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

Figure 4. Overexpression of IRS4 enhanced IFN-a-induced activation of Jak/STAT signalling.

Figure 4

(A, B) Overexpression of IRS4 enhanced IFN-a stimulated p-STAT1 level. 293T cells (A) or Huh-7.5.1 cells (B) were transfected with Flag (M, mock control), Flag-IRS4 or untreated(un), the cells were treated with 100 IU/mL IFN-α for 30 mins at 48 hours after transfection. Protein lysates were harvested, separated by SDS-PAGE, 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 (bottom). (C) Overexpression of IRS4 promoted IFN-a-stimulated ISRE activity. Huh7.5.1 cells were co-transfected with Flag-IRS4 plasmid or Flag empty plasmid and pISRE-luc (expressing firefly luciferase) and pRL-TK (expressing Renilla luciferase) as described in Materials and Methods. (D) Overexpression of IRS4 promoted IFN-a-induced ISG expression. Expression levels of IFIT1/VIPERIN/MX1 were examined by real time PCR in Huh7.5.1 cells transfected with IRS4 plasmid, empty plasmid (M, mock control) 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, **P<0.01, ***P<0.001.