The long term objectives of the proposed research ar to understand mechanisms of DNA repair and mutagenesis. The cellular process of DNA repair is important in maintenance of genetic information in the cell for both expression and inheritance. It is clear that some cellular processes promote mutagenesis, perhaps to allow cell survival, while other processes appear to prevent mutagenesis. Cells have apparently evolved processes which allow survival with some toleration of mutagenesis. We have demonstrated that a group of mutants of E. coli which can survive without a functional DNA polymerase III shows striking alterations in DNA repair and mutagenesis. These cells offer an unique opportunity to study these polymerase I is supplying critical replicative functions. We will take advantage of these cells by development of an in vitro system for repair synthesis in which the holoenzyme containing DNA polymerase I is functional. We will investigate the interactions of the replicative complex with the RecA and UmuCD proteins to analyze bypass synthesis. We will also use the in vitro synthesis system to analyze response to MMS and hydrogen peroxide damage, two types of damage to which these cells are particularly sensitive. We will also focus on the uvrC gene and its regulation. It is not clear if this gene product supplied other functions in addition to repair. We will analyze the regulation and function of the uvrC gene product by a combination of genetic and recombinant DNA technology.
