Varicella Viruses Inhibit Interferon-Stimulated JAK-STAT Signaling through Multiple Mechanisms
Fig 3
SVV prevents IFN-mediated phosphorylation of STAT2 and reduces expression levels of STAT2 and IRF9.
(A) TRFs were infected with SVV.eGFP (ratio 5:1) for 48 hours and stimulated for 20 minutes with 5000 U/ml uIFN. Lysates were analyzed for expression of various members (non-phosphorylated as well as phosphorylated (p)) of the JAK-STAT pathway using SDS-PAGE and western blot with antibodies specific for the indicated proteins. ORF31 expression confirmed productive viral infection and GAPDH was used as a loading control. (B) Relative expression of STAT1, STAT2 and IRF9 in SVV-infected cells compared to unstimulated mock-infected cells (set at 100%). Shown are the averages of four independent experiments. (C) Mock- and SVV.eGFP-infected cells (ratio 5:1) were stimulated with 5000 U/ml uIFN at 48 hours p.i. for 20 minutes. Cytoplasmic and nuclear fractions were isolated and analyzed for IRF9 expression by SDS-PAGE and western blotting. An antibody directed against ORF62 confirmed productive SVV-infection. Purity of fractionation was confirmed using GAPDH (cytosolic) and p84 (nuclear). Results from one of three three independent experiments is shown.