Coordination and propagation of cell fate choice in neural circuit assembly

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Project Summary How vast numbers of neurons are specified into correct cell fates and connected with proper targets during development represents a fascinating area of developmental neuroscience. Mechanisms of stochastic and deterministic cell specification programs to achieve neuronal diversity have been extensively studied. Over the last decades, a number of cell surface molecules have also been identified that mediate axon guidance and connectivity. However, little is known about the coordination between neuronal specification and specific connectivity patterns, especially when two synaptic partners undergo two different modes of cell specification (stochastic vs. deterministic). The Drosophila color vision circuit is an appealing model to address this question due to our deep knowledge of its development, its precise neuronal connectivity, and the availability of powerful genetic tools for cell-type specific manipulations. In the fly retina, pale (p) and yellow (y) subtypes of color photoreceptors (R7 and R8) are stochastically specified, whereas their synaptic partners in the optic lobe are produced through highly deterministic programs. How do stochastically determined p/y R7 and R8 find their targets that are deterministically specified in the optic lobes? How is this decision propagated to their downstream targets during circuit formation? What molecules direct these events? Previous work from our lab has identified Dpr11 and DIPg, which are members of an interacting network of immunoglobulin superfamily proteins, as critical regulators of the synaptic connection between yR7 and its downstream target. We hypothesize that different pairs of cell adhesion molecules mediate the matching of other synaptic partners. By using single-cell RNA sequencing technology, CRISPR gene editing, and sophisticated genetic manipulation in the Drosophila color vision circuit, we aim to identify cell adhesion molecules that direct synaptic partner matching and the molecular logic for coordinating between cell-type specification and the synaptic connectivity at the system level. We will define the synaptic connectivity as well as generate cell-type specific transgenic reagents and higher-depth transcriptomes of relevant cell types (Aim 1). We will use a candidate approach combined with transcriptome analysis of sorted targeted neurons of color photoreceptors to identify the molecules required for synaptic partner matching (Aim 2). We will compare whether a given neuron uses the same or different molecular codes for matching its pre- and post-synaptic partners. Finally, we will study how the synaptic partner choices propagate to neurons further downstream by perturbating the cell fates of R7 and R8 (Aim 3). Successful completion of this proposal will uncover novel molecular mechanisms regulating synaptic pairing and probe the fundamental principles underlying the propagation of cell fate choices during circuit assembly. The principles identified here will be significant and applicable to other neuronal circuits facing similar developmental challenges, such as the olfactory system in rodents and color vision in humans.