Fig 1. STING interacts with NLRP3 to facilitate the inflammasome activation.

(A–C) HEK293T cells were co-transfected with pFlag-STING and pHA-NLRP3 (A and B), or with pFlag-STING, pHA-NLRP3, pHA-ASC, and pHA-Casp-1 (C). (D) Diagrams of NLRP3, PYRIN, NACHT, and LRR. (E, F) HEK293T cells were co-transfected with pHA-STING and pFlag-NLRP3, pFlag-PYRIN, pFlag-NACHT, or pFlag-LRR (E), or with pFlag-STING and pHA-NLRP3, pHA-PYRIN, pHA-NACHT, or pHA-LRR (F). (G) Diagrams of STING and its truncated proteins. (H, I) HEK293T cells were co-transfected with pHA-NLRP3 and pFlag-STING, truncated proteins (H) or TM5-deleted STING(delTM5) (I). (J, K) HEK293T cells were co-transfected with plasmids encoding NLRP3, ASC, pro-Casp-1, and pro-IL-1β to generate a pro-IL-1β cleavage system (NACI), and transfected with pFlag-STING, truncated proteins (J), TM2-deleted STING(delTM2), TM5-deleted STING(delTM5) or TM2 and TM5-deleted STING(delTM2 and 5) (K). IL-1β in supernatants was determined by ELISA. (L) HEK293T cells were co-transfected with pFlag-ASC, pHA-NLRP3, or pFlag-STING. (A–C, E, F, H, I, and L) Cell lysates were subjected to co-immunoprecipitation (Co-IP) using IgG (Mouse) and anti-Flag antibody (C, F), IgG (Rabbit) and anti-HA antibody (L), anti-HA antibody (B), and anti-Flag antibody (A, E, H and I), and analyzed by immunoblotting using anti-HA or anti-Flag antibody (top) or subjected directly to Western blot using anti-Flag or anti-HA antibody (input) (bottom). (M) HEK293T cells were co-transfected with pFlag-ASC, pHA-NLRP3, and pMyc-STING. Pellets were subjected to ASC oligomerization assays (top) and lysates were prepared for Western blots (bottom). (N, O) HEK293T cells (N) or Hela cells (O) were co-transfected with pFlag-STING and/or pHA-NLRP3. Sub-cellular localization of Flag-STING (green), HA-NLRP3 (red), and DAPI (blue) were examined by confocal microscopy. Data shown are means ± SEMs; **p < 0.01, ***p < 0.0001; ns, no significance.