The long-term goal of this proposal is to investigate the molecular mechanism of eukaryotic DNA replication. The lower eukaryotic organism, yeast, will be used as the model system. An active component of yeast DNA replication, CDC8, is selected for further study. We have analyzed the CDC8 gene and identified its gene product as yeast thymidylate kinase. We will further examine the subcellular localization of the CDC8 protein and investigate its functions through the cell division cycle. The relationship between the CDC8 protein and other replication components will be explored. The result will be compared to the functions of other eukaryotic replication systems. This study will provide information about the temporal and spatial events of yeast replication components and the mechanism of action of the CDC8 in DNA replication. Genetic approach to the roles of CDC8 will further substantiate the above studies. A CDC8 suppressor (SOC8-1) will also be studied. The gene and its gene product will be characterized, and the result will be integrated into information from the CDC8 studies, providing us information to deduce the suppression mechanism and role of CDC8 in DNA replication. We believe that this research will contribute to our understanding of the metabolic coordination of DNA precursor enzymes and the complex processes involved in the propagation and regulation of yeast DNA replication. We expect that the elucidation of yeast enzymes will lead to understanding the functions of corresponding enzymes in other eukaryotic systems.
