The Ras and Rafl proto-oncogenes transduce extracellular signals that promote cell growth. Cdc25 phosphatases activate the cell division cycle by dephosphorylation of critical threonine and tyrosine residues within the cyclin-dependent kinases. We show here that Cdc25 phosphatase associates with rafl in somatic mammalian cells and in meiotic frog oocytes. Furthermore, Cdc25 phosphatase can be activated in vitro in a Rafl-dependent manner. We suggest that activation of the cell cycle by the Ras/Rafl pathways might be mediated in part by Cdc25.

During oogenesis, oocytes are arrested in prophase and resume meiosis by activating the kinase Cdk1 upon hormonal stimulation. In all vertebrates, release from prophase arrest relies on protein kinase A (PKA) downregulation and on the dephosphorylation of a long-sought but still unidentified substrate. Here we show that ARPP19 is the PKA substrate whose phosphorylation at serine 109 is necessary and sufficient for maintaining Xenopus oocytes arrested in prophase. By downregulating PKA, progesterone, the meiotic inducer in Xenopus, promotes partial dephosphorylation of ARPP19 that is required for the formation of a threshold level of active Cdk1. Active Cdk1 then initiates MPF autoamplification loop that occurs independently of both PKA and ARPP19 phosphorylation at serine 109 but requires the Greatwall-dependent phosphorylation of ARPP19 at serine 67. Therefore, ARPP19 stands at a crossroads in the meiotic M-phase control network by integrating differential effects of PKA and Greatwall, two essential kinases for meiosis resumption.

Entry into mitosis or meiosis relies on the coordinated action of kinases and phosphatases that ultimately leads to the activation of Cyclin B-Cdk1, also called MPF for M-phase promoting factor. Vertebrate oocytes are blocked in prophase of the first meiotic division, an arrest tightly controlled by a high PKA activity. Reentry into meiosis depends on Cdk1 activation that obeys a two steps mechanism: a catalytic amount of Cdk1 is generated in a PKA and protein synthesis-dependent manner; then a regulatory network called MPF auto-amplification loop is initiated. This second step is independent of PKA and protein synthesis. However, none of the molecular components of the auto-amplification loop identified so far acts independently of PKA. Therefore, the protein rendering this process independent of PKA in oocytes remains unknown. Using a physiological intact cell system, the Xenopus oocyte, we show that the phosphorylation of ARPP19 at S67 by the Greatwall kinase promotes its binding to the PP2A-B55 δ phosphatase, thus inhibiting its activity. This process is controlled by Cdk1 and plays an essential role within the Cdk1 auto-amplification loop for entry into the first meiotic division. Moreover, once phosphorylated by Greatwall, ARPP19 escapes the negative regulation exerted by PKA. It also promotes MPF activation independently of protein synthesis, provided a small amount of Mos is present. Taken together, these findings reveal that PP2A-B55δ, Greatwall and ARPP19 are not only required for entry into meiotic divisions, but are also pivotal effectors within the Cdk1 auto-regulatory loop responsible for its independence toward PKA negative control.