Periodic expression of the cyclin-dependent kinase inhibitor p57(Kip2) in trophoblast giant cells defines a G2-like gap phase of the endocycle

Endoreduplication is an unusual form of cell cycle in which rounds of DNA s ynthesis repeat in the absence of intervening mitoses. How G1/S cyclin-depe ndent kinase (Cdk) activity is regulated during the mammalian endocycle is poorly understood. We show here that expression of the G1/S Cdk inhibitor p 57(Kip2) is induced coincidentally with the transition to the endocycle in trophoblast giant cells. Kip2 mRNA is constitutively expressed during subse quent endocycles, but the protein level fluctuates. In trophoblast giant ce lls synchronized for the first few endocycles, the p57Kip2 protein accumula tes only at the end of S-phase and then rapidly disappears a few hours befo re the onset of the next S-phase. The protein becomes stabilized by mutatio n of a C-terminal Cdk phosphorylation site. As a consequence, introduction of this stable form of p57(Kip2) into giant cells blocks S-phase entry. The se data imply that p57(Kip2) is subject to phosphorylation-dependent turnov er. Surprisingly, although this occurs in endoreduplicating giant cells, p5 7(Kip2) is stable when ectopically expressed in proliferating trophoblast c ells, indicating that these cells lack the mechanism for protein targeting and/or degradation. These data show that the appearance of p57(Kip2) punctu ates the completion of DNA replication, whereas its turnover is subsequentl y required to initiate the next round of endoreduplication in trophoblast g iant cells. Cyclical expression of a Cdk inhibitor, by terminating G1/S Cdk activity, may help promote the resetting of DNA replication machinery.