AIM: PKR is important in the formation of antiviral stress granules (AvSGs) and subsequent induction of IFN-β, however it is unclear if PKR or AvSG is important in the formation of the transcription factor complex IFN-β enhanceosome. People with chronic obstructive pulmonary disease (COPD) are more susceptible to influenza infection, the mechanisms of which remain unclear. In this study we aim to investigate if PKR is essential in AvSG, IFN-β enhanceosome formation, and subsequent IFN-β protein induction. We also aimed to determine if these responses are impaired in people with COPD, leading to deficient antiviral responses to influenza infection.

METHODS: Primary bronchial epithelial cells (pBECs) from healthy controls and subjects with COPD were infected with human influenza A/H1N1 at MOI of 5. AvSG proteins PKR, TIAR, G3BP were detected by confocal microscopy, and by western blotting. IFN-β enhanceosome proteins pIRF3, pATF2, and p300 were also measured. IFN-β protein was measured by western blotting. Viral replication was measured by plaque assay. PKR was silenced by specific siRNA 24 before infection. Co-localisation test was performed using ImageJ according to instructions.

RESULTS: Influenza infection resulted in reduced AvSG component PKR, TIAR, and G3BP expression and decreased formation of AvSG in COPD pBECs. The formation of IFN-β enhanceosome was also significantly decreased in COPD pBECs with reduced expression of enhanceosome components pIRF3, pATF2, and p300. We then determined that PKR was important in the formation of AvSG, as PKR inhibition by siRNA led to impaired AvSG formation and also IFN-β enhanceosome. This then led to reduced IFN-β induction and increased influenza viral replication in COPD pBECs.

CONCLUSION: PKR is important in the formation of AvSGs and IFN-β enhanceosome, and was impaired in COPD pBECs, leading to reduced antiviral responses and heightened viral replication to influenza infection.