Fig. 3: HRD1 and TLR3 signaling form a positive feedback loop between each other.

a qPCR analysis of Hrd1 mRNA levels in primary macrophages pre- and post-poly(I:C) (50 μg/ml for 3 h) treatment. n = 4 each. b–c Immunoblot analysis of ERK1/2 phosphorylation levels (b) and qPCR analysis of Ets1 and Hrd1 mRNA levels (c) in macrophage cells pre-treated with LS-102 (5 μM) or ERK1/2 inhibitor PD98059 for 24 h, and then with poly(I:C) (50 μg/ml) for 3 h. n = 4 each in (c). d–e Immunoblot analysis of ERK1/2 phosphorylation levels (d) and qPCR analysis of Ets1 and Hrd1 mRNA levels (e) in macrophage cells pre-treated with LS-102 (5 μM for 24 h), and stimulate with poly(I:C) (50 μg/ml) and indicated cytokines for 3 h. n = 4 each in (e). f Immunoblot analysis of indicated proteins in macrophages treated with poly(I:C), TNFα, IL1β, IL6 or IFNβ for 12 h. g qPCR analysis of Tlr3 mRNA levels in macrophages pre- and post-poly(I:C) (50 μg/ml for 3 h) treatment. n = 4 each. h Immunoblot analysis of indicated proteins in macrophages pre-treated with vehicle or LS-102 for 24 h, followed by poly(I:C) (50 μg/ml) treatment for the indicated times. i–j Immunoblot analysis of STAT1 phosphorylation levels (i) and qPCR analysis of Irf1, Irf2 and Tlr3 mRNA levels (j) in macrophages pre-treated with LS-102 (5 μM) or STAT1 inhibitor Fludarabine for 24 h, and then stimulated with poly(I:C) (50 μg/ml) for 3 h. n = 4 for (j). k–l Immunoblot analysis of STAT1 phosphorylation levels (k) and qPCR analysis of Irf1, Irf2 and Tlr3 mRNA levels ( l) in macrophages pre-treated with LS-102 (5 μM) for 24 h, followed by stimulation with poly(I:C) (50 μg/ml) and IFNβ for 3 h. n = 4 for (l). All experiments were repeated for at least three times. Quantitation of the ratio of phosphorylated to total protein (p/t) and indicated protein is shown below each blot. Values represent mean ± SEM, NS, not significant, by unpaired, two-tailed, Student’s t-test (a, c, e, g, j, l). Source data are provided as a Source Data file.