mRNA transport, translation, and decay in adult mammalian central nervous system axons

Localized mRNA translation regulates synapse function and axon maintenance, but how compartment-specific mRNA repertoires are regulated is largely unknown. We developed an axonal transcriptome capture method that allows deep sequencing of metabolically-labeled mRNAs from retinal ganglion cell axon terminals in mouse. Comparing axonal-to-somal transcriptomes and axonal translatome-to-transcriptome enables genome-wide visualization of mRNA transport and translation and unveils potential regulators tuned to each process. FMRP and TDP-43 stand out as key regulators of transport, and experiments in Fmr1 knock-out mice validate FMRP's role in the axonal transportation of synapse-related mRNAs. Pulse-and-chase experiments enable genome-wide assessment of mRNA stability in axons and reveal a strong coupling between mRNA translation and decay. Measuring the absolute mRNA abundance per axon terminal shows that the axonal transcriptome is stably maintained in adulthood by persistent transport. Our datasets provide a rich resource for unique insights into RNA-based mechanisms in maintaining presynaptic structure and function in vivo. Overall design: Poly-adenylated RNAs in mouse retinal ganglion cell axons terminating at the superior colliculus or lateral geniculate nucleus. For the total RNA, EU (5'-Ethynyl Uridine) was injected into the vitreous to label newly transcribed RNAs and EU-labeled RNAs were affinity purified from the superior colliculus. For the ribosome-bound RNA, axon-TRAP was performed.