Membrane rearrangement by positive-sense RNA viruses: Molecular mechanisms and cellular responses

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Lipid membranes mediate intracellular compartmentalization and play a key role in organelle architecture and physiology. Approximately one third of proteins are membrane-embedded, and their activities are regulated by the physiochemical properties of the lipid membrane. Cellular structure and function therefore depend on proper regulation of membrane physiology and homeostasis, yet the underlying mechanisms are poorly understood. Viruses have long been considered excellent tools to dissect fundamental cellular biology. Positive-sense RNA viruses (or (+)ssRNA viruses) are a large group of viruses that extensively remodel host intracellular membranes. We use (+)ssRNA viruses as tools to understand fundamental mechanisms regulating lipid membrane remodeling and homeostasis, which are crucial for the maintenance of normal cellular function. One theme of our research is to understand how (+)ssRNA viruses co-opt host factors and cellular mechanisms to rearrange membranes. Using hepatitis C virus (HCV) as a model (+)ssRNA virus, we identified the host protein cyclophilin A (CypA) as a key player in HCV-induced intracellular membrane rearrangement. We now seek to define how CypA regulates membrane rearrangements. CypA interacts with the HCV protein NS5A, which has a central role in membrane rearrangement. We hypothesize that the CypA-NS5A interaction regulates lipid trafficking to drive intracellular membrane remodelling. In order to maintain cellular functionality, membrane perturbation must be "buffered" by the cell. A second theme of our research is to characterize how cells respond to membrane rearrangements. We hypothesize that cells sense membrane perturbation (such as that induced by (+)ssRNA viruses) as a stress signal and activate cellular responses that restore membrane homeostasis. Our recent findings suggest a role for CypA in regulating protein kinase R (PKR)-dependent stress responses. Strikingly, CypA inhibition appears to activate a PKR-dependent antiviral program that is particularly potent against (+)ssRNA viruses (i.e., viruses that rearrange membranes). We will now determine how CypA regulates PKR-dependent responses that impact membrane rearrangement. Furthermore, we will probe the role of other stress-related proteins in sensing intracellular membrane perturbation. Significance: We anticipate that our findings will provide new understanding of the mechanisms regulating lipid membrane homeostasis and remodelling of intracellular membranes. By advancing our understanding of these fundamental cellular processes, we expect our findings will be important for researchers studying many aspects of cell biology, ranging from lipid metabolism to membrane proteins to cell intrinsic antiviral responses. More broadly, our work will benefit Canada by training several HQP in state-of-the-art techniques in cellular and molecular biology, providing our future scientific leaders with the required expertise to excel in natural science careers.