Herpes simplex virus (HSV) is a human pathogen that is capable of both rapid productive infections, which lead to destruction of the infected host cells, and latent infections where the viral genome is relatively silent, persisting in the neurons of sensory ganglia for many years. HSV specifies a number of transacting proteins that alter the regulatory pathways and transcriptional machinery of the cell. The ICP4 protein of HSV is required for viral growth and is involved in the regulated transcription of nearly every viral gene. Like many transcriptional regulatory proteins, the ICP4 protein of herpes simplex virus (HSV) is modular in composition, consisting of multiple discrete domains that collectively contribute to its function during viral infection. The proposed research program includes the identification of functional domains of the ICP4 molecule in the context of viral mutants, the isolation of mutant proteins, and the characterization of the mutant proteins with respect to activity in vitro, phosphorylation, DNA binding and the interaction with cellular transcription factors. Given our knowledge of the domain composition of ICP4, one aim of this proposal is to generate and study mutants of the ICP4 protein that specify a subset of functional domains and are transdominant inhibitors of viral growth. We will explore inhibitory effects of such proteins in vivo by constructing transgenic mice that express transdominant mutants in response to infection. The experiments with transgenic mice will also help determine the stage/s in the pathogenesis of HSV most susceptible to intervention through disruption of the interactions discussed above. Another aspect of the research is dedicated to examining the relevant signal transduction pathways which affect transcriptional regulation in infected cells in lytic and latent infection. this involves exploring the interaction of specific cellular protein kinases with specific sites in ICP4 and the consequences the ensuing modifications on activity, molecular interactions and pathogenesis. Enclosed in a research plan that was previously reviewed and funded, which contains a subset of the experiments comprising the total research program. The immediate goals of the program are to develop a comprehensive understanding of transcriptional regulation in HSV infection. The long term goals of the program are to develop an understanding of the various mechanisms by which HSV interacts with host cells resulting in the alteration of host cell macromolecular metabolism. This understanding may allow for the rational derivation of novel antivirals.
