THE MAIN AREA OF FOCUS of the laboratory is the regulation of the G1 to S phase transition of the cell cycle in yeast and human cells. Most cells coordinate growth and division in this phase of the cell cycle. Elucidation of the mechanisms of cell cycle control and cell commitment to DNA replication is important for determining the etiology of a number of diseases, especially cancer, in which the regulation is altered. Yeast (Saccharomyces cerevisiae), as an eukaryotic microorganism is an excellent model system to study the cell cycle because facile molecular genetic techniques can be used in combination with classical biochemical and genetic methods. We use human cells in culture as a way to study the defects that are present in the cell cycle of cancer cells.
Yeast: Our current studies focus on the regulation of the yeast Cdc7/Dbf4 protein kinase, which regulates the initiation of DNA replication during the somatic cell cycle. Our current model is that Cdc7 kinase is activated by binding the Dbf4 protein during the G1 to S phase transition (Sclafani, 2000). Dbf4 protein levels are regulated by Rad53 protein kinase, which is also involved in DNA-checkpoint regulation, and by the APC (anaphase promotion complex). Active CDC7 kinase then phosphorylates the MCM complex, a multi-subunit protein complex found at origins of DNA replication, to initiate the S phase. Both Cdc7 and Cdk1 proteins are needed for mtuliple steps during initiation (Sclafani et al., 2002). We have begun structure-function studies of the MCM complex with Dr. X. Chen by comparing Archaea MCM to Yeast MCM (Fletcher et al., 2003; Sclafani et al., 2004).
We have also defined a role of Cdc7/Dbf4 kinase in error-prone tyranslesion synthesis, which is important for mutagenesis (Pessoa-Brandão and Sclafani, 2004).
Humans: Our human studies are focused on the deregulation of the cell cycle which occurs in cancer cells. We are determining the mode of action of several drugs found in natural products that function in cancer chemoprevention. We have shown that the target of silibinin from Milk Thistle is CDK-inhibitors (Agarwal et al., 2003). We are also investigating the action of drugs that activate the DNA damage checkpoint pathway such as Reseveratrol from red grape skins/wine (Tyagi et al., 2005). These studies are in collaboration with Dr. Rajesh Agarwal of the UCHSC School of Pharmacy.