Microinjection of EGTA into prophase-blocked oocytes does not inhibit hormone-induced meiosis reinitiation, although it prevents oocyte activation by fertilization, by ionophore A23187, or by subsequent microinjection of otherwise efficient Ca2+ buffers. In contrast microinjection of Ca2+ buffers inhibits 1-methyladenine-induced meiosis reinitiation. Oocytes can be released from Ca2+ inhibition by raising hormone concentration or by the subsequent transfer of cytoplasm taken from maturing oocytes. Ca2+-microinjected oocytes remain inhibited up to 1 h after microinjection, although free Ca2+ concentration comes back to its resting value less than 30 sec after microinjection. Cyanide, which decreases ATP content and depresses Ca2+-pumping activity, reversibly inhibits 1-methyladenine-induced meiosis reinitiation. These results do not support the hypothesis that Ca2+ is the second messenger of the hormone in meiosis reinitiation of starfish oocytes, although they support the view that elimination of Ca2+ from some component of the oocyte cortex (perhaps the plasma membrane) might be a compulsory event for transduction of the hormonal message.

1-methyladenine stimulates the transfer of the γ-phosphoryl group from ATP to high molecular weight proteins which have been purified by column chromatography. SDS gel electrophoresis shows rather similar pattern of 32P-labelled polypeptides excepted for a 45,500 daltons component which becomes phosphorylated only in hormone-treated oocytes.

Dramatic changes of cell organisation occur at onset of mitosis. Genetic analysis of fission yeast and physiological studies of vertebrate and invertebrate oocytes showed that activation of cyclin B-cdc2 kinase triggers mitosis. Nevertheless, upstream mechanisms responsible for this activation remain largely unknown in somatic cells of higher eukaryotes. This review discusses possible pathways and mechanisms involved in triggering onset of mitosis in such cells, including inhibitory checkpoint mechanisms that detect defects in structural organisation of the cell.

Dramatic changes of cell organisation occur at onset of mitosis. Genetic analysis of fission yeast and physiological studies of vertebrate and invertebrate oocytes showed that activation of cyclin B-cdc2 kinase triggers mitosis. Nevertheless, upstream mechanisms responsible for this activation remain largely unknown in somatic cells of higher eukaryotes. This review discusses possible pathways and mechanisms involved in triggering onset of mitosis in such cells, including inhibitory checkpoint mechanisms that detect defects in structural organisation of the cell.

The resumption of meiosis in Xenopus arrested oocytes is triggered by progesterone, which leads to polyadenylation and translation of Mos mRNA, then activation of MAPK pathway. While Mos protein kinase has been reported to be essential for re-entry into meiosis in Xenopus, arrested oocytes can undergo germinal vesicle breakdown (GVBD) independently of MAPK activation, leading us to question what the Mos target might be if Mos is still required. We now demonstrate that Mos is indeed necessary, although is independent of the MAPK cascade, for conversion of inactive pre-MPF into active MPF. We have found that Myt1 is likely to be the Mos target in this process, as Mos interacts with Myt1 in oocyte extracts and Mos triggers Myt1 phosphorylation on some sites in vivo, even in the absence of MAPK activation. We propose that Mos is involved, not only in the MAPK cascade pathway, but also in a mechanism that directly activates MPF in Xenopus oocytes.

At the G2/M transition of the cell cycle, the cdc25c phosphatase dephosphorylates inhibitory residues of cdc2, and cyclin‐B–cdc2 kinase (MPF) is activated. Phosphorylation of cyclin B1 induces its nuclear accumulation, and, since cdc25c is also believed to accumulate and activate shortly before G2/M in the nucleus, it has been proposed that this induces cyclin‐B1–cdc2 kinase activation. We demonstrate that cyclin B1 phosphorylation has another essential function in vivo: it is required for cdc25c and MPF activation, which does not require nuclear accumulation of cyclin B1, and occurs in the cytoplasm.

The idea that Cdc2 and cyclins play a key role in the control of the G2/M transition of the cell cycle came largely from genetic analysis of fission yeast and physiological studies of clam, frog, sea urchin and starfish eggs and oocytes. However, it took a long time to realise that Cdc2 and cyclins form a stoichiometric complex and that a cyclin subunit is necessary for the Cdc2 subunit to gain its protein kinase activity.Cyclins were first recognized as proteins whose abundance oscillates during the early cell cycles of marine invertebrate eggs and their connection with MPF (maturation-promoting factor), the entity defined in frog and starfish oocytes whose activity controls entry into M phase, was far from clear at first. Indeed, it was a long time before MPF was shown to be a protein kinase, and direct proof that MPF is a heterodimer comprising one molecule of cyclin and one molecule of Cdc2 was finally obtained only when the Cdc2-associated component of purified starfish MPF was sequenced and found to be cyclin B. When this fundamental discovery was confirmed in vertebrates and mammalian members of the Cdc2 family were also shown to bind cyclins, Cdc2 became Cdk1, the first cyclin-dependent protein kinase.

Throughout oocyte maturation, and subsequently during the first mitotic cell cycle, the MAP kinase cascade and cyclin-B-Cdc2 kinase are associated with the control of cell cycle progression. Many roles have been directly or indirectly attributed to MAP kinase and its influence on cyclin-B-Cdc2 kinase in different model systems; yet a principle theme does not emerge from the published literature, some of which is apparently contradictory. Interplay between these two kinases affects the major events of meiotic maturation throughout the animal kingdom, including the suppression of DNA replication, the segregation of meiotic chromosomes, and the prevention of parthenogenetic activation. Central to many of these events appears to be the control by MAP kinase of cyclin translation and degradation.

Leland H. Hartwell, Américain, est né le 30 octobre 1939 à Los Angeles (Californie, États-Unis). Entré en 1996 au Fred Hutchinson Cancer Research de Seattle (États-Unis), il en est depuis 1997 le Président directeur général et l’administrateur. Membre de l’académie américaine des sciences depuis 1987, il a rejoint en 1968 l’Université de l’État de Washington où il enseigne la génétique depuis 1973. En novembre 2000, il a reçu à Paris le Prix Leopold Griffuel décerné par l’Association française pour la recherche sur le cancer.R. Timothy Hunt, Britannique, né le 19 février 1943, a obtenu son PhD de biologie à l’Université de Cambridge (Grande-Bretagne), il a ensuite poursuivi ses études post-doctorales à l’École de médecine Albert-Einstein de New York (États-Unis), avant de revenir au département de biochimie de l’Université de Cambridge, puis a rejoint en 1990 l’Imperial Cancer Research Fund, à Londres (Grande-Bretagne) où il exerce encore aujourd’hui. Paul M. Nurse, Britannique, né le 25 janvier 1949, est depuis septembre 1996 le Directeur général de l’Imperial Cancer Research Fund de Londres (Grande-Bretagne). Titulaire d’un PhD de biologie de l’Université d’East Anglia, il a travaillé au département de zoologie de l’Université d’Edinburgh (de 1974 à 1980), à l’École de biologie de l’Université du Sussex (de 1980 à 1987), puis à l’Imperial Cancer Research Fund, non sans être passé par l’Université d’Oxford (de 1987 à 1996). Lauréat du Prix Lasker en 1998, il est membre de l’académie américaine des sciences.

Living cells reproduce by duplicating their content and dividing in two. At a minimum, the cell cycle comprises the set of processes that a cell must go through in order to faithfully replicate its DNA and to segregate duplicated chromosomes (mitosis) in two separate daughter cells. Recent data suggest that a central system operates, which is distinct from the machinery performing the essential processes of the mitotic cell cycle. This central clock is regulated at specific checkpoints where feedback signals from the downstream processes can delay the triggering of the next step before the previous one is achieved. This biochemical device of cell cycle control is based on the regulation of the activity of cyclin-dependent protein kinases (cdks). Cyclins bind to cdks and regulate their ability to phosphorylate specific substrates. Moreover, the activity of some cdks, and therefore the ability of a given cell to divide, seems to depend on its "history". This information could play a crucial role in various processes such as differenciation, organogenesis and senescence.

Les transitions du cycle, lors desquelles chaque cellule prend la decision de poursuivre ou de s'arreter, sont controlees par une serie de proteine-kinases dependantes des cyclines. L'activite de ces kinases depend d'une variete de mecanismes reversibles ou irreversibles qui orientent et limitent la proliferation des cellules au cours du temps. Ces mecanismes jouent un role crucial dans le developpement, puisque l'historique des cycles precedents semble fournir a chaque cellule de l'organisme un indicateur de position jouant un role decisif dans la differenciation et l'organogenese. Nos cycles cellulaires sont comptes, ils font l'objet de regulations programmees au cours du developpement embryonnaire, et participent a la senescence et a la mort cellulaire. Les cancers resultent souvent d'un dysfonctionnement de leurs mecanismes de regulation.

Here we show that segregation of homologous chromosomes and that of sister chromatids are differentially regulated in Xenopus and possibly in other higher eukaryotes. Upon hormonal stimulation, Xenopus oocytes microinjected with antibodies against the anaphase-promoting complex (APC) activator Fizzy or the APC core subunit Cdc27, or with the checkpoint protein Mad2, a destruction-box peptide or methylated ubiquitin, readily progress through the first meiotic cell cycle and arrest at second meiotic metaphase. However, they fail to segregate sister chromatids and remain arrested at second meiotic metaphase when electrically stimulated or when treated with ionophore A34187, two treatments that mimic fertilization and readily induce chromatid segregation in control oocytes. Thus, APC is required for second meiotic anaphase but not for first meiotic anaphase.

Members of the polo-like family of protein kinases have been involved in the control of APC (anaphase-promoting complex) during the cell cycle, yet how they activate APC is not understood in any detail. In Xenopus oocytes, Ca2+-dependent degradation of cyclin B associated with release from arrest at second meiotic metaphase was demonstrated to require the polo-like kinase Plx1. The aim of the present study was to examine, beyond Ca2+-dependent resumption of meiosis, the possible role of Plx1 in the control of cyclin degradation during the early mitotic cell cycle. Plx1 was found to be dispensable for MPF to turn on the cyclin degradation machinery. However, it is required to prevent premature inactivation of the APC-dependent proteolytic pathway. Microcystin suppresses the requirement for Plx1 in both Ca2+-dependent exit from meiosis, associated with degradation of both cyclin B and A downstream of CaMK2 activation, and prevention of premature APC(Fizzy) inactivation in the early mitotic cell cycle. These results are consistent with the view that Plx1 antagonizes an unidentified microcystin-sensitive phosphatase that inactivates APC(Fizzy).

During Xenopus oocyte maturation, the Mos protein kinase is synthesized and activates the MAP kinase cascade. In this report, we demonstrate that the synthesis and activation of Mos are two separable processes. We find that Hsp90 function is required for activation and phosphorylation of Mos and full activation of the MAP kinase cascade. Once Mos is activated, Hsp90 function is no longer required. We show that Mos interacts with both Hsp90 and Hsp70, and that there is an inverse relationship between association of Mos with these two chaperones. We propose that Mos protein kinase is activated by a novel mechanism involving sequential association with Hsp70 and Hsp90 as well as phosphorylation. We also present evidence for a two‐phase activation of MAP kinase in Xenopus oocytes.

We have examined the expression of glycogen synthase kinase-3beta in oocytes and early embryos of Xenopus and found that the protein is developmentally regulated. In resting oocytes, GSK-3beta is active and it is inactivated on maturation in response to progesterone. GSK-3beta inactivation is necessary and rate limiting for the cell cycle response to this hormone and the subsequent accumulation of beta-catenin. Overexpression of a dominant negative form of the kinase accelerates maturation, as does inactivation by expression of Xenopus Dishevelled or microinjection of an inactivating antibody. Cell cycle inhibition by GSK-3beta is not mediated by the level of beta-catenin or by a direct effect on either the MAP kinase pathway or translation of mos and cyclin B1. These data indicate a novel role for GSK-3beta in Xenopus development: in addition to controlling specification of the dorsoventral axis in embryos, it mediates cell cycle arrest in oocytes.

MAP kinase activation occurs during meiotic maturation of oocytes from all animals, but the requirement for MAP kinase activation in reinitiation of meiosis appears to vary between different classes. In particular, it has become accepted that MAP kinase activation is necessary for progesterone-stimulated meiotic maturation of Xenopus oocytes, while this is clearly not the case in other systems. In this paper, we demonstrate that MAP kinase activation in Xenopus oocytes is an early response to progesterone and can be temporally dissociated from MPF activation. We show that MAP kinase activation can be suppressed by treatment with geldanamycin or by overexpression of the MAP kinase phosphatase Pyst1. A transient and low-level early activation of MAP kinase increases the efficiency of cell cycle activation later on, when MAP kinase activity is no longer essential. Many oocytes can still undergo reinitiation of meiosis in the absence of active MAP kinase. Suppression of MAP kinase activation does not affect the formation or activation of Cdc2-cyclin B complexes, but reduces the level of active Cdc2 kinase. We discuss these findings in the context of a universal mechanism for meiotic maturation in oocytes throughout the animal kingdom.