Regulation of Cdc2/cyclin B activation in Xenopus egg extracts via inhibitory phosphorylation of Cdc25C phosphatase by Ca(2+)/calmodulin-dependent protein [corrected] kinase II.
Activation of Cdc2/cyclin B kinase and entry into mitosis requires dephosphorylation of inhibitory sites on Cdc2 by Cdc25 phosphatase. In vertebrates, Cdc25C is inhibited by phosphorylation at a single site targeted by the checkpoint kinases Chk1 and Cds1/Chk2 in response to DNA damage or replication arrest. In Xenopus early embryos, the inhibitory site on Cdc25C (S287) is also phosphorylated by a distinct protein kinase that may determine the intrinsic timing of the cell cycle. We show that S287-kinase activity is repressed in extracts of unfertilized Xenopus eggs arrested in M phase but is rapidly stimulated upon release into interphase by addition of Ca2+, which mimics fertilization. S287-kinase activity is not dependent on cyclin B degradation or inactivation of Cdc2/cyclin B kinase, indicating a direct mechanism of activation by Ca2+. Indeed, inhibitor studies identify the predominant S287-kinase as Ca2+/calmodulin-dependent protein kinase II (CaMKII). CaMKII phosphorylates Cdc25C efficiently on S287 in vitro and, like Chk1, is inhibited by 7-hydroxystaurosporine (UCN-01) and debromohymenialdisine, compounds that abrogate G2 arrest in somatic cells. CaMKII delays Cdc2/cyclin B activation via phosphorylation of Cdc25C at S287 in egg extracts, indicating that this pathway regulates the timing of mitosis during the early embryonic cell cycle.
D000595:Amino Acid Sequence; D000818:Animals; D001381:Azepines / Q000494:pharmacology; D042846:CDC2-CDC28 Kinases / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism*; D000071877:Checkpoint Kinase 1; D064447:Checkpoint Kinase 2; D003001:Cloning, Molecular; D019926:Cyclin B / Q000378:metabolism*; D004249:DNA Damage; D004261:DNA Replication; D005260:Female; D016195:G2 Phase / Q000502:physiology; D007399:Interphase; D008938:Mitosis; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009865:Oocytes / Q000201:enzymology*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011758:Pyrroles / Q000494:pharmacology; D019311:Staurosporine / Q000031:analogs & derivatives* / Q000494:pharmacology; D029867:Xenopus Proteins; D014982:Xenopus laevis; D020687:cdc25 Phosphatases / Q000378:metabolism*
D001381:Azepines; D018797:Cell Cycle Proteins; D019926:Cyclin B; D011758:Pyrroles; D029867:Xenopus Proteins; C097660:debromohymenialdisine; C054852:7-hydroxystaurosporine; D011494:Protein Kinases; D064447:Checkpoint Kinase 2; D000071877:Checkpoint Kinase 1; C000605875:Chek1 protein, Xenopus; D017346:Protein Serine-Threonine Kinases; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D042846:CDC2-CDC28 Kinases; D020687:cdc25 Phosphatases; D019311:Staurosporine; D002118:Calcium