Fig. 8: Hydroxylation of IRF3 at proline 10 enhances IRF3 activation and nuclear translocation in cellular antiviral immune responses.

a IFN-β promoter activity and ISRE reporter activity in HEK293T transfected with the Myc empty vector (EV) or the plasmid expressing wild-type IRF3 (WT) or its mutant (P10A). b IFN-β promoter activity and ISRE reporter activity in IRF3^−/− H1299 cells transfected with the Myc empty vector or the plasmid expressing wild-type IRF3 or its mutant (P10A). c qPCR analysis of IFN-β mRNA in HEK293T transfected with the empty vector or the plasmid expressing wild-type IRF3 (WT) or its mutant (P10A), followed by infected without (UI) or with SeV for 8 h. d qPCR analysis of IFN-β, CXCL10 and ISG15 mRNA in IRF3^−/− H1299 transfected with the empty Flag vector or the plasmid expressing Flag-tagged wild-type IRF3 or its mutant (P10A), together with the empty Myc vector or the plasmid expressing Myc-tagged EGLN1 (EGLN1). e qPCR analysis of Ifn-β mRNA in Irf3 and Irf7-deficient MEF (Irf3^−/−Irf7 ^−/−) transfected with the empty Flag vector or the plasmid expressing Flag-tagged wild-type Irf3 or its mutant (P10A), together with the empty Myc vector or the plasmid expressing Myc-tagged Egln1 (Egln1). f IRF3^−/− H1299 were transfected with the plasmid expressing Flag-tagged wild-type IRF3 or its mutant (P10A) for 24 h, followed by infection without or with VSV-GFP viruses for 12 h, and viral infectivity was detected by fluorescence microscopy or flow cytometry analysis (bottom panels). g IRF3^−/− H1299 were transfected with the plasmid expressing HA-tagged wild-type IRF3 or P10A mutant (P10A), followed by infection without or with SeV for 8 h. Confocal microscopy image of exogenous IRF3 was detected by immunofluorescence staining with anti-HA antibody. Scale bar = 25 µm. BR, bright field. Data in (a, b) are presented as mean ± S.D., two-tailed Student’s t test; n = 3 biological independent experiments. Data in (c–e) are presented as mean ± S.D., two-way ANOVA; n = 3 biological independent experiments. Data in (f, g) are representative from three independent experiments. See also Supplementary Fig. 13. Source data are provided as a Source data file.