By identifying pseudouridine as a biochemical address tag — written by PUS1 and read by MYL6 — that selects specific RNAs for extracellular export, this paper reveals the first chemical code governing intercellular RNA communication.
Almost every cell secretes RNAs, including neurons, and yet the molecular mechanisms governing extracellular RNA (exRNA) export remain poorly understood. Using a novel genome-wide screen, we identified two unexpected classes of exRNA regulators: domesticated retroviral proteins, hijacked for neuronal RNA communication, and RNA-modifying enzymes. We discovered that pseudouridine (Ψ), installed by the pseudouridine synthase PUS1, is necessary and sufficient for the export of a subset of exRNAs that are recognized and packaged for export by the non-canonical RNA-binding protein MYL6. Our findings revealed a biochemical code that guides RNAs across cellular boundaries in the nervous system and beyond, representing an essential step toward deciphering the biological functions of exRNAs.
Pseudouridine modification by PUS1 functions as a biochemical code that selects specific RNAs for extracellular export, with the pseudouridine-binding protein MYL6 recognizing this modification to mediate RNA loading into extracellular vesicles, establishing a novel mechanism for intercellular RNA communication in neurons and other cell types.
RNAs move through the extracellular space to transmit information between cells, including mammalian neurons, yet how specific RNAs are channeled into these extracellular routes is unknown. Using genome-wide CRISPR screening, proteomics, and high-sensitivity transcriptomics in a neuronal model system, we identify domesticated retroviral proteins and RNA-modifying enzymes that regulate RNA loading into and transportation via extracellular vesicles. We show that the pseudouridine synthase PUS1 is a key determinant of RNA trafficking, and that its catalytic product in RNA, pseudouridine, is enriched in extracellular RNAs from transformed and primary neurons. Furthermore, the presence of pseudouridine on select RNAs is both necessary and sufficient for their extracellular export. Finally, we show that myosin light chain 6 (MYL6) is a pseudouridine-binding protein required for secretion of synthetic and endogenous RNAs. These findings reveal a biochemical code linking chemical RNA modification to extracellular transport, and establish a framework to study the function of extracellular RNAs in the nervous system and beyond.