Cell entry-independent role for the reovirus μ1 protein in regulating necroptosis and the accumulation of viral gene products

Abstract

The reovirus outer capsid protein 1 regulates cell death in infected cells. To distinguish between the role of incoming, capsid-associated and newly synthesized 1, we used siRNA-mediated knockdown. Loss of newly synthesized 1 protein does not impact apoptotic cell death in HeLa cells but enhances necroptosis in L929 cells. Knockdown of 1 also impacts aspects of viral replication. We found that while 1 knockdown results in diminished release of infectious viral progeny from infected cells, viral minus strand RNA, plus strand RNA, and proteins that are not targeted by the 1 siRNA accumulate to a greater extent when compared to control siRNA-treated cells. Furthermore, we observe a decrease in sensitivity of these viral products to inhibition by GuHCl (which targets minus strand synthesis to produce dsRNA) when 1 is knocked down. Following 1 knockdown, cell death is also less sensitive to treatment with GuHCl. Our studies suggest that the absence of 1 allows enhanced transcriptional activity of newly synthesized cores and the consequent accumulation of viral gene products. We speculate that enhanced accumulation and detection of these gene products due to a 1 knockdown potentiates RIP3 dependent cell death. IMPORTANCEWe use mammalian reovirus as a model to study how virus infections result in cell death. Here, we sought to determine how viral factors regulate cell death. Our work highlights a previously unknown role for reovirus outer capsid protein 1 in limiting the induction of a necrotic form of cell death called necroptosis. Induction of cell death by necroptosis requires the detection of viral gene products late in infection. 1 limits cell death by this mechanism because it prevents excessive accumulation of viral gene products that trigger cell death.