Gene Regulatory Networks of Synaptic Specificity

Project Summary/Abstract Neuronal type identity is central to the development and function of neural circuits, as it instructs both the connectivity of neurons as well as their synaptic and electrophysiological properties. Neuronal fates are thought to be controlled by combinations of transcription factors (TF) known as terminal selectors, but very little is known about the gene regulatory mechanisms that link differential TF expression to specific neuronal features. The Drosophila visual system, a well-characterized brain region that has an organization analogous to the vertebrate retina and cortex, provides the ideal balance of complexity and accessibility to investigate these mechanisms. The first aim of this project will be to decipher the terminal selector TF codes that establish and maintain the unique identity of approximately 200 neuronal types that make the Drosophila optic lobes. Using a single-cell RNA sequencing (scRNA-seq) dataset I generated from developing optic lobes, I identified the combinations of TFs that are stably maintained in each neuronal type throughout their differentiation. Under the mentorship of Claude Desplan (K99 phase), I will test the hypothesis that these TFs function as terminal selectors by modifying the TF codes of specific optic lobe neurons in vivo, with the goal of predictably transdifferentiating them into other cell types. This will demonstrate the sufficiency of terminal selectors to confer neuronal identity and benefit the field of regenerative medicine. The conserved mechanisms in mammalian systems could be exploited to induce differentiation of pluripotent cells into specific neurons that could be transplanted to treat blindness or neurodegeneration. The second aim of this project will link the terminal selector TFs to their downstream targets. In collaboration with Richard Bonneau, I will learn to use the “Inferelator” algorithm to generate computational models of gene regulatory networks by combining my existing scRNA-seq data with new chromatin accessibility (scATAC-seq) data I will acquire. During the K99 phase, I will test the effects of perturbating key predicted downstream effectors on the morphology and connectivity of a select group of neurons to establish proof-of- concept. I will then generalize this approach in the R00 phase by inferring gene regulatory networks for all optic lobe neurons at multiple developmental stages. The third aim will be performed in my independent lab (R00) to utilize the network models for engineering precise modifications in visual circuits. I will seek to selectively uncouple the circuit that computes wide-field motion from the one that detects small moving objects. I will use synaptic tracing methods as well as intravital calcium imaging to demonstrate the functional consequences of developmental perturbations. Altogether, this project will establish direct mechanistic links between the encoding of neuronal identity and the molecules that mediate intercellular interactions during synaptic partner selection, which are commonly affected in neurodevelopmental disorders. The mentorship I will receive from Dr. Desplan and Dr. Bonneau, combined with the impressive resources of New York University provide the ideal environment for preparing me to build a successful independent research program that link gene regulation to brain wiring.