Effect of C-29EA on expression of viral proteins in replicon cell lines.

<p>The Huh7 replicon cell lines derived from genotype 1b (A) and 2a (B) were incubated with C-29EA at 37°C for 72 h. The treated cells were harvested and then subjected to Western blotting. Treatment with DMSO corresponds to ‘0′. The bicistronic gene is transcribed under the control of the elongation factor 1α (EF1α) promoter. The upstream cistron encoding <i>Renilla</i> luciferase (RLuc) is translated by a cap-dependent mechanism. The downstream cistron encodes the fusion protein (Feo), which consists of the firefly luciferase (Fluc) and neomycin phosphotransferase (Neo^r), and is translated under the control of the EMCV IRES (C) or HCV IRES (D). The Huh7 cell line transfected with the plasmid (each above the panel in C and D) was established in the presence of G418. The cells were incubated for 72 h without (control) and with 15 µg/ml of C-29EA. Firefly or <i>Renilla</i> luciferase activity was measured by the method described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0048685#s4" target="_blank">Materials and Methods</a> and was normalized by the protein concentration. F/R: relative ratio of firefly luciferase activity to <i>Renilla</i> luciferase activity. F/R is presented as a percentage of the control condition. Error bars indicate standard deviation. The data represent three independent experiments.