Genetic dissection of virion maturation and epithelial cell tropism in human cytomegalovirus

Human cytomegalovirus is the archetypal member of the Betaherpesvirinae subfamily. Cytomegalovirus normally persists for the life of its host as an asymptomatic, latent infection in hematopoietic progenitor cells of the myeloid lineage. While pathological manifestations in adults are restricted solely to immuno-compromised populations, congenital and neonatal cytomegalovirus infections routinely impact their hosts with severe complications. Congenital infection is the leading infectious cause of birth defects, and neonatal infection most frequently results in severe hearing loss. To cause these pathological outcomes, infection must spread from the site of introduction to various organs and tissues throughout the host, thereby obligating cytomegalovirus to complete its lytic infectious cycle in a variety of cell types. These include epithelial cells, fibroblasts, smooth muscle cells, and endothelial cells, each of which facilitates viral replication in a different intracellular milieu. Epithelial cells, in particular, serve an important role during in vivo infection, as mucosal epithelia are thought to be involved in the establishment of infection, while ductal and secretory epithelia have been implicated in the dissemination of infectious viral progeny into the environment. Employing a genome-wide genetic screen in a clinical isolate of cytomegalovirus, I identified the viral genes RL4/5, UL39, UL83, UL132, UL148, and US22, as well as a group of contiguous genes spanning the UL109-111a loci, as novel determinants of epithelial cell tropism.;Also underlying the in vivo spread of cytomegalovirus infection are the cytoplasmic processes of viral assembly and egress. These cytoplasmic processes are coordinated in the viral Assembly Compartment, a dense, juxtanuclear collection of membranes, virion proteins and cellular proteins that serves as the site of final virion tegumentation and envelopment. In the absence of efficient virion assembly and egress, the spread of infection to neighboring cells and tissues is severely impared. Furthermore, dissemination of cytomegalovirus throughout the host population requires the release of extracellular infectious viral progeny. I have identified the function of cytomegalovirus protein UL71, a previously uncharacterized component of the virion tegument, to be involved in the cytomegalovirus-induced reorganization of host cell membranes that results in the formation of the viral Assembly Compartment. Additionally, I have found pUL71 to be required for efficient viral trafficking and for delaying the formation of large intracytoplasmic inclusions that are believed to function in the degradation of cytoplasmic virions at late times post infection.;Together these findings contribute to the overall understanding of human cytomegalovirus replication and, in particular, specific aspects of the replication cycle that are required for the in vivo spread of infection. Based on the hypothesized importance of epithelial cells to the intra- and inter-host spread of infection, we believe therapies that target host or viral constituents of pathways required for epithelial cell tropism could prevent the in vivo spread of infection, as well as the dissemination of the virus throughout the host population. Likewise, because of the large defect in viral spread observed when the processes of assembly and egress are disrupted, we believe there may be clinical benefit from the development of therapies that target host and viral constituents of these pathways.