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. 2017 Nov 18;8(62):105923–105935. doi: 10.18632/oncotarget.22510

Figure 5. IRS4 knock-down suppressed the IFN-a-induced activation of Jak/STAT signaling.

Figure 5

(A) IRS4 knock-down inhibited IFN stimulated p-STAT1 level. Huh-7.5.1 cells were transfected with siIRS4, negative control (NC) or untreated (un), the cells were treated with 100 IU/mL IFN-α for 30 min at 48 hours after transfection. Protein lysates were harvested, separated electrophoretically, and probed for STAT1-phospho701. Blots then were stripped and probed for total STAT1 expression. The ratio of p-STAT1 to total STAT1 from 3 independent experiments was quantified (right). (B) IRS4 knock-down inhibited IFN stimulated ISRE activity. Huh7.5.1 cells were co-transfected siRNA and negative control with pISRE-luc (expressing firefly luciferase) and pRL-TK (expressing Renilla luciferase) as described in Materials and Methods. 24 hours later, cells were treated with 100 IU/mL IFN-α for 24 hours before the cells were lysed for dual luciferase reporter gene assay. (C) IRS4 knock-down inhibited IFNa-induced ISG expression. Expression levels of IFIT1/MX1 were examined by real time PCR in Huh7.5.1 cells transfected with siRNA and negative control (NC) or untreated (un), 24 hours after transfection and then treated with 100IU/mL IFN-α for 24 hours. Error bars indicated mean±SD, *P<0.05.