A Molecular Approach to Identify Determinants of Synaptic Specificity

Information processing in the nervous system relies on precise patterns of synaptic connections between neurons. The level of synaptic specificity within the nervous system is remarkable given its tremendous complexity. How neurons can distinguish their correct synaptic partners from many other neurons during circuit assembly remains a central question in neuroscience. Although important progress has been made on molecular mechanisms regulating neural circuit assembly, the cellular recognition mechanisms mediating synaptic specificity are still poorly understood. The Drosophila visual system is well suited to uncovering the molecular mechanisms underlying synaptic specificity, because of the availability of diverse genetic tools, cell type specific markers and electron microscopic reconstruction data. In the medulla neuropil of the Drosophila visual system, different neurons form synaptic connections in different layers. Within a layer, neurons form synapses with a unique set of multiple neuronal types, which represents only a subset of neurons with processes in that layer. In my thesis research, I sought to identify candidate cell recognition molecules underlying this specificity. I did RNA sequencing on closely related neurons with different layer-specific synaptic specificities, lamina neurons L1-L5 and photoreceptor R7 and R8, at the onset of synapse formation. I showed that each of the seven cell types expresses a unique set of hundreds of genes encoding cell surface and secreted proteins. Using these data and additional localization studies on proteins tagged through modification of the endogenous locus, I demonstrated that 21 paralogs of the Dpr family, a subclass of Immunoglobulin (Ig) domain containing cell surface proteins, are expressed in unique combinations in each of the seven cell types during synapse formation. Dpr interacting proteins (DIPs), comprising nine paralogs of another subclass of Ig-containing proteins, are expressed in a complementary layer-specific fashion in subsets of synaptic partners for each of the seven cell types. Thus, I demonstrated that interacting Dprs and DIPs are expressed in synaptic partners during synapse formation in the Drosophila visual system. Furthermore, I generated null mutants of Dprs and DIPs via CRISPR-based methods, and showed that mutants of two DIPs and their cognate Dprs had similar phenotypes: neurons normally expressing these DIPs had reduced number of cells in corresponding DIP and cognate Dpr mutants. These data suggested that cognate Dprs and DIPs play important roles in the development of synaptic partners expressing them