Uncovering DIP-α signaling pathways underlying circuit assembly - PROJECT SUMMARY / ABSTRACT All animal behaviors and cognition require precise assembly of neural circuits. Despite a highly complex environment in the central nervous system, neurons faithfully recognize their precise partners and establish synaptic connections. Precise connectivity has been well demonstrated across many organisms, but the mechanisms underlying this specificity remain unclear. Cell surfaces proteins (CSPs) have been implicated in establishing correct connectivity, specifically by serving as “identification tags”. In Drosophila, two CSP subfamilies of the immunoglobulin superfamily (IgSF), the Dprs and DIPs, have garnered significant attention due to their multifaceted roles in nervous system development. The 32 members of the Dpr and DIP subfamilies are GPI-anchored, and several interacting pairs were demonstrated to have roles in instructing connectivity in several circuits. For example, DIP-α is required for instructing connectivity between motor neurons (MNs) and muscles in the motor system and between interneurons (INs) in the visual circuit. Our preliminary data suggests that DIP-α localizes to the dendrites of MNs as well, suggesting a potential role in IN-MN recognition. Despite their fundamental roles in various circuits, the signaling mechanism(s) underlying DIP/Dpr functions remains unclear. This proposal will test two non-mutually exclusive hypotheses: Aim 1) DIPs and Dprs instruct IN-MN connectivity and Aim 2) DIPs and Dprs interact with other CSP co-receptors to transduce cellular signals. I will focus on DIP-α because of its implications in connectivity, cell survival, and synaptic development; however, I hypothesize that some signaling components will be shared between Dpr/DIP members. In Aim 1, I will reconstruct MN dendrite morphology and synaptic connectivity between a MN and its presynaptic INs. I will determine if known DIP-α interactors, Dpr6/10, are required and investigate the functional outcome of disrupting DIP-α-dependent connectivity. In Aim 2, I will use proximity labeling to uncover candidate DIP-α co-receptors in an unbiased manner, and I will validate them biochemically and genetically. This proposal will combine interdisciplinary and innovative approaches, including optogenetics, electrophysiology, biochemistry, microscopy, proteomics, and bioinformatics to elucidate fundamental mechanisms underlying synaptic connectivity. The proposed work will address significant knowledge gaps in central motor circuit connectivity and signaling mechanisms of GPI-anchored proteins. The vertebrate orthologs of DIPs/Dprs, the IgLONs, are also GPI-anchored and are associated with various diseases, including Alzheimer’s disease and autism spectrum disorder. Thus, our proposed study may also contribute to our understanding of the molecular processes disrupted in specific neurological disorders.
