Viral RNA at Two Stages of Reovirus Infection Is Required for the Induction of Necroptosis

Abstract

Necroptosis, a regulated form of necrotic cell death, requires the activation of the RIP3 kinase. Here, we identify that infection of host cells with reovirus can result in necroptosis. We find that necroptosis requires sensing of the genomic RNA within incoming virus particles via cytoplasmic RNA sensors to produce type I interferon (IFN). While these events that occur prior to the synthesis of viral RNA are required for the induction of necroptosis, they are not sufficient. The induction of necroptosis also requires late stages of reovirus infection. Specifically, efficient synthesis of double-stranded RNA (dsRNA) within infected cells is required for necroptosis. These data indicate that viral RNA interfaces with host components at two different stages of infection to induce necroptosis. This work provides new molecular details about events in the viral replication cycle that contribute to the induction of necroptosis following infection with an RNA virus. An appreciation of how cell death pathways are regulated following viral infection may reveal strategies to limit tissue destruction and prevent the onset of disease. Cell death following virus infection can occur by apoptosis or a regulated form of necrosis known as necroptosis. Apoptotic cells are typically disposed of without activating the immune system. In contrast, necroptotic cells alert the immune system, resulting in inflammation and tissue damage. While apoptosis following virus infection has been extensively investigated, how necroptosis is unleashed following virus infection is understood for only a small group of viruses. Here, using mammalian reovirus, we highlight the molecular mechanism by which infection with a dsRNA virus results in necroptosis.

Loss of IKK subunits limits NF-κB signaling in reovirus infected cells

“…To determine the stage of the reovirus replication cycle that is required for the loss of the IKK complex and the inhibition of NF-κB, we treated cells with ribavirin, which diminishes viral gene expression (13, 22). We found that ribavirin treatment prevented T3A mediated loss of IKKβ (Fig.…”

“…This outcome, at least in part, is due an inability of p50−/− mice to produce IFNβ. Despite the fact that the viral genomic RNA remains within the two concentric protein shells that comprise the reovirus capsid, the current model posits that the innate immune response is initiated when genomic RNA from incoming virions is sensed by the RLRs - RIG-I and MDA5 (9, 13). The sensing of the RNA leads to the activation of IRF3 and NF-κB, which lead to the production of IFN and other inflammatory cytokines (14).…”

Apoptosis and Necroptosis as Host Defense Strategies to Prevent Viral Infection

“…Additional sensors that initiate necroptosis following virus infection likely exist, as infection of L929 cells with the T3D strain of respiratory enteric orphan virus (reovirus) induces caspase-independent cell death [52] (Figure 1). Cell death following T3D infection is dependent on de novo synthesis of viral dsRNA and requires RIPK1, but is independent of the known initiators of necroptosis, TNFR, DAI or TLR3 [81]. In addition, this response requires production of IFN and signaling through the type I IFN receptor, suggesting an IFN stimulated gene(s) might sense or regulate reovirus-mediated cell death [81].…”

“…Cell death following T3D infection is dependent on de novo synthesis of viral dsRNA and requires RIPK1, but is independent of the known initiators of necroptosis, TNFR, DAI or TLR3 [81]. In addition, this response requires production of IFN and signaling through the type I IFN receptor, suggesting an IFN stimulated gene(s) might sense or regulate reovirus-mediated cell death [81]. Further identification and characterization of this response remains an important area of future study.…”

Enhanced Killing of Triple-Negative Breast Cancer Cells by Reassortant Reovirus and Topoisomerase Inhibitors

“…Reovirus induces programmed cell death in vitro and in vivo (23–30). Although both Type 1 and Type 3 reovirus can induce apoptosis, Type 3 reoviruses induce apoptosis and necroptosis more efficiently in most cells (18, 23, 24). Serotype-dependent differences in apoptosis induction segregate with the S1 and M2 gene segments (31–33).…”