mTRACE: Modification-enabled Tracking of RNA-associated Cellular Events

However, existing methods face limitations in continuously recording these events in living cells. Here we introduce mTRACE (Modification-enabled Tracking of RNA-associated Cellular Events), a molecular recording framework that leverages RNA modification enzymes to capture the history of RNA-associated cellular processes.

By systematically screening 34 candidate enzymes, we identified N6-adenosine methyltransferase M.EcoGII as a highly efficient and unbiased modifier. mTRACE exploits M.EcoGII to install natural and non-disruptive RNA modification, writing durable marks on transcripts undergoing specific molecular events in living cells. We demonstrate that mTRACE enables retrospective tracking of RNA-protein interactions, quantification of translational activity and efficiency by coupling M.EcoGII to ribosomal proteins, and spatial recording of RNA residence across multiple subcellular compartments using engineered M.EcoGII variants. mTRACE provides a general framework for dissecting RNA dynamic regulation and supports multi-dimensional recording of RNA-associated cellular events with spatiotemporal resolution.