The cell cycle is a universal process by which cells grow, propagate and differentiate. Cell cycle transitions are controlled by cyclin-dependent kinases (CDKs), which in turn are tightly regulated by protein-protein interactions and phosphorylations. Checkpoint mechanisms maintain proper cell cycle progression and genetic stability. One of the mechanisms that regulates CDKs is through inhibitory phosphorylation of two residues within the catalytic cleft. I showed that inhibitory phosphorylations of CDC2 controlled the timing of their activities in the unperturbed cell cycle. Expression of non-phosphorylatable mutants of CDC2 or CDK2 was cytotoxic. The non-phosphorylatable mutants bypassed the G₂ DNA damage checkpoint and induced histone H3 phosphorylation. Moreover, expression of non-p...[ Read more ]

The cell cycle is a universal process by which cells grow, propagate and differentiate. Cell cycle transitions are controlled by cyclin-dependent kinases (CDKs), which in turn are tightly regulated by protein-protein interactions and phosphorylations. Checkpoint mechanisms maintain proper cell cycle progression and genetic stability. One of the mechanisms that regulates CDKs is through inhibitory phosphorylation of two residues within the catalytic cleft. I showed that inhibitory phosphorylations of CDC2 controlled the timing of their activities in the unperturbed cell cycle. Expression of non-phosphorylatable mutants of CDC2 or CDK2 was cytotoxic. The non-phosphorylatable mutants bypassed the G₂ DNA damage checkpoint and induced histone H3 phosphorylation. Moreover, expression of non-phosphorylatable CDC2 induced DNA replication and cytolunesis in the presence of DNA damage. Thus, inhibitory phosphorylations of CDC2 and CDK2 may play different role in the normal cell cycle but both are important for the DNA damage checkpoint. In a parallel study, I examined the DNA damage responses during the activation of spindle-assembly checkpoint. DNA damage during mitotic block reversed cells back to a G₂ DNA damage checkpoint-like state, including CDC2 inactivation, histone H3 dephosphorylation, and chromosome decondensation. The mitotic DNA damage responses were dependent on the Ataxia Telangiectasia mutated kinase ATM. Ectopic expression of CDC25A and nonphosphorylatable CDC2 inhibited histone H3 dephosphorylation after DNA damage. Thus, the signal from DNA damage could overcome the signal from spindle-assembly checkpoint. These studies demonstrated that inhibitory phosphorylation sites of CDC2 and CDK2 play essential roles in the precise regulation of cell cycle and checkpoints.