Herpesvirvses are significant human pathogens and cause a variety of disease syndromes which range in severity from inapparent infection to life-threatening disease. A hallmark of herpesvirvses is their ability to establish persistent infection. This ability is especially evident in diseases such as oral or genital herpes simplex virus infections, in which periods of latency are punctuated with episodes of active, localized lesions. Very little is known about the alterations in virus/cell interactions which result in herpesviral persistent infection rather than productive, lytic infection. As a model system for persistent infection by a cytocidal herpesvirus, persistently infected/oncogenically transformed cell lines were developed by infection of primary hamster embryo cells with equine herpesvirus type I (EHV-1) preparations rich in defective interfering particles (DIPs). Small populations (<20%) of the cells continuously produce both standard (STD) EHV-1 and DIPs; the remaining cells do not produce viral particles, but have been shown to contain portions of the EHV-1 genome. Utilizing this model system, the overall goal of the current research is to provide an understanding of the molecular biology of persistent infection by examining the regulation of viral gene expression and characterizing the structure and function of viral proteins in persistently infected cells as compared to STD EHV-1 infected and DIP-rich infected cells. Specifically, the aims are to: 1) characterize the structure and function of the EHV-1 immediate early proteins (IEPs) and define further the alterations in their regulation, synthesis, and localization; 2) determine the location and function of the virion protein, ICP130, and define the mechanism for its reduced synthesis in DIP-rich infected and persistently infected cells; and 3) characterize persistently infected and DIP-rich infected cells for other deviations from STD EHV-1 lytic infection patterns. Methods include: development of anti-IEP monoclonal antibodies and monospecific antisera against other viral proteins, radioisotopic cell labeling, immunoprecipitation and immunoblotting assays, immunofluorescence studies, phosphorylation and DNA-binding assays, in vitro translation, transcriptional mapping, plasmid construction, and transient expression assays. These studies will yield information not only about herpesviral gene regulation, but also about the functions of critical viral gene products in determining the outcome of herpesvirus infections.
