Translational control of a complete developing sensory circuit

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Project Summary Development of the nervous system requires the generation of diverse neuronal types that subsequently drastically alter their shape and physiology to form vast interconnected networks. Each process requires precise gene regulation. Technological innovations have drastically increased the scale at which we can distinguish neuronal identities or stages based on transcriptomics or epigenetics. However, principles guiding post-transcriptional control of gene expression, which is essential for neural development and homeostasis, by regulatory factors including RNA-binding proteins (RBP) and miRNAs has not been defined at a similar systematic level. As numerous neurodegenerative disorders have been linked to impaired RBP-RNA interactions, and there is a huge need for improved cell type engineering strategies for therapeutics, insight into this process is critically important. I propose to use the Drosophila visual system as a model to study post- transcriptional regulation during specification and wiring of an entire neural sensory system. Specifically, this proposal focuses on the regulation of mRNA translational repression, which is a conserved feature of both neuronal fate diversification and differentiation in vertebrate and invertebrates. A single-cell transcriptomic atlas of the developing visual system, or optic lobe, was recently described which defines the transcriptome of each neuronal type in the optic lobe throughout development. Consistent with results from other model systems, many genes associated with terminal neuronal function were detected at transcripts in the immature neurons well before they are functionally required, and the corresponding protein was absent for two selected genes studied. I aim to define the scope of translational regulation in this system by first performing whole-tissue and select cell-specific ribosome profiling of the optic lobe over development, and determine the upstream control of this process using a combination of bioinformatics and genetics (Aim 1). Next I will tease apart the molecular and cellular mechanism by which two genes associated with different neuron signaling pathways are repressed at the translational level to assess how their expression is coordinated (Aim 2). Finally, I will adapt a single-cell translation profiling technique, scRibo-STAMP, in the Drosophila visual system to profile translation of all optic lobe neurons during specification and wiring. Combined with machine-learning based analysis, I will predict RBP/miRNA-RNA target interactions to gain fundamental insight into how RNA regulatory networks are shaped (Aim 3). Together this study will provide significant insights into the role of translational regulation during formation of the nervous system relevant to both human development and health.