MITOTIC AND G(2) CHECKPOINT CONTROL - REGULATION OF 14-3-3-PROTEIN-BINDING BY PHOSPHORYLATION OF CDC25C ON SERINE-216

Human Cdc25C is a dual-specificity protein phosphatase that controls e ntry into mitosis by dephosphorylating the protein kinase Cdc2. Throug hout interphase, but not in mitosis, Cdc25C was phosphorylated on seri ne-216 and bound to members of the highly conserved and ubiquitously e xpressed family of 14-3-3 proteins. A mutation preventing phosphorylat ion of serine-216 abrogated 14-3-3 binding. Conditional overexpression of this mutant perturbed mitotic timing and allowed cells to escape t he G(2) checkpoint arrest induced by either unreplicated DNA or radiat ion-induced damage. Chk1, a fission yeast kinase involved in the DNA d amage checkpoint response, phosphorylated Cdc25C in vitro on serine-21 6. These results indicate that serine-216 phosphorylation and 14-3-3 b inding negatively regulate Cdc25C and identify Cdc25C as a potential t arget of checkpoint control in human cells.