p21 Inhibits Thr(161) phosphorylation of Cdc2 to enforce the G(2) DNA damage checkpoint

The cyclin-dependent kinase inhibitor p21 is required for a sustained G(2) arrest after activation of the DNA damage checkpoint. Here we have addresse d the mechanism by which p21 can contribute to this arrest in G(2). We show that p21 blocks the activating phosphorylation of Cdc2 on Thr(161). p21 do es not interfere with the dephosphorylation of two inhibitory phosphorylati on sites on Cdc2, Thr(14) and Tyr(15), indicating that p21 targets a differ ent event in Cdc2 activation as the well described DNA damage checkpoint pa thway involving Chk1 and Cdc25C. Taken together our data show that a cell i s equipped with at least two independent pathways to ensure efficient inhib ition of Cdc2 activity in response to DNA damage, influencing both positive and negative regulatory phosphorylation events on Cdc2.