Phosphoinositide Regulation of Intracellular Nucleic Acid Sensors

Abstract

Innate immune sensing relies on pattern recognition receptors (PRRs) to bind molecular motifs unique to pathogens to initiate inflammation and the adaptive immune response. These PRRs are tightly regulated, particularly in regard to their localization and access to ligands. This spatial regulation of PRRs is well characterized for transmembrane proteins, such as those of the Toll-like receptor (TLR) family. However, little is known about the subcellular positioning of intracellular PRRs that lack transmembrane domains, which generally are assumed to be residents of the cytosol. To determine if these receptors, like their transmembrane counterparts, had specific subcellular positioning, we interrogated the localization of the intracellular nucleic acid sensors cyclic GMP-AMP synthase (cGAS) and Retinoic acid inducible gene I (RIG-I) and the role of this localization in regulating their sensory capacity. Our studies revealed that cGAS, an integral sensor of cytosolic DNA, localizes to the plasma membrane in the absence of stimuli in human and murine monocytes. This subcellular positioning occurs through an electrostatic interaction between the cGAS N terminus and the plasma membrane lipid phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2). Loss of the cGAS N terminus led to a loss of membrane localization and heightened interferon (IFN) signaling in the absence of infection. Mislocalized cGAS also generated heightened responses to genotoxic stress, but not viral infection. These data reveal that the subcellular positioning of this receptor serves as a mechanism of self-nonself discrimination by specifically inhibiting sensing of self- DNA. Similar to cGAS, we found that RIG-I, a key sensor of viral RNA, interacts with the membrane lipid PI(3,5)P2 in vitro and that this interaction inhibited RIG-I RNA binding and ATPase activity. Together, these data suggest that subcellular positioning through electrostatic interactions with phosphatidylinositol phosphates (PIPs) may be a common mechanism of localization and regulation for intracellular PRRs lacking transmembrane domains.