Nonspecific DNA Binding of cGAS N Terminus Promotes cGAS Activation

The cytosolic DNA sensor Cyclic GMP-AMP Synthase (cGAS) mediates innate immune responses against invading pathogens, or against self-dsDNA, which causes autoimmune disorders. Upon nonspecific binding of cytosolic B-form DNA, cGAS synthesizes the second messenger 2'3'-cGAMP and triggers STING-dependent signaling to produce type I IFNs. The cGAS comprises less-conserved N-terminal residues and highly conserved nucleotidyltransferase/Mab21 domains. The function and structure of the well-conserved domains have been extensively studied, whereas the physiological function of the N-terminal domain of cGAS is largely uncharacterized. In this study we used a single-molecule technique combined with traditional biochemical and cellular assays to demonstrate that binding of nonspecific dsDNA by the N-terminal domain of cGAS promotes its activation. We have observed that the N terminus of human cGAS (hcGAS-N160) undergoes secondary structural change upon dsDNA binding in solution. Furthermore, we showed that the hcGAS-N160 helps full length hcGAS to expand the binding range on λDNA and facilitates its binding efficiency to dsDNA compared with hcGAS without the 160 N-terminal residues (hcGAS-d160). More importantly, hcGAS-N160 endows full length hcGAS relatively higher Enzyme activity and stronger activation of STING/IRF3-mediated cytosolic DNA signaling. These findings strongly indicate that the N-terminal domain of cGAS plays an important role in enhancing its function.