Extended Data Fig. 5: Syntenin-1 and syntenin-2 inhibit TLR7 signalling.

a, Syntenin-1 is selectively recruited to UNC93B1 after TLR7 stimulation, but not TLR3 stimulation. Syntenin-1 binding to UNC93B1 was measured by Flag immunoprecipitation followed by immunoblot for syntenin-1 from RAW macrophage lines stimulated with R848 (0.5 μg ml⁻¹) or poly(I:C) (10 µg ml⁻¹) for the indicated times. Levels of syntenin-1 and UNC93B1–Flag in cell lysates are also shown. b, Syntenin-1 associates selectively with the TLR7–UNC93B1 complex, but not with TLR9. Syntenin-1 binding to TLR7–HA or TLR9–HA was measured by haemagglutinin immunoprecipitation followed by immunoblot for syntenin-1 from indicated RAW macrophage lines stimulated with R848 (0.5 μg ml⁻¹) or CpG-B (0.5 µM) for the indicated times. Levels of syntenin-1 and TLR7–HA or TLR9–HA in cell lysates are also shown. c, NF-κB activation in HEK293T cells transiently expressing syntenin-1 and stimulated with TNF (10 ng ml⁻¹). d, NF-κB activation in HEK293T cells transiently expressing TLR7 and increasing amounts of syntenin-2. Cells were stimulated with R848 (50 ng ml⁻¹) for 16 h before collection. Data in c and d were measured using a dual luciferase reporter assay, normalized to Renilla expression and expressed as RLUs. Data are mean ± s.d., n = 3 biological replicates. *P < 0.05, **P < 0.01 and ***P < 0.001, one-way ANOVA followed by a Tukey’s post-test (95% confidence interval). All data are representative of at least three independent experiments.