Disrupting IFIT1-STAT2 viral evasion synergy yields live-attenuated flavivirus with enhanced immune priming

Diverse human viruses deploy a common immune-evasion strategy: viral 2’-O-methylation coupled with STAT2 antagonism. Yet the importance of this coupling and whether targeting the coupled functions offers benefits in vaccine development, remains unknown. Using deep mutational scanning of the dengue NS5 protein, we discovered that the two immune-evasion functions work synergistically to cripple interferon signaling. Disrupting this synergy exposes a reciprocal regulatory relationship between the host proteins IFIT1 and STAT2 that controls both viral restriction and immune cell activation. Dengue viruses with mutations that disrupt the synergy are potently attenuated yet trigger antigen-presenting cell activation that exceeds a licensed dengue vaccine benchmark. Our results establish a generalizable principle: targeting synergies between viral immune evasion mechanisms can simultaneously achieve maximal viral attenuation and innate immune priming. This work provides a rational design framework for developing high-performance vaccines against many human viruses that exploit the IFIT1-STAT2 axis.