Extended Data Fig. 1: Virus infection or 2’3’-cGAMP induces STING ER-associated condensate formation.
From: The STING phase-separator suppresses innate immune signalling
a, STING expression in HeLa TMEM173−⁄−-STING-GFP cell (hereon HeLa-STING-GFP), COS-7-STING-GFP, THP-1 cell and HT1080 cell was analyzed by western blot using antibodies against STING and GAPDH. b, HeLa-STING-GFP and COS-7-STING-GFP cells were stimulated with VACV or cGAMP as in Fig. 1a. The indicated proteins were analyzed at the indicated times by western blot with indicated antibodies. c, Representative images of STING condensate induced by cGAS activation. HeLa -STING-GFP were treated with 2 μg/ml 2’3’-cGAMP for 1 h or infected with indicated virus for 18 h. d, e, COS-7 cells expressing STING-GFP, mCherry-KDEL and LAMP1-iRFP (d) and COS-7 cells expressing STING-GFP, mCherry-KDEL and LC3B-iRFP (e) were activated by cGAS-transfection for 72 h. Representative confocal images showed STING condensates were co-localized with lysosomes (d) and autophagosomes (e). 293T cells transfected with plasmid expressing cGAS and STING-GFP for 72 h were stained by OTO method and analyzed by TEM. Representative images of puzzle-like structures digested by lysosomes (d) and autophagosome (e) were shown. f, 3D reconstruction of STING condensates in different cells. 293T and COS-7 cells were transfected with plasmid expressing cGAS and STING-GFP for 18 h. HeLa TMEM173−⁄−-STING-GFP and COS-7-STING-GFP cells were treated with VACV or 2’3’-cGAMP for 18 h or 1 h respectively. Series z-stack images of live cells were captured by confocal microscope and then 3D reconstruction was performed. Typical optical sections of z-stack were shown. g, COS-7 and 293T cells were transfected with plasmid expressing cGAS, STING-GFP and mCherry-KDEL. Live cell confocal images were taken. For all Extended Data Fig. 1, data were representatives of at least three independent experiments. Confocal images were representatives of fields, in which >95% of the cells displayed similar pattern. TEM images represented at least five cells with the same pattern. Scale bar, as indicated. Nuc, Nucleus. See uncropped gel images in source data.
