Human cytomegalovirus (HCMV) is a significant human pathogen that infects the majority of the world's population. Primary lytic replication or reactivation of latent infections causes severe disease in patients with suppressed immune function. Furthermore, many studies link HCMV infection with proliferative diseases such as cancer, restenosis, and transplant-associated vasculopathy. Because the HCMV pp71 protein initiates the lytic replication cycle, stimulates cell cycle progression, and may control latency, it is a potential target for antiviral therapy. We plan to examine the molecular mechanisms behind the different activities of pp71, and the role that these different activities play during lytic and latent viral replication. We have already discovered and published the mechanism through which pp71 activates viral immediate early (IE) gene expression during lytic replication: it degrades the cellular transcriptional represser Daxx. Here we will generate HCMV mutants that are resistant to Daxx-mediated repression, and use them to further our understanding of IE gene expression during lytic infection, and determine if the same mechanism applies during the establishment and/or reactivation of latency. We have previously shown that pp71 stimulates cell cycle progression by degrading the Rb protein. Here we will examine how pp71 and other viral proteins cooperate to modulate the activity of Rb (and the related p130 protein) during lytic and latent infections. Recently, we discovered that pp71 degrades another cellular protein that modulates transcription, the cell cycle, and innate immunity. Here we will determine how the ability of pp71 to degrade this protein affects each of those processes in HCMV-infected cells. This proposal builds upon our published and unpublished discoveries, and will use molecular and genetic approaches to determine not only the mechanisms used by pp71 to promote viral transcription, induce cell cycle progression, and potentially evade innate immunity, but also the roles that these activities of pp71 play during both lytic and latent infections. Thus our long-term goal is to determine the molecular mechanisms behind each function of pp71 in HCMV-infected cells, and the contribution that the different activities of pp71 make during lytic and latent viral replication cycles. This information should allow us to design antiviral therapies targeted to the functions of pp71 required for viral replication to treat both proliferative and non-proliferative diseases associated with HCMV infection.
