Effect of methyl DNA adducts on stimulation of human cytoplasmic DNA-sensor cyclic GMP-AMP synthase (cGAS)

The immune system uses a variety of DNA sensors, including endo-lysosomal Toll-like receptors 9 (TLR9) and cytosolic DNA sensor cyclic GMP-AMP (cGAMP) synthase (cGAS). These sensors activate immune responses by inducing the production of a variety of cytokines, including type I interferons (IFN). Activation of cGAS requires DNA-cGAS interaction. Accumulation of cGAMP activates the stimulator of interferon genes (STING), ultimately leading to pathogen clearance by type I IFN production. To prevent the sensing of endogenous nuclear DNA, cGAS is usually localized in the cytoplasm. In this work, we studied the interaction and activation of cGAS by DNA containing non-CpG methyl adducts N3-methyl-C (3mC) and 7-methyl-G (7mG). We report that while DNA with 3mC and 7mG interacts with cGAS, it fails to stimulate its activity in vitro. To gain mechanistic insight, we used synthetic oligonucleotides containing 3mC and 7mG for cGAS activation. We observed that the presence of these adducts was inhibitory to cGAS-catalyzed cGAMP production and type I IFN response in human monocyte cell line THP1. Thus, our study reveals that the specific DNA base methylation adducts 3mC and 7mG contribute to the regulation of cGAS activation and provide a potential strategy for delivering DNA without activating the cGAS pathway.