Extended Data Fig. 9. 3′2′-cGAMP induces the expression of dSTING-regulated genes.

a–d, Injection of 3′2′-cGAMP into D. melanogaster has a dose-dependent effect on the expression of Sting-regulated genes (srgs). 2′3′-cGAMP was used as positive control as previously characterized^23,26. Synthetic nucleotide was injected into the body cavity of WT (w¹¹¹⁸) flies and gene expression was measured after 24 h. RNA levels were measured relative to the control gene RpL32, and nucleotide concentrations are displayed in μg μl⁻¹. Note that for srg2 measurement after injection of 9E−7 μg μl⁻¹ 3′2′-cGAMP, there was one outlier replicate with a value of 0.5977 (data not shown, included in mean analysis). e–k, As in Fig. 4a, RNA expression analysis of Sting-regulated genes (srgs) 24 h after injection with synthetic 3′2′-cGAMP or 3′3′-c-di-GMP. RNA levels are shown as fold induction compared with buffer control in WT flies. dSTING^Mut = RXN mutant; Relish^Mut = Relish^E20 mutant, as previously characterized^23,26. All data in a–k represent the mean ± s.e.m. of n = 3 independent experiments and each point represents a pool of 6 flies. P value ns (>0.05) unless otherwise noted: ^****P < 0.0001 (e); ^***P = 0.0006, ^*P = 0.0404 (f); ^****P < 0.0001, ^***P = 0.0002 (g); ^****P < 0.0001 (h); ^**P = 0.0015, ^*P = 0.0117 (i); ^***P = 0.0002, ^**P = 0.0076 (j); ^***P = 0.0009 (k).

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