ACTIVATED POLO-LIKE KINASE PLX1 IS REQUIRED AT MULTIPLE POINTS DURINGMITOSIS IN XENOPUS-LAEVIS

Entry into mitosis depends upon activation of the dual-specificity pho sphatase Cdc25C, which dephosphorylates and activates the cyclin B-Cdc 2 complex. Previous work has shown that the Xenopus polo-like kinase P lx1 can phosphorylate and activate Cdc25C in vitro. In the work presen ted here, we demonstrate that Plx1 is activated in vivo during oocyte maturation with the same kinetics as Cdc25C. Microinjection of wild-ty pe Plx1 into Xenopus oocytes accelerated the rate of activation of Cdc 25C and cyclin B-Cdc2. Conversely, microinjection of either an antibod y against Plx1 or kinase-dead Plx1 significantly inhibited the activat ion of Cdc25C and cycliu B-Cdc2. This effect could be reversed by inje ction of active Cdc25C, indicating that Plx1 is upstream of Cdc25C. Ho wever, injection of Cdc25C, which directly activates cyclin B-Cdc2, al so caused activation of Plx1, suggesting that a positive feedback loop exists in the Plx1 activation pathway. Other experiments show that in jection of Plx1 antibody into early embryos, which do not require Cdc2 5C for the activation of cyclin B-Cdc2, resulted in an arrest of cleav age that was associated with monopolar spindles. These results demonst rate that in Xenopus laevis, Plx1 plays important roles both in the ac tivation of Cdc25C at the initiation of mitosis and in spindle assembl y at late stages of mitosis.