Dynamic, Single‐cell Monitoring of RNA Modifications Response to Viral Infection Using a Genetically Encoded Live‐cell RNA Methylation Sensor

Abstract RNA modifications, such as N6‐methylation of adenosine (m 6 A), serve as key regulators of cellular behaviors, and are highly dynamic; however, tools for dynamic monitoring of RNA modifications in live cells are lacking. Here, we develop a genetically encoded live‐cell RNA methylation sensor that can dynamically monitor RNA m 6 A level at single‐cell resolution. The sensor senses RNA m 6 A in cells via affinity‐induced cytoplasmic retention using a nuclear location sequence‐fused m 6 A reader. It allows for simultaneously measure RNA m 6 A dynamics and viral invasion at single‐cell level. Based on the single‐cell analytical tool, we found that SARS‐CoV‐2 infection enhances host‐cell RNA m 6 A level, and high‐level RNA m 6 A modification in host cells, in turn, facilitates viral infection. Particularly, Omicron, a variant of SARS‐CoV‐2, that features as high infection capacity, however, exhibits a reduced facilitation of m 6 A modification in host cells. In addition, the sensor can estimate viral inhibition via measuring cellular m 6 A level, that was explored for screening potential antiviral drugs. The methylation sensor can serve for elucidating the interplay between pathogens and host‐cell epigenetics at single‐cell level.