The survival of p53 defective tumor cells to agents that inhibit DNA replication or damage DNA may be largely dependent on checkpoints that regulate the onset of mitosis. The mitosis inducing kinase, Cdc2/Cyclin B, is inhibited by phosphorylation of threonine-14 and tyrosine-15. Disruption of these phosphorylation sites abrogates checkpoint-mediated regulation of Cdc2 and renders tumor cells highly sensitive to killing by DNA damage or by inhibition of DNA replication. These findings establish the importance of inhibitory phosphorylation of Cdc2 in G2 checkpoint control and in the survival mechanism used by human cells when exposed to some of the most common forms of anti-cancer therapy. A molecular understanding of this checkpoint is essential as the rationale basis for the development of therapies that target the approximately 50 percent of human cancers that are defective for p53 function. The phosphorylation state of Cdc2 is controlled by the opposing activities of the Wee1/Myt1 kinases and the Cdc25 phosphatase. The regulation of these enzymes is undoubtedly crucial for the functioning of the G2 checkpoint, however, a direct effect on the activity of any of these enzymes has not been reported. We have evidence suggesting that Cdc25 activity is down-regulated in vivo following irradiation and that a similar reduction in Cdc25 activity is seen following phosphorylation by Chk1 in vitro. Furthermore the radiation induced down-regulation of Cdc25 is dependent on the product of the Ataxia Telangiectasia gene. These observations provide the framework of a model by which DNA damage might delay mitotic progression in human cells. The primary objective of this proposal is therefore to provide a detailed understanding of the regulation of Cdc25, Wee1, Myt1 and Chk1 in human checkpoint control responses.
| Scorah, Jennifer; McGowan, Clare H (2009) Claspin and Chk1 regulate replication fork stability by different mechanisms. Cell Cycle 8:1036-43 |
| MacLaren, Ann; Slavin, Daniela; McGowan, Clare H (2009) Chk2 protects against radiation-induced genomic instability. Radiat Res 172:463-72 |
| Scorah, Jennifer; Dong, Meng-Qiu; Yates 3rd, John R et al. (2008) A conserved proliferating cell nuclear antigen-interacting protein sequence in Chk1 is required for checkpoint function. J Biol Chem 283:17250-9 |
