Fig. 5: TNIP1Q333P regulates NF-κB signaling but not IFNβ and alters the size of TNIP1 puncta.
From: A TNIP1-driven systemic autoimmune disorder with elevated IgG4
a–c, NF-κB activity (ratio of NF-κB firefly to Renilla luciferase in relative light units) 24 h after lipofectamine transfection of human TNIP1 plasmids (WT, Q333P and D472N) into HEK 293 cells cotransfected with MyD88 (a), TBK1 (b) and TRAF6 (c). Data are shown as the mean and s.d. of n = 3 biological replicates or transfections, shown as individual dots and representative of three independent experiments (a–c). d, Immunoblots (IBs) of splenic B cell lysates from 20-week-old WT (+/+) and vikala homozygote (vik/vik) mice probed for TNIP1 and actin proteins after stimulation with CpG-B, R848, BCR (αIgM), BCR and CpG-B, BCR and R848, CD40 and BAFF. Data are representative of two independent experiments. e–h, IFNβ (e–g) and NF-κB (h) activity (ratio of IFNβ or NF-κB firefly to Renilla luciferase in relative light units) 24 h after lipofectamine transfection with human TNIP1 plasmids (WT, Q333P and D472N) into HEK 293 cells cotransfected with MyD88 (e) or TBK1 (f–h). Data are shown as the mean and s.d. of n = 3 biological replicates or transfections shown as individual dots and representative of three independent experiments (e–h). Statistical significance was calculated using a one-way ANOVA with Tukey’s correction for multiple comparisons (a–c,e–h). Exact P values are shown. i, ELISA-determined serum levels of IFNβ protein from vikala BM-pDCs; Tnip1+/+ (n = 5) and Tnip1vik/vik (n = 5), untreated or stimulated with CpG-A, R848, R837 and Escherichia coli. Data are representative of three experiments. The error bars indicate the mean with the s.e.m.; each dot represents a single mouse (biological replicate). j, Immunofluorescence staining of HEK 293 cells expressing TNIP1WT, TNIP1Q333P or TNIP1D472N. k, Quantification of WT (n = 301), Q333P (n = 305) and TNIP1D472N (n = 268) mean particle size. Scale bar, 10 μm. Data are representative of n = 3 independent experiments. Statistical significance was performed using a two-way ANOVA with Šidák’s correction for multiple comparisons (i) and a one-way ANOVA with Tukey’s correction for multiple comparisons (k). The error bars indicate the mean with the s.d. Exact P values are shown.
