DNA-PK controls cyclic dinucleotide-associated type I Interferon responses

Cytosolic dsDNAs are potent immune-stimulatory molecules that trigger inflammation in several human pathologies ^1,2 . A major pathway for the detection of cytosolic dsDNA relies on the cyclic GMP-AMP (cGAMP) synthase (cGAS) that produces the 2’3’-cGAMP cyclic dinucleotide (CDN) for activation of the Stimulator of Interferon Genes (STING) adaptor protein that subsequently drives type I Interferon (IFN) responses ^3,4 . Here, we investigated the mechanism regulating intracellular 2’3’-cGAMP levels. We show that the DNA-dependent protein kinase catalytic subunit (DNA-PKcs), a major player in the repair of double-strand breaks, directly regulates intracellular levels of 2’3’-cGAMP, thereby reducing STING activation. We describe that the binding of 2’3’-cGAMP to DNA-PKcs occurs in its catalytic cleft, impeding its kinase function. Contrary to other CDN regulatory mechanisms that have been shown to primarily regulate extracellular 2’3’-cGAMP, we show that DNA-PKcs also interacts with the 3’3’-cGAMP bacterial CDN, limiting its capacity to activate STING signaling. Furthermore, we found that DNA-PKcs decreases the potency of pharmacological STING activators. As STING is a major target for therapeutic interventions aiming to boost inflammatory responses in immunosuppressed contexts ⁵ , our data bear important implications for drug development and deepens our understanding of inflammatory regulation in response to CDNs.