Abstract

Innate immune activation is a major driver of unmodified in vitro–transcribed (IVT) mRNA degradation; however, how modified IVT mRNAs are degraded, and the related regulation mechanisms, remain poorly understood. Through a focused screen of viral- and host-derived immune suppressors, we identify 13 factors that enhance mRNA performance, with SOCS1 and the coronaviral membrane protein (M) emerging as the most potent. Multi-omics analyses reveal that pseudouridine-modified IVT mRNA undergoes rapid deadenylation and predominant 3′−5′ decay, followed by bidirectional degradation, closely resembling endogenous mRNA decay kinetics, and is extensively associated with canonical mRNA decay machineries. Mechanistically, IVT mRNA activates IFN-β signaling, which promotes processing body (P-body) formation and XRN1-mediated 5′−3′ degradation. Suppression of IFN signaling by SOCS1 or M markedly enhances mRNA expression across diverse cell types, organoid systems, and murine disease models. Together, these findings define a type I interferon–P-body–XRN1 axis that constrains modified IVT mRNA stability and provides a framework for enhancing mRNA therapeutics. How innate immune signaling affects the stability of modified mRNA therapeutics remains to be explored. The authors here identify a type I interferon–P-body–XRN1 axis that limits modified IVT mRNA stability.