Extended Data Fig. 4: IFI16 binds RIG-I promoter and enhances RIG-I transcription.
a, RIG-I expression in A549 cells transfected with IFI16-Flag vectors was examined. b, HEK293 cells co-transfected with indicated plasmids for 24 h were treated with cycloheximide (CHX) for 12 and 24 h. Expression of RIG-I and IFI16 in cell lysates was detected. c, RIG-I expression in ifnar1−/−A549 cells transfected with IFI16-Flag vectors was examined. d, RIG-I mRNA expression in ifnar1−/− A549 cells transfected with IFI16-Flag plasmids for 24 h was determined. e, RIG-I expression in A549 cells transfected with IFI16-Flag plasmids for 24 h was determined. f, RIG-I expression in lung tissues from virus-infected WT and p204−/− mice (n = 3) was determined. g, Schematic diagram of the biotinylated probe sequences from the promoter of RIG-I gene and corresponding mutants. h, IB analysis of the binding ability between wild type (p1 and p2) and mutated (p2-mut1 to p2-mut5) probes, and IFI16 proteins. i, IFI16+/+ and IFI16−/− A549 cells were infected with PR8 virus for 12 h, followed by ChIP assay. j, Schematic diagram of full-length IFI16 vectors and truncated mutants. k, A549 cells were transfected with Flag-tagged full-length IFI16 vectors and truncated mutant plasmids. After 24 h transfection, nuclear extracts were incubated with non-biotinylated or biotinylated promoter sequence of RIG-I for 4 h. Nuclear extracts were examined for IFI16 and truncated mutant expression by Western blotting. l, A549 cells after Flag-tagged IFI16 vectors and truncated mutants transfection and PR8 infection was determined. d to f and i, Data are representative of three independent experiments (mean ± SD). (a), (b), (c), (h), (k), and (l), Data are representative of three independent experiments. Statistical significance in (d) to (f) and (i) was determined by unpaired two-tailed Student’s t-test. ns = non-significant.
