The Early Interferon Response to Rotavirus Is Regulated by PKR and Depends on MAVS/IPS-1, RIG-I, MDA-5, and IRF3

In mouse embryonic fibroblasts (MEFs), the bovine rotavirus (UK strain) but not the simian rhesus rotavirus (RRV) robustly triggers beta interferon (IFN-beta) secretion, resulting in an IFN-dependent restriction of replication. We now find that both rotavirus strains trigger antiviral transcriptional responses early during infection and that both transcriptional responses and IFN-beta secretion are completely abrogated in MAVS/IPS-1(-/-) MEFs. Replication of UK virus could be rescued in MAVS/IPS-1(-/-) MEFs, and synthesis of viral RNA significantly increased early during virus infection. UK virus induced IFN-beta secretion and transcription of IFN-stimulated genes (ISGs) in both RIG-I-/- and MDA-5(-/-) MEFs, and neither receptor was essential by itself for the antiviral response to UK rotavirus. However, when receptors RIG-I and MDA-5 were depleted using RNA interference, we found that both contribute to the magnitude of the IFN response. IRF3 was found to be essential for MAVS/IPS-1-directed ISG transcription and IFN-beta secretion during rotavirus infection. Interestingly, absence of the double-stranded RNA-dependent protein kinase PKR led to a profound defect in the capacity of host cells to secrete IFN-beta in response to virus. Both PKR and IRF3 restricted the early replication of UK as indicated by significant increases in viral RNA in fibroblasts lacking either gene. Despite the loss in IFN-beta secretion in PKR-/- MEFs, we did not observe decreased IRF3- or NF-kappa B-dependent early ISG transcription in these cells. Levels of transcripts encoding IFN-alpha 4, IFN-alpha 5, and IFN-beta were high in infected PKR-/- MEFs, indicating that during rotavirus infection, PKR functions at a stage between IFN gene transcription and subsequent IFN-beta secretion. These findings reveal that activation of the antiviral response by rotavirus is dependent on MAVS/IPS-1 and IRF3 and involves both RIG-I and MDA-5 and that IFN-beta secretion during rotavirus infection is regulated by PKR.