Herpesvirus 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 herpesviruses in their ability to establish persistent infection. This ability is especially evident in diseases such as oral or genital herpes simplex 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, cytocidal infection. As a model for persistent infection, transformed hamster cell lines have been obtained by infection with equine herpesvirus type 1 (EHV-1) preparations rich in defective interfering (DI) particles; small populations (3-20%) of the cells continuously produce both standard EHV-1 and DI particles. The remaining majority of cells do not produce viral particles but have been shown to contain portions of the EHV-1 genome and to express viral antigens. Inoculation of these cell lines into syngeneic hamsters result in tumor production, and the resulting tumor cell lines are virus nonproducers. The overall goal of this proposal is to gain insight into the molecular biology of persistent infection by comparing the regulation of viral protein synthesis in these persistently infected cells to that in cytocidal EHV-1 infection and in cells infected with DI-rich virus. Specifically, the aims are to: 1) further define the alpha/beta/gamma temporal regulation of viral protein synthesis in EHV-1 cytocidal infection by using metabolic inhibitors such as cyclohexamide, actinomycin D, Ara C during infections; 2) characterize, through the use of isotopic labeling, SDS-PAGE, immunoprecipitation and immunoblotting, the proteins comprising each class with respect to physical properties (glycosylation, phosphorylation), temporal appearance, stability, cellular localization, and function; 3) examine DI-rich infections by the same methods to detect any aberrant regulation (underproduced, overproduced, novel proteins); 4) characterize viral proteins produced in persistently infected cells to discern any altered regulation patterns; and 5) identify and characterize viral proteins expressed in cloned virus non-producer transformed cells and in tumor cells to gain information as to which viral gene products may be necessary for transformation versus persistent infection.
