Antiviral Effect Of PKR-eIF2α Against Newcastle Disease Virus

Newcastle disease is a highly contagious and fatal viral disease affecting most species of poultry. Itis caused by Newcastle disease virus (NDV) which belongs to the family Paramyxoviridae, subfamilyParamyxovirinae with the single stranded, negative genomic RNA.Interferon, one of the most important components of the cellular antiviral innate immune, couldimmediately enhance the antiviral immune through activating some interferon-stimulated genes (ISG).double-stranded RNA-activated protein kinase (PKR), one of Interferon inducing protein, couldrecognize double stranded RNA (dsRNA) and initiate antiviral signaling pathway by phosphorylatinginitial factor2(eIF2α).NDV is known to have the ability to inhibit the translation of host protein. However, antiviraleffect of PKR against NDV still has limited knowlege. In this study, we demonstrated that NDV couldactivate PKR by generating viral dsRNA. The antiviral effect of PKR against NDV was evaluated byover-expression or knock-down of PKR in HeLa cells. We also confirmed that eIF2α participated inNDV replication and PKR was essential in Interferon production induced by NDV.1. The influence of NDV infection on the PKR-eIF2α signaling pathwayWe found that when NDV replication viral dsRNA was generated in cytoplasm. The significantphosphorylation of PKR and eIF2α suggested that PKR-eIF2α signaling pathway could be activated byviral dsRNA compared to the inactivated NDV by UV treatment.2. The antiviral role of PKR against NDVTo determine whether PKR induces an antiviral effect, HeLa cells with over-expression orknock-down of PKR were infected with NDV strain Herts/33. We found that over-expression of PKRcould block the synthesis of viral protein and inhibit the replication of NDV. However, NDV replicationwas significantly enhanced in the PKR knockdown cells.3. The function of eIF2α in NDV replicationTo determine whether PKR inhibits the NDV replication via phosphorylation of eIF2α and toconfirm the role of eIF2α in NDV replication, eIF2α knockdown HeLa cells were infected with NDV.The results showed that NDV replication was inhibited in eIF2α knockdown cells by decreasing viralproteins expression and virus amount. Moreover, phosphorylation of eIF2α was enhanced by Okadaicacid in HeLa cells with NDV infection. The results suggested that Okadaic acid significantly inhibitedNDV replication. Above all, NDV replication in HeLa cells depends on the host translation system andphosphorylation of eIF2α could suppress its replication.4. The function of PKR in the production of interferon induced by NDVTo explore whether PKR involves in the production of interferon induced by NDV, PKRknockdown HeLa cells were infected with NDV Herts/33and the levels of IFN-β mRNA was evaluated.Compared to the controls, the levels of FN-β mRNA in PKR knockdown cells were greatly reduced inaccordance to the levels of PKR. These results demonstrated that PKR plays a role in production ofNDV inducing interferon.