Figure 9.

NS2 inhibits interferon-β (IFN-β) induced by H1C. (A) Investigation of the effect of NS2 on IFN-β stimulated by H1C. A549 cells were transfected with pCAGGS and pCAGGA-NS2 individually, or co-transfected with p3xflag or H1C, then stimulated by poly (I:C) (20 nmol/mL) for 6 h, and IFN-β mRNA was detected by real-time PCR. (B) NS2 affects IRF3 phosphorylation and H1C–IRF3-P interaction. (a) Human embryonic kidney 293 T (HEK293T) cells co-expressed with NS2 and H1C or its mutants were infected with Sendai virus (Sev) for 10 h. Next, the cells were lysed, and IRF3 and IRF3-P levels were detected by western blotting, respectively; (b) cells were treated as described above, and co-immunoprecipitation (Co-IP) experiments were performed to detect the effect of NS2 on the H1C–IRF3-p interaction; (c) detection of the effect of NS2 on IRF3 and IRF3 phosphorylation. Human embryonic kidney 293 T cells were transfected with pCAGGS or pCAGGS-NS2 and infected with Sev for 12 h. IRF3 and IRF3 phosphorylation levels were detected by western blotting. (C) Analysis of the effects of NS2 on the ability of IRF3 to bind to the IFN-β promoter by chromatin immunoprecipitation and quantitation PCR. HEK293T cells co-transfected with IRF3, NS2, and H1C or its mutants and stimulated by Sev, and the chromatin immunoprecipitation assay was performed using anti-IRF3-P antibody. The negative control was performed using the mouse IgG antibody, and the positive control was performed using the anti-poly II antibody (*p < 0.05, ***p < 0.001, the data were generated from one of three independent experiments).