Fig 2: cGAS and STING inhibition prevents neuronal loss and rescues the transcriptional activation of IEGs upon neuronal excitatory input. (a‐d) WT and A‐T BOs were treated for one month with aspirin (4mM) or H‐151 (3 µM) starting at 5 months of development. (a) Immunofluorescence representative images of BO sections stained for NeuN (green) and GFAP (red). Scale bar, 0.7 mm. (b,c) BO sections stained for NeuN (b) and GFAP & p21 (c) were quantified. Each point in the scatter plot represents a single BO section analysed. Error bars represent SD; n = 4 independent experiments. ***p < 0.001; one‐way ANOVA with Tukey's multiple‐comparison post hoc corrections. (d) A‐T BOs were treated with NMDA (100 µM), KCl (55 mM) and Bicuculline (50 µM) for 30 min. Immediately after, BOs were collected for RNA extraction. RT‐qPCR analysis of the indicated IEGs was performed, and B2M mRNA was used as normalizer. Bar graphs show fold induction of IEG levels in stimulated organoids relative to untreated (no neuronal‐stimulating drug, depicted by a grid line). Error bars represent SD; n = 3 independent experiments. *p < 0.05; **p < 0.01, Each bar was compared to grid line values. Student's t test

Fig 3: Inhibition of cGAS and STING reduces tissue degeneration and cellular senescence in brain organoids. (a–e) WT and A‐T BOs where treated for one month with aspirin (4mM) or H‐151 (3 µM) starting at 5 months of development. (a) Each point in the scatter plot shows the organoid size of a single BO. At least 8 organoids per condition were analysed. ***p < 0.001; one‐way ANOVA with Tukey's multiple‐comparison post hoc corrections. (b) Representative images from quantifications shown in c. Scale bar, 0.7 mm. (c) SA‐β‐gal assays were performed on WT and A‐T BOs with the indicated treatments. Each point in the scatter plot represents a single BO section analysed. n = 3 independent experiments. *p < 0.05; ***p < 0.001; one‐way ANOVA with Tukey's multiple‐comparison post hoc corrections. (e) Total RNA from WT and A‐T BOs were used for RT‐qPCR analysis. B2M mRNA was used as normalizer. n = 3 independent biological samples. Exact P values can be found in Figure S4

Fig 4: Structural basis for phospho-regulation of STING recognition by AP-1.a, Three-dimensional (3D) reconstructions of the complex in two different orientations. The atomic models of dimeric human LBD-STING (Protein Data Bank (PDB) code: 4KSY) and the AP-1 complex (PDB 6DFF) were fitted into the maps (grey) through rigid-body docking. b, High-resolution 3D reconstruction from focused refinement on the AP-1 core and pSTING tail at 2.34-Å resolution contoured at 3s. c, Ribbon representation of the AP-1 pSTING complex structure. d–f, Detailed views of the binding interface. e, Potentialhydrophobic interactions around the EXXXLI motif are indicated with dotted lines. The density map (grey mesh) of pSTING tail is contoured at 3s. f, Potential hydrogen bonds around pS366 indicated with dotted lines. The density map (grey mesh) of the pSTING tail is contoured at 3s. g, HEK293T cells transfected with Flag-tagged STING and HA-tagged wild-type AP-1s or AP-1s mutants sKR (K60A/R61A), s(I103S) and s(V88D) were stimulated with diABZI for 2 h. Cell lysates were extracted, immunoprecipitated with anti-Flag antibody and analysed by western blot. h, WCL and CCV fractions from untreated and diABZI-treated HeLa STING KO cells reconstituted with Flag-tagged wild-type STING and the indicated STING mutants STINGLI, STINGLR(L374A/R375A) or STING(S358A) were analysed by western blot. CHC and GAPDH were used as loading controls. i, HeLa STING KO cells reconstituted with Flag-tagged STING and the indicated STING mutants STING(S358A), STING(S366A) or STING(S358/366A) were stimulated with diABZI for 2 h or left untreated. After immunoprecipitation with anti-Flag antibody, samples were analysed by western blot. j, Schematic diagram of the function of AP-1 in the termination of STING signalling. One representative example of at least three (g–i) independent experiments is shown.

Fig 5: AP-1 depletion boosts STING-dependent immune activation in different cell types.a,b, HaCaT cells (a) or THP-1 cells (b) transfected with NC siRNA or AP-1 siRNA for 3 days were treated with 2.5 µM diABZI for indicated time and analysed by western blot. GAPDH was used as a loading control. c,d, Human primary epithelial cells incubated with NC siRNA or AP-1 siRNA for 3 days were treated with 2.5 µM diABZI and were analysed by RT–qPCR (c) and western blot (d). Vinculin was used as a loading control. e–j, Fibroblast cells derived from patients with SAVI characterized by STING1 N154S mutation (e,f), H72N (g,h) or V147M (i,j) were incubated with NC siRNA or AP-1 siRNA for three days and then analysed by western blot. Vinculin was used as a loading control in f,h,j. e,g,i, Induction of IFNB1, IFIT1, IFIT2, IFIT3 expression was assessed by RT–qPCR in HeLa cells transfected with NC siRNA or AP-1 siRNA for 3 days and then stimulated with 1 µM diABZI. Mean ± s.d. of three (c,e,g,i) technical replicates. P values based on two-tailed Student’s t-tests (c,e,g,i). Source Data