Mechanisms and functions of the transcription factor broad as a terminal selector in the fly visual system - ABSTRACT It has been recently hypothesized that it is the persistent expression of certain transcription factors (TFs), called ‘terminal selectors’ that specify a neuron’s identity. Terminal selectors are thought to be expressed throughout development and adulthood, and determine a cell’s terminal characteristics, such as morphology, gene expression, and connectivity. However, in the majority of cell populations, the identities of terminal selectors, the characteristics they control, and the mechanisms by which they control each characteristic are still largely unknown. This proposal aims to establish mechanisms by which terminal selectors determine identity during development by using the well-studied model organism Drosophila melanogaster. Utilizing a single-cell RNA sequencing (scRNA-seq) transcriptional atlas of the fly visual system and knockdown/overexpression experiments, we identified a novel terminal selector, broad, which seemingly differentiates identity between two closely related visual projection neuron cell types. The terminal characteristics that differentiate these two cell types, including connectivity and function, are well established, and their experimental accessibility makes them an ideal model to study principles of terminal identity. We hypothesize that broad acts as a terminal selector between two cell types, acting to determine morphology, gene expression, and connectivity between two genetically similar cell types. To evaluate our hypothesis, in Aim 1 we first establish TFs that act as terminal selectors by (1) using knock-down and overexpression constructs to determine impacts on morphology, (2) utilizing scRNA-seq to determine impacts on gene expression and (3) performing whole-cell electrophysiology after hypothesized presynaptic inputs have changed to determine impacts on connectivity. In Aim 2 we will identify mechanisms underlying the changes in wiring patterns following terminal selector perturbations, utilizing (1) scRNA-seq, (2) DNA adenine methyltransferase identification (DamID) sequencing, and (3) knock- down/overexpression experiments. This fundamental research will expand the scientific community’s understanding of how neuronal identity arises and the minimal requirements for establishing identity characteristics. This work will also provide insight on how failures to appropriately specify neuronal cell types can contribute to neurodevelopmental and neuropsychiatric disorders.
