FIG 4.
3Cpro of EV68 reduces the transcription of IFN-β (A), ISG54 (B), IL-6 (C), ISG56 (D), IP10 (E), and RANTES (F) by poly(I·C). HeLa cells were transfected with GFP or GFP-3C. Twenty-four hours after transfection, cells were treated with or without poly(I·C). Two, 4, 8, and 12 h after poly(I·C) treatment, total RNAs extracted from cells and the expression of the IFN-β, ISG54, ISG56, IL-6, RANTES, IP10, and GAPDH genes were evaluated by quantitative real-time PCR using SYBR green. Results are expressed as increases in mRNA levels relative to those in cells in the absence of poly(I·C) and were normalized by using the GAPDH housekeeping gene. *, P < 0.05; **, P < 0.01; ***, P < 0.001. (G) 3Cpro of EV68 inhibits poly(I·C)-induced phosphorylation of IRF3. HeLa cells were transfected with GFP or GFP-3C. Twenty-four hours after transfection, cells were treated with or without poly(I·C) for 4 h. Cell lysates were subjected to Western blot analysis with antibodies against IRF3 or phosphorylated IRF3. (H) Densitometry analysis. Phosphorylated IRF3 protein bands from three independent experiments as shown in panel G were quantified and normalized to IRF3 by using Odyssey image software. *, P < 0.05.