RNA and Protein Networks That Locally Control Brain Wiring During Development

Abstract

The molecular machineries of growth cones control the formation of neural circuits in the developing brain. Although great progress has been made in elucidating axon guidance cues and their growth cone receptors, we still lack an understanding of the projection-specific RNA and protein networks in growth cones that likely control the wiring of specific circuits in vivo. To understand how specific projection neurons make wiring decisions, I focus on callosal projection neurons (CPN), which connect the two cerebral hemispheres through the corpus callosum. I developed an approach to profile and quantify the full-depth transcriptomes and proteomes of CPN growth cones and their parent cell bodies isolated in vivo. Using this comparative approach, I uncover general patterns of RNA and protein subcellular localization, with several previously unrecognized features, that might control the wiring of specific brain circuits. First, while most transcripts are expressed at similar levels in cell bodies and growth cones, a select subset are more than 10-fold enriched in growth cones compared to cell bodies, indicating active localization of those transcripts to the growth cone. By then correlating transcriptomic and proteomic data, I characterize the spatial relationship between coding transcripts and their encoded proteins. Intriguingly, many of the growth cone-enriched transcripts are noncoding RNA with unknown function. Further, growth cones appear to have distinct ribosomes. These ribosomes lack several large subunit proteins, raising the intriguing possibility of growth cone-specific translational mechanisms for selective mRNA expression. This approach is readily adaptable to other projection types in the brain, enabling high-throughput, quantitative investigation of RNA and protein controls over circuit development and, potentially, the regeneration of damaged circuitry. In addition, the approach is scalable to include epigenetic profiling, enabling full investigation of DNA, RNA, and protein networks that collectively coordinate brain wiring during development. The insights derived from this approach exemplify its capacity to quantify and characterize the molecular and translational mechanisms that control specific brain wiring at the subcellular level in vivo.