Figure 5

PKR contributed to the upregulation of IFN-β after NDV infection. A. HeLa cells were either stimulated with poly (I:C) (5 μg/mL) or infected with N strain Herts33 at an MOI of 1 for 12 h. The cell lysates were collected and analyzed by western blot analysis with anti-PKR, anti-p-PKR (T446), anti-eIF2α, and anti-p-eIF2α (S51) antibodies. β-actin was used as a protein loading control. B. HeLa cells were treated with poly (I:C) (5 μg/mL) or infected with NDV strain Herts33 at an MOI of 1 for 12 h. IFN-β mRNA levels in the cell lysates were quantified by real-time PCR. The folds of mRNA expression were calculated by the formula of comparative C_T method. Data are presented as means from three independent experiments. Significance was analyzed using One-way ANOVA followed by Tukey’s test. **, p ≤ 0.01; ***, p ≤ 0.001. C. HeLa cells were transfected with PKR-specific siRNA (100 pmol/mL) or control siRNA for 48 h and then inoculated with NDV strain Herts/33 at an MOI of 1. The cells were harvested and analyzed by western blot analysis to determine the efficiency of siRNAs. D. HeLa cells transfected with siPKRs were infected with NDV strain Herts/33 at an MOI of 1 for 12 h and then harvested for qualification of IFN-β mRNA levels by real-time PCR. The folds of mRNA expression were calculated using the comparative C_T method. Data are presented as means from three independent experiments. Significance was analyzed using two-tailed Student’s t-test. ***, p ≤ 0.001.