NORTH CAROLINA STATE UNIVERSITY
Our overall goal is to identify novel components of the host innate antiviral response and novel mechanisms by which viruses subvert this response to cause disease. Cardiac myocytes are non-replenishable; thus the heart is unusually dependent on this first-line defense. Indeed, we have shown that the cardiac Type I interferon (IFN) response is unique, and that in reovirus-induced murine myocarditis the IFN response differs between virus strains and is critical for protection. We continue to use this powerful tool-kit of viruses to probe the innate response in a highly vulnerable organ, the heart, and have made two new discoveries. First, we found that reovirus inhibits IFN signaling by a mechanism not previously shown for any virus. Specifically, reovirus protein mu2 represses IFN signaling, and repression is virus strain-specific and is associated with unusual nuclear accumulation of transcription factor IRF9, likely reflecting concomitant interference with its normal participation in induction of IFN-stimulated genes. We hypothesize that reovirus protein mu2 modulates IRF9 structure / function to inhibit IFN signaling, and that mu2 repression of IFN signaling in cardiac cells is required for myocarditis. In Specific Aim 1, we will: i) determine whether mu2 alters IRF9 structure or cellular binding partners; ii) identify mu2 and IRF9 domains required for IRF9 effects, and iii) determine whether mu2 modulation of IRF9 is required for reovirus-induced myocarditis. Second, using a proteomic discovery approach, we identified a new IFN-independent protective response which can be subverted by virus. We found that reoviruses induce phosphorylation (non-myocarditic reovirus) or degradation (myocarditic reovirus) of Heat Shock Protein-25 (HSP25) in cardiac myocytes, and that this is cell type-specific and IFN-independent. Members of seven virus families induce HSP25 expression or phosphorylation but not degradation, yet only one study has addressed HSP25 antiviral effects. Many cardiac insults induce HSP25 phosphorylation but not degradation, and over-expressed HSP25 protects against stress-induced cardiac damage. We hypothesize that HSP25 plays a cell type-specific antiviral role, that phosphorylated HSP25 is antiviral, and that viruses have evolved mechanisms to evade this innate response. In Specific Aim 2, we will determine: i) cell type-specificity for reovirus alteration of HSP25, ii) mechanisms by which reovirus alters HSP25, and iii) mechanisms by which HSP25 inhibits reovirus infection. The broader impact of our studies is to increase the catalog of protective host factors that can be sabotaged by viruses, potentially providing new therapeutic targets, particularly for viral myocarditis which remains an intractable disease.