FIG. 2.

Effect of NS5A expression on IFN-induced posttranslational modifications and upregulation of gene expression. (A) Effect of NS5A expression on IFN induction of STAT-1 phosphorylation and PKR expression. FL-NS5A-NR and FL-NS5A-CR proteins were repressed or induced by growing cells in the presence or absence of doxycycline (Dox) for 24 h and treated with IFN, and protein extracts were harvested at various times thereafter. Protein extracts were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, respectively, followed by Western blotting with antibodies to NS5A (first panels), phosphorylated STAT-1 (second panels), PKR (third panels), and β-tubulin (fourth panels) as described in Materials and Methods. (B) Effect of NS5A expression on IFN-induced upregulation of MHC antigen. HeLa cells expressing FL-NS5A-NR (left panels) and FL-NS5A-CR (right panels) proteins for 24 h were treated with IFN, and cells were prepared for flow cytometry as described in Materials and Methods. Top panels are from cells grown in the presence of doxcycline (NS5A expression repressed), while bottom panels are from cells grown in the absence of doxcycline (NS5A expression induced). The gray histogram on the left represents a control staining with an isotype-matched control. To the right of the control histogram in each panel are the specific staining patterns. IFN-treated cells are depicted by the gray curve, while cells not treated with IFN are depicted by the black curve.