The overall goal of this research proposal is to examine in a comprehensive fashion, the structural properties of the bovine papillomavirus (BPV) E2 protein that mediate its transcription functions. The E2 gene product is central to two pivotal events in the viral like cycle. E2 coordinates transcription and is necessary for viral E1 protein. The 410 amino acid E2 protein has distinct domains: it's amino-terminal third forms a dimer that binds a specific papillomavirs DNA sequence with high affinity. The first specific aim of the proposal involves continued characterization of the structure and function of the E2 protein's carboxy terminus. The specific amino acids that mediate nucleic acid recognition and dimerization as well as a newly-identified function needed for transactivation represent the focus of attention. The second specific aim has as its goal the genetic characterization of the structure and functions of the E2 amino terminus in transactivation and, to a limited extent viral DNA replication and E1 binding. Experiments proposed in aim 3 use the results and resources provided by the experiments in the first two aims to identify cellular proteins that bind E2 and to determine how E2 functions are regulated in the cell. For example, biochemical and mutational studies of the cysteine located in the center of the DNA contact region and conserved among papillomavirus E2 proteins suggest that its role may be to control E2 activity perhaps through a nuclear localization pathway. The investigations proposed in this application are designed to elucidate the molecular mechanisms through which E2 stimulates transcription, binds DNA, and interacts with cellular transcription factors and thereby provide insight into the papillomavirus life cycle as well as into basic methods of controlling gene expression.
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