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

To validate that cell-derived cGAMP could be detected by this assay, a time-course 197 experiment was conducted by transfecting HEK 293E cells expressing WT hcGAS 198 with salmon sperm DNA, revealing the time-dependent increase in cGAMP (Figure 199 4J). Furthermore, we found that transfected cGAMP was rapidly turned over within 200 . 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 It is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the .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 hours (Fig. 4K ), most likely via the ENPP1 phosphodiesterase previously reported to 201 turnover cGAMP in mammalian cells (39) . Collectively, these results indicate that if 202 cGAMP is produced in response to RNA virus infection, it may be produced at levels 203 below the limit of our detection and/or rapidly turned over. 204 cGAS binds mitochondrial DNA at steady state and during RNA virus 205 infection. Given that cGAS DNA binding activity was also required for RNA virus 206 restriction, we sought to identify DNA ligands of cGAS during RNA virus infection. 207