Innate Immune Priming by cGAS as a Preparatory Countermeasure Against RNA Virus Infection

Michael T. Parker; Smita Gopinath; Corey E. Perez; Melissa M. Linehan; Jason M. Crawford; Akiko Iwasaki; Brett D. Lindenbach

For RNA viruses that replicate in the cytosol without a DNA intermediate, the 98 specific ligands that activate cGAS remain unclear. At present, the prevailing 99 hypothesis is that RNA viruses induce release of mitochondrial DNA (mtDNA) into 100 the cytosol, thereby activating innate immune responses (7, (33) (34) (35) (36) . However, it is 101 unclear whether mitochondrial damage is a conserved feature of RNA virus 102 infection, nor is it clear that cGAS-STING activation follows the same pathway for 103 both RNA and DNA viruses. 104 In this study, we investigated whether the DNA binding and cGAMP synthesis 105 activities of human cGAS (hcGAS) are required for RNA virus restriction. While both 106 activities were required, the amount of cGAMP produced during virus infection was 107 too low to detect. We also confirmed that hcGAS binds mtDNA in both uninfected 108 and infected cells but did not observe increased cytosolic or cGAS-associated 109 mtDNA in response to RNA virus infection. We found that cGAS stimulated 110 smoldering, low-level innate immune activation, most likely in response to 111 endogenous DNA ligands, suggesting that cGAS-STING can passively restrict 112 incoming RNA viruses. 113 . CC-BY-NC-ND 4.0 International license author/funder. It is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/434027 doi: bioRxiv preprint virus infection or, as a positive control, DNA transfection, by using liquid 172