Single-cell RNA sequencing of motoneurons identifies regulators of synaptic wiring in Drosophila embryos

The correct wiring of neuronal circuits is one of the most complex processes in development, since axons form highly specific connections out of a vast number of possibilities. Circuit structure is genetically determined, but the mechanisms guiding each axon to precisely innervate a unique target cell are poorly understood. We investigated Drosophila embryonic motoneurons using single cell genomics, imaging and genetics. We show that a cell specific combination of homeodomain transcription factors and downstream Immunoglobulin domain proteins is expressed in individual cells and plays an important role in determining specific connections between motoneurons and muscles. We provide genetic evidence for a functional role of five homeodomain transcription factors and four immunoglobulins in the neuromuscular wiring. Knockdown and ectopic expression of these homeodomain transcription factors induces synaptic wiring defects specific to single cells that are in line with their spatial expression patterns and partly phenocopied by genetic modulations of their Immunoglobulin targets. Thus, our data strongly suggest that homeodomain transcription factor and Immunoglobulin molecule expression are directly linked and a major determinant of neuronal circuit structure.