High-throughput identification of the spatial origins of Drosophila optic lobe neurons using single-cell mRNA-sequencing

The medulla is the largest neuropil of the Drosophila optic lobe. It contains about 100 neuronal types that have been comprehensively characterized morphologically and molecularly. These neuronal types are specified from a larval neuroepithelium called the Outer Proliferation Center (OPC) via the integration of temporal, spatial, and Notch-driven mechanisms. Although we recently characterized the temporal windows of origin of all medulla neurons, as well as their Notch status, their spatial origins remained unknown. Here, we isolated cells from different OPC spatial domains and performed single-cell mRNA-sequencing to identify the neuronal types produced in these domains. This allowed us to characterize in a high-throughput manner the spatial origins of all medulla neurons. Overall design: We drove the expression of nuclear GFP in cells produced from the various mOPC spatial domains using spatial region-specific Gal4 drivers. We isolated the labeled cells by Fluorescence-Activated Cell Sorting, performed scRNA-seq on the sorted cells, and annotated the cells from each population using a neural network classifier. Optix: nuclear GFP positive optic lobe cells from the Optix-TS-MC>GFP line dOptix: nuclear GFP positive optic lobe cells from the dOptix>GFP line vOptix: nuclear GFP positive optic lobe cells from the vOptix-MC>GFP line hh: nuclear GFP positive optic lobe cells from the hh-TS-MC>GFP line dpp: nuclear GFP positive optic lobe cells from the dpp>GFP line pxb: nuclear GFP positive optic lobe cells from the pxb-MC>GFP line