| Apoptosis of virus-infected cells often contributes to disease pathogenesis. Therefore, understanding virus-host interactions that cause apoptosis is of great interest for development of improved strategies for virus control and therapeutics. The baculovirus Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) induces apoptosis in insect cells. The goal of my thesis research was to identify the baculoviral factors that induce apoptosis. By using RNA interference (RNAi) to reduce expression of essential baculoviral genes, I blocked the virus lifecycle at different stages and analyzed the effect on induction of apoptosis. I determined that baculovirus replicative late expression factors ( lefs) were necessary for viral DNA replication during Ac MNPV infection. Detailed analysis of the transactivator ie-1 showed that this replicative lef was required for multiple stages of the baculoviral lifecycle, including transactivation of select viral genes and viral DNA replication. RNAi-mediated ablation of ie-1 or any of the replicative lefs, but not other early genes, prevented AcMNPV-induced apoptosis. Because the replicative lefs were required for apoptosis, I concluded that the process of viral DNA replication was directly or indirectly responsible for inducing apoptosis. Additionally, because the nonreplicative lefs were not required for apoptosis, I concluded that viral late gene expression did not trigger apoptosis.;My thesis research also showed that cellular inhibitor-of- apoptosis (IAP) proteins are central regulators of baculovirus-induced apoptosis. I demonstrated that AcMNPV infection depleted endogenous IAP levels through a caspase-independent mechanism. When intracellular IAP levels were increased by iap overexpression, virus-induced proteolytic caspase activation and cell death were blocked. My research supports a model whereby AcMNPV infection promotes proteasome-dependent depletion of cellular IAPs by a mechanism that is independent of direct interactions with the Reaper-Hid-Grim family of pro-apoptotic proteins. My studies have demonstrated that intracellular IAP levels are a critical determinant of cell survival during AcMNPV infection. By studying baculovirus-induced apoptosis in insect cells, I have shown that the mechanism by which DNA viruses induce apoptosis in vertebrate and invertebrate cells is conserved. |
