SARS-CoV-2 Nsp1-Resistant Modified RNA for the Creation of Nsp1-Responsive Systems

Modified RNA (modRNA) facilitates the introduction of complex synthetic genetic circuits into cells without the risk of genomic integration, opening up the implementation of synthetic circuits as therapeutics. However, the number of protein-RNA interfaces that are suitable for the construction of protein-responsive modRNA switches as well as the lack of protein-responsive exclusive selector systems stifles the development of RNA-based synthetic circuits. Here, we present the creation of a modRNA capable of resisting the effects of Nsp1 for the reliable expression of its coding sequence. Using both the subgenomic viral RNA 5'UTR and two modified nucleosides, we observed efficient exogenous protein expression even in Nsp1-transfected cells. To demonstrate its utility, we developed a barnase-barstar system capable of conditional transcript suppression in the presence of Nsp1. Altogether, the resistance to Nsp1-mediated translational suppression and the resulting Nsp1-sensing system we present in this study provide an invaluable opportunity to develop a new class of protein-sensing systems for the construction of more complex RNA-based genetic circuits.