FUNCTIONAL-ANALYSIS OF THE P-BOX, A DOMAIN IN CYCLIN-B REQUIRED FOR THE ACTIVATION OF CDC25

Cyclin B-cdc2 complexes are kept inactive by inhibitory phosphorylatio ns on Thr-14 and Tyr-15 of cdc2 until they are dephosphorylated at the end of G2 by the phosphatase cdc25. Recent work has suggested that a small region of cyclin B, which we call the P box, may contribute part of a phosphatase-activating domain to cdc25. Individual conservative substitutions at three invariant residues within the P box yield mutan t cyclin 8 proteins that bind cdc2 in vitro and then show the predicte d cell cycle arrest, with cdc25 remaining in the low activity interpha se form and cyclin B-cdc2 complexes remaining phosphorylated and inact ive. While the low activity interphase form of cdc25 cannot act on cdc 2 complexed with a mutant P box cyclin, the high activity M phase form of cdc25 can. These results demonstrate that the P box domain of cycl in B is required for cdc25 activation and support a two-step mechanism for the cdc25-dependent activation of cyclin B-cdc2.