| The mammalian cell cycle consists of a series of highly organized steps that ensure the faithful replication of DNA and the production of two identical progeny or daughter cells. Ordered progression through the cell cycle is governed by key regulatory elements, the CDKs and cyclins. The discovery of the CDKs or c&barbelow;yclin d&barbelow;ependent k&barbelow;inases in the early 1990s has enabled scientists to begin to delineate the specific steps required for the transition from G1 to S phase of the cell cycle, and thus cell proliferation. Upon mitogenic stimulation, CDK2, which is essential for normal cell cycle progression, is activated in late G1. Many aspects of CDK2 regulation are well established including the effects of a variety of post-translational events including cyclin binding, phosphorylation and dephosphorylation and association with inhibitory protein. However, the regulation of the enzymes that govern these regulatory events is not known, and requires further investigation for a more complete understanding of both the function and regulation of CDK2.; This dissertation is concerned with the mitogenic activation, regulation and function of CDK2. In this dissertation, we further elucidate the regulation of CDK2 necessary for progression from late G1 to S phase of the cell cycle. Specifically, we show that ERK activation regulates the nucleocytoplasmic translocation of CDK2. Furthermore, while nucleocytoplasmic translocation of CDK2 is necessary, it is not sufficient for CDK2-cyclin E activation. Growth factor stimulation is also necessary for CAK activity and phosphorylation of CDK2 at Thr160. Surprisingly, while the nuclear activation of CDK2-cyclin E is sensitive to inhibition of MEK activation of ERK, the activational phosphorylation of Thr160 is independent of ERK. Finally, we provide evidence that CDK2-cyclin E activity results in Rb inactivation by phosphorylation. Interestingly, the actions of CDK2-cyclin E do not require prior substrate phosphorylation by CDK4-cyclin D1. |
