Fig 1: Both the UBA-like and NBR1-like domains of ILRUN are necessary for inhibition of IRF3 signalling, while the disordered region is dispensable. (A) Schematic of ILRUN mutants, including wild-type ILRUN, ΔUBA, lacking residues 1–75, ΔNBR1, lacking residues 105–170, and Δdis, lacking residues 194–298. (B) HeLa cells were transfected with plasmids expressing the indicated proteins (or empty vector/pCAGGS) for 24 h before treatment with transfected poly(I:C) (5 μg/mL) for 6 h, with cells collected and analyzed for mRNA expression of the listed cytokines by qRT-PCR. (C) HeLa cells were transfected with plasmids expressing the indicated proteins and stimulated with poly(I:C), and the nuclear proteins were extracted using a hypotonic lysis method. Nuclear proteins (10 μg) were analyzed for IRF3–DNA binding. (D) Cell supernatants from cells used in (C) were analysed for IFN-b protein by ELISA assay. Error bars denote ±SEM of three independent experiments, and asterisks show significant changes compared with controls as measured by one- or two-way ANOVA with Bonferroni post-test (∗∗∗, p < 0.001; ∗∗, p < 0.01).