Calcium/calmodulin-dependent phosphorylation and activation of human cdc25-c at the G(2)/M phase transition in HeLa cells

The human tyrosine phosphatase (p54(cdc25-c)) is activated by phosphorylati on at mitosis entry. The phosphorylated p54(cdc25-c) in turn activates the p34-cyclin B protein kinase and triggers mitosis. Although the active p34-c yclin B protein kinase can itself phosphorylate and activate p54(cdc25-c), we have investigated the possibility that other kinases may initially trigg er the phosphorylation and activation of p54(cdc25-c). We have examined the effects of the calcium/calmodulin-dependent protein kinase (CaM kinase II) on p54(cdc25-c). Our in vitro experiments show that CaM kinase II can phos phorylate p54(cdc25-c) and increase its phosphatase activity by 2.5-3-fold. Treatment of a synchronous population of HeLa cells with KN-93 (a water-so luble inhibitor of CaM kinase II) or the microinjection of AC3-I (a specifi c peptide inhibitor of CaM kinase II) results in a cell cycle block in G(2) phase. In the KN-93-arrested cells, p54(cdc25-c) is not phosphorylated, p3 4(cdc2) remains tyrosine phosphorylated, and there is no increase in histon e H1 kinase activity. Our data suggest that a calcium-calmodulin-dependent step may be involved in the initial activation of p54(cdc25-c).