INTERACTIONS AMONG REPLICATION AND MUTATION PROTEINS

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Genetic damage by environmental and endogenous agents can lead to birth defects, heritable diseases, cancer, and possibly contributes to aging. Thus, understanding how mutations are produced is fundamental to understanding and, perhaps, preventing many human disorders. Mutations are created when damaged DNA fails to be repaired, but, instead, is replicated. The long-term goal of this research is to understand what happens When the DNA replication complex encounters a DNA lesion, and to elucidate what cellular factors determine whether DNA repair or mutation prevails. In Escherichia coli, DNA damage induces about 20 proteins, which together are called the SOS response. The research proposed will investigate the interactions between the DNA replication complex and RecA, UmuD, and UmuC, the SOS proteins that are required for mutagenic replication past DNA lesions. This mutagenic activity is mediated by protein-protein interactions, probably resulting in a transient modification of the replication complex, DNA polymerase III holoenzyme (Pol III HE). The proofreading activity of Pol III HE is performed by a distinct subunit, epsilon. High cellular levels of epsilon inhibit SOS mutagenesis, but this inhibition can be relieved by concurrent overproduction of UmuDC. These and other results suggest that the SOS proteins may interact with epsilon as part of their alteration of Pol III HE. There are three specific aims of this research. (i) to identify any interactions between epsilon and the SOS proteins and to determine the functional significance of any interaction; (2) to generate and characterize new defects in the replication subunits that interfere with SOS mutagenesis; and (3) to investigate if the SOS proteins interact with replication subunits other than epsilon. These studies will elucidate the complex interactions that take place when damaged DNA is replicated and may provide a model for similar processes occurring in higher organisms.