Fig 1: cGAS/STING pathway activation is caused by general deficiency in COPI-mediated retrograde trafficking.a Schematic illustration of coatomer complexes COPII (green) and COPI (blue) mediating intracellular trafficking between endoplasmic reticulum (ER) and Golgi compartments. The COPII complex mediates anterograde transport from ER to Golgi, whereas COPI mediates retrograde transport from Golgi to ER as well as within cis-Golgi compartments. COPI vesicle formation occurs at the Golgi membrane through interaction of 7 subunits: a (COPA), ß (COPB1), ß’ (COPB2), d (COPD), e (COPE), ? (COPG), ? (COPZ). After vesicle budding, the coatomer coats are shed off and released into the cytoplasm to allow vesicle fusion at the target membrane2. b Representative IF images of parental, COPAdeficient, COPG1deficient and COPDdeficient HeLa cells co-stained for KDEL (green), GM130 (magenta), indicated COPI subunit (cyan) and DAPI (blue). Images are presented as maximum intensity projection. Scale bar 10 µm. c Quantification of IF co-localization of KDEL and GM130 signal in COPI subunit-deficient HeLa cells shown in (b). Results are quantified as percentage area of KDEL localization (ER-specific retention signal) inside cis-Golgi (GM130). Each dot represents one single cell. Parental cells stained for different subunits were combined for quantification analysis. Statistical significance was assessed by one-way ANOVA with Dunnett’s multiple comparison test using n = 35 (parental), n = 8 (COPAdeficient), n = 15 (COPG1deficient) and n = 18 (COPDdeficient) biologically independent cells examined in 2 independent experiments. Line at median. d Immunoblot analysis of THP-1 cells after CRISPR/Cas9-mediated genetic deletion of COPI-subunit proteins COPA, COPG1, COPD and COPE following treatment with STING inhibitor H-151 (2.5 µM). Results are shown as a representative of n = 3 independent repeats e qRT-PCR analysis of proinflammatory genes and ISGs in COPI-subunit deficient THP-1 cells following H-151 treatment (2.5 µM). Data are shown as mean ± SEM from n = 3 independent experiments and statistical significance was assessed by two-tailed ratio paired Student’s t-test comparing Veh ctrl and H-151 treatment individually for each cell line. P values are indicated by numbers or as * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. Source data are provided as a Source Data file for Fig. 5c, d.