RILP cleavage links exosome secretion to inflammasome activation. (A) Exosome release is increased in HCV-infected cells. cRILP expression alone increases exosome output, while ncRILP blocks HCV-induced exosome secretion. When indicated, JNK inhibitor SP600125 (25 µM) was added 24 h before collection. (B) HCV infection activates the inflammasome as noted by the appearance of cleaved (active) caspase-1. (C) The caspase-1 inhibitor VI (Z-YVAD-FMK, 100 µM) blocks HCV-induced exosome secretion. Cont., control; Veh., vehicle. (D) IL-18 secretion in HCV-infected Huh7.5 cells. (E) NLRP3, in the context of HCV infection, increases exosome secretion nearly 25-fold, and this secretion is blocked by the expression of an ncRILP. (F) IL-18 secretion in LPS/ATP-treated THP-1 cells. (G) RILP is cleaved only after full inflammasome activation using both LPS and ATP. Ut, untreated. (H) Inhibition of either RILP function (ncRILP) or inflammasome activation (BHB) decreases LPS/ATP-induced exosome secretion. (I) The caspase-1 inhibitor VI (Z-YVAD-FMK, 100 µM) blocks LPS/ATP-induced exosome secretion. The caspase-4 inhibitor (Ac-LEVD-CHO, 100 µM) has no effect. (J) shRNA to NLRP3 inhibits LPS/ATP-induced exosome secretion, while shRNA to Gasdermin D has no effect. Data are shown as mean ± SD; *, P ≤ 0.05 for n = 3–8.