Supplementary Figure 2: RNF128 positively regulates IFN-β signaling.
(a) Quantitative RT-PCR analysis of Rnf128 mRNA expression (upper) and western blot analysis of RNF128 protein expression (lower) in mouse peritoneal macrophages transfected with control siRNA or mice RNF128 siRNA 1, 2 or 3 for 48 h. (b) mRNA expression of Ifnb1, Cxcl10, Mx1, Ccl5, Il6 and Tnfa in mouse peritoneal macrophages transfected with control siRNA or mice RNF128 siRNA 1, 2 or 3 and then stimulated with poly(I:C) and ISD or infected with SeV and HSV-1. (c) Quantitative RT-PCR analysis of RNF128 mRNA expression (upper) and western blot analysis of RNF128 protein expression (lower) in THP-1 cells transfected with control siRNA or human RNF128 siRNA 1, 2 or 3 for 48 h. (d) Quantitative RT-PCR analysis of IFNB1, CXCL10, MX1, CCL5, IL6 and TNFA mRNA expression in THP-1 cells transfected with control siRNA or human RNF128 siRNA 1, 2 or 3 for 48 h and then stimulated with poly(I:C) and ISD or infected with SeV and HSV-1. (e) Quantitative RT-PCR analysis of Ifnb1, Cxcl10, Mx1, Ccl5, Il6 and Tnfa mRNA expression in WT or Rnf128−/− mouse peritoneal macrophages stimulated with LPS for the indicated time. (f,g) Luciferase activity of IFN-β promoter reporter and mRNA expression of IFNB1 in HEK293 cells transfected with expression plasmids for TRIF, cGAS+STING and RIG-IN along with V5-RNF128 expression plasmid or control vector for 24 h. *p <0.05,# p <0.01 (two-tail Student's t-test). Data are from three independent experiments (a-f; mean+S.D. of triplicate assays) or are representative of three independent experiments with similar results (a,b).
