Figure 2.

IRF5 negatively modulates expression of HCV protein(s) and HCV RNA replication. A, MH-14(c) cells were co-transfected with 80 ng of GFP-IRF5 and the HCV IRES-Luc reporter. -Fold change in luciferase activity is shown after normalization to Renilla. B, same as in A except GFP-IRF5 and HCV IRES-Luc were transfected to Feo1b and HCV2a cells. A representative Western blot of GFP-IRF5 expression in transfected cells is shown. C, levels of HCV viral proteins in HCV replicon cells overexpressing IRF5 were determined by Western blot analysis. D, effect of ectopic IRF5 expression on replicating HCV RNA levels in C-5B cells. HCV RNA expression was analyzed by real-time qPCR 36 h after transfection of 0.1 μg of empty FLAG vector (V) or FLAG-IRF5 (IRF5). Data were normalized to β-actin levels and presented relative to empty vector control. E, Huh7 cells were transfected with scrambled (SCR) or IRF5 siRNAs (siRF5) for 48 h before infecting with purified virus stock for 3 days. IRF5 and HCV RNA levels were determined by real-time qPCR. F, representative Western blot from E showing IRF5 knockdown and HCV protein levels at days 0 and 2 of virus infection. Live cell images are from day 2 postinfection in Huh7 cells transfected with SCR or siIRF5. Images are representative of three independent transient transfections in each cell line. Images were taken at ×40 magnification. Data are representative of duplicate data points from three independent replicates; plotted values are means ± S.D. (error bars) (*, p < 0.05; **, p < 0.001; ***, p < 0.001).