Grant: Discovery Grants Program - Individual

The cell division process is tightly regulated and closely coordinated with copying of its genetic material (DNA). In mammalian cells, the genome must replicate only once in its entirety during a single cell division cycle. Derangement of this process can be fatal for the cell or result in genetic instability. The once-per-cell cycle replication is achieved mainly by strictly regulating the stepwise formation and activation of pre-replication complex (pre-RC). At final stage of this regulation, Cdc7 protein functions as a 'molecular switch' for the activation of DNA replication by phosphorylating critical licensing components of the pre-RC. It is well known that Cdc7 itself is regulated by Dbf4 protein. However, it is not well-understood the mechanism how Cdc7 and the Dbf4 regulatory subunit bind each other to activate the pre-RC at the replication start-site. Our recent data shows that the Cdk1-mediated Cdc7 phosphorylation causes an increase in Cdc7 affinity to CRM1 protein. As a result, phosphorylated Cdc7 is dissociated from replication start-sites and bound by CRM1, by which Cdc7 is sequestered in the cytoplasm. Thus, Cdc7 phosphorylation prevents DNA re-replication during G2-M, which is critical for maintaining genetic stability. As a cell progresses from G1 to S phase during the next round of cell division cycle, the function of Cdc7 as replication activator is 'restored' by removing its phosphate group by an enzyme called phosphatase. In effect, the enzyme responsible for Cdc7 dephosphorylation functions as a key molecular switch toward the activation of DNA replication by Cdc7-Dbf4. Our main objective is to identify and characterize this important dephosphorylation enzyme. In addition, we also plan to determine whether phosphorylated Cdc7 has other functions. Data from this proposed work will shed exciting new light on the regulation of DNA replication in the context of cell cycle progression. Our data will also provide tremendous insight into the control mechanism about how a cell maintains its genetic stability. If successful, this work can have very significant and lasting impact on the DNA replication and cell cycle research fields.