Genotype 2 Strains of Porcine Reproductive and Respiratory Syndrome Virus Dysregulate Alveolar Macrophage Cytokine Production via the Unfolded Protein Response

Porcine reproductive and respiratory syndrome virus (PRRSV) infects alveolar macrophages (AM phi), causing dysregulated alpha interferon (IFN-alpha) and tumor necrosis factor alpha (TNF-alpha) production through a mechanism(s) yet to be resolved. Here, we show that AM phi infected with PRRSV secreted a reduced quantity of IFN-alpha following exposure of the cell to synthetic double-stranded RNA (dsRNA). This reduction did not correlate with reduced IFNA1 gene transcription. Rather, it coincided with two events that occurred late during infection and that were indicative of translational attenuation, specifically, the activation of eukaryotic translation initiation factor 2 alpha (eIF2 alpha) and the appearance of stress granules. Notably, the typical rapid production of TNF-alpha by AM phi exposed to lipopolysaccharide (LPS) was suppressed or enhanced by PRRSV, depending on when the LPS exposure occurred after virus infection. If exposure was delayed until 6 h postinfection (hpi) so that the development of the cytokine response coincided with the time in which phosphorylation of eIF2 alpha by the stress sensor PERK (protein kinase RNA [PKR]-like ER kinase) occurred, inhibition of TNF-alpha production was observed. However, if LPS exposure occurred at 2 hpi, prior to a detectable onset of eIF2 alpha phosphorylation, a synergistic response was observed due to the earlier NF-kappa B activation via the stress sensor IRE1 alpha (inositol-requiring kinase 1 alpha). These results suggest that the asynchronous actions of two branches of the unfolded protein response (UPR), namely, IRE1 alpha, and PERK, activated by ER stress resulting from the virus infection, are associated with enhancement or suppression of TNF-alpha production, respectively. IMPORTANCE The activation of AM phi is controlled by the microenvironment to deter excessive proinflammatory cytokine responses to microbes that could impair lung function. However, viral pneumonias frequently become complicated by secondary bacterial infections, triggering severe inflammation, lung dysfunction, and death. Although dysregulated cytokine production is considered an integral component of the exacerbated inflammatory response in viral-bacterial coinfections, the mechanism responsible for this event is unknown. Here, we show that PRRSV replication in porcine AM phi triggers activation of the IRE1 alpha branch of the UPR, which causes a synergistic TNF-alpha response to LPS exposure. Thus, the severe pneumonias typically observed in pigs afflicted with PRRSV-bacterial coinfections could result from dysregulated, overly robust TNF-alpha production in response to opportunistic pathogens that is not commensurate with the typical restrained reaction by uninfected AM phi. This notion could help in the design of therapies to mitigate the severity of viral and bacterial coinfections.