Activation of IFN pathway by Candid#1 JUNV infection.

<p>(A) Vero and A549 cells were mock-infected or infected (MOI of 1) with Candid#1 JUNV for 24 h and then mock-treated or treated with IFN-α (1000 IU/ml) for 1 h. Cell lysates were prepared and subjected to western blotting analysis for phosphorylated STAT1 (p-STAT1), STAT1 protein (STAT1), ISG15 (ISG15), virus NP protein (NP) and human β-actin (actin). (B) Vero cells were co-transfected with an IFN responsive IRES reporter plasmid pISRE-luc (expressing firefly luciferase) together with a plasmid constitutively expressing the Renilla luciferase (RL) reporter gene under the Simian Virus 40 promoter (pRL-SV40). After 24 h of transfection, cells were infected with Candid#1 JUNV (Candid) or mock infected (mock) for 24 h followed by mock- (IFN−) or IFN-α treatment (1000 IU/ml) (IFN+) for 6 hr. Cells lysates were prepared and subjected to dual luciferase assay. The ISRE-driven firefly luciferase activity was normalized to the RL activity. Data shown are the mean of three independent experiments (error bar, S.D.). (C) A549 cells were mock infected (mock) or infected with Candid#1 JUNV (Candid). At various time points as indicated, cytoplasmic extract and nuclear extract were prepared for measurement of IRF3 protein level in cytoplasm and nuclear fractions by SID-SRM analysis.