Initiation and Regulation of Antiviral Innate Immunity

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Project Summary The goal of this proposal is to identify how the pattern recognition receptor cyclic GMP-AMP synthase (cGAS) operates as a high-fidelity DNA sensor that drives interferon (IFN) and inflammatory responses. In humans and mice, cGAS synthesizes the second messenger cyclic GMP-AMP (cGAMP) upon binding DNA. This second messenger stimulates immune responses upon binding the downstream regulatory protein STING. As such, cGAMP is considered a next-generation adjuvant. Because cGAS detects the sugar-phosphate backbone of double stranded DNA, a major question relates to how this enzyme is regulated to ensure inflammatory responses are selectively generated in response to viral (but not self) DNA. All cGAS proteins in nature contain a C-terminal catalytic domain that is necessary and sufficient for DNA binding and cGAMP production. All cGAS proteins also contain an N-terminal domain of poorly defined function. In the previous funding period, we discovered that the cGAS N-terminal domain functions to prevent self-DNA reactivity within human cells. Despite this insight into cGAS regulation, much remains unclear about its activities in infectious and non-infectious contexts. Indeed, we have discovered striking species-specific diversity of regulatory mechanisms that control cGAS activities. Understanding regulatory events that drive cGAS-mediated inflammatory responses will enable discussions of how protective immunity can be stimulated against infection and cancer. In this application, we propose to leverage natural cGAS diversity to explain the mechanisms of DNA-induced IFN responses (Aim 1), and leverage synthetic biology to further diversify cGAS into a “guard protein”, which provides a novel means of probing host-virus interaction (Aim 2).