ASSEMBLY AND FUNCTION OF THE HERPES SIMPLEX ENVELOPE

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Herpes viruses are associated with a range of human disease; Burkitts lymphoma and nasopharyngeal carcinoma (Epstein-Barr virus), cytomegalic inclusion disease (Cytomegalovirus), chickenpox and shingles (Varicella- Zoster virus) and oral or genital mucocutaneous lesions (Herpes simplex virus, HSV types 1 and 2). Their ability to remain latent, then reactivate means they present a serious source of infection following immune suppression in organ transplant, chemotherapy of AIDS patients. The recent appearance of acyclovir-resistant HSV strains in CD4-suppressed subjects underlines the importance of developing new therapeutic strategies to combat herpes virus infections. Little is known of how herpes viruses enter and leave cells. Envelope/plasma membrane fusion during infection requires a large number of viral proteins, and the molecular role of each is unknown. Similarly, the mechanism by which newly assembled progeny capsids become enveloped then released from the cell remains unclear. The long term objective of these studies is a molecular dissection of these phenomena, revealing much about the basic biology of the herpes viruses and potentially yielding a wealth of new virus-specific drug targets. The specific aims are as follows: (1). To investigate the kinetics and morphology of HSV-1 egress from infected cells. A single, synchronized wave of egressing virions will be assayed for DNA packaging, envelopment, acquisition of infectivity and rate of secretion from the cell. Antibodies against the viral capsid will be used to correlate these events with morphological phenomena. (2). To determine the origin of the HSV-1 envelope. The essential HSV glycoprotein gH will be targeted to organelles within HSV infected cells to identify the site of envelopment. Passage of virions through subcellular compartments will be monitored and the kinetics of egress determined. (3). To characterize the defect in the gH mutant tsQ26. Pulse chase experiments will be used to test the hypothesis that the tsQ26 allele of gH is unable to traffic to the site of HSV envelopment at the nonpermissive temperature. (4). To identify regions of glycoprotein gH required for fusion. Regions of the glycoprotein required for envelope/plasma membrane fusion and cell/cell fusion will be determined using a transient expression/complementation assay.