivo the level of tyrosine phosphorylation of p34cdc2.
Asunto(s)
Proteína Quinasa CDC2/metabolismo , Factor Promotor de Maduración/fisiología , Meiosis/fisiología , Oocitos/metabolismo , Proteínas/metabolismo , Tirosina/metabolismo , Adenina/análogos & derivados , Adenina/farmacología , Animales , Éteres Cíclicos/farmacología , Femenino , Immunoblotting , Ionóforos/farmacología , Ácido Ocadaico , Oocitos/citología , Fosforilación/efectos de los fármacos , Xenopus laevisRESUMEN
We have characterized a serine/threonine protein kinase from Xenopus metaphase-II-blocked oocytes, which phosphorylates in vitro the microtubule-associated protein 2 (MAP2). The MAP2 kinase activity, undetectable in prophase oocytes, is activated during the progesterone-induced meiotic maturation (G2-M transition of the cell cycle). p-Nitrophenyl phosphate, a phosphatase inhibitor, is required to prevent spontaneous deactivation of the MAP2 kinase in crude preparations; conversely, the partially purified enzyme can be in vitro deactivated by the low-Mr polycation-stimulated (PCSL) phosphatase (also termed protein phosphatase 2A2), working as a phosphoserine/phosphothreonine-specific phosphatase and not as a phosphotyrosyl phosphatase indicating that phosphorylation of serine/threonine is necessary for its activity. S6 kinase, a protein kinase activated during oocyte maturation which phosphorylates in vitro ribosomal protein S6 and lamin C, can be deactivated in vitro by PCSL phosphatase. S6 kinase from prophase oocytes can also be activated in vitro in fractions known to contain all the factors necessary to convert pre-M-phase-promoting factor (pre-MPF) to MPF. Active MAP2 kinase can activate in vitro the inactive S6 kinase present in prophase oocytes or reactivate S6 kinase previously inactivated in vitro by PCSL phosphatase. These data are consistent with the hypothesis that the MAP2 kinase is a link of the meiosis signalling pathway and is activated by a serine/threonine kinase. This will lead to the regulation of further steps in the cell cycle, such as microtubular reorganisation and S6 kinase activation.