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. 2023 Feb 20;12(1):2178238. doi: 10.1080/22221751.2023.2178238

Figure 2.

Figure 2.

NSUN2 inhibits the expression level of IRF3. (a) Dual-luciferase assay analysing a luciferase reporter plasmid for the IRF3-responsive promoter containing positive regulatory domains III and I of the IFN-β promoter (PRDIII-I-Luc) in HEK293T cells in 24-well plates transfected for 36 h with the RIG-N, MDA5-N, MAVS, TBK1, and IRF3-5D expression plasmids, as indicated, with co-transfection with empty vector or NSUN2. (b) Dual-luciferase analysis of PRDIII-I-Luc in HEK293T cells in 24-well plates transfected for 36 h with the indicated RIG-N, MDA5-N, MAVS, TBK1, and IRF3-5D expression plasmids with co-transfection with siControl or siNSUN2-1. (c) Immunoblot analysis in HEK293T cells transfected with vector or NSUN2 for 36 h, with or without infection by SeV for another 12 h. (d) Immunoblot analysis in wild-type HEK293T cells or NSUN2−/− HEK293T cells with or without infection by SeV for 12 h. (e) Immunoblot analysis in wild-type A549 cells or NSUN2−/− A549 cells with infection by SeV for 0, 4, 8, and 12 h. (f) Immunoblot analysis in wild-type HEK293T cells or NSUN2−/− HEK293T cells, with infection by SeV for 0, 4, 8, and 12 h. (g) qPCR analysis of IFNB1 mRNA in wild-type HEK293T cells or NSUN2−/− HEK293T cells transfected with vector or NSUN2, with co-transfection with vector or IRF3-FL, as indicated, with infection by SeV for 12 h. (h) qPCR analysis of VSV-G or HSV-1-UL-30 RNA in Irf3−/−Irf7−/− MEFs transfected with vector or NSUN2, with co-transfection with vector or IRF3-FL, as indicated, with infection by VSV or HSV-1 for 12 h. Data are representative of three independent experiments and were analysed by two-tailed unpaired t test (or by two-factor ANOVA test for 2 h and 2i). Graphs show the mean ± SD (n = 3) derived from three independent experiments. NS, not significant for P > 0.05, *P < 0.05, **P < 0.01, ***P-< 0.001.