Single Cell Profiling in the Sox10Dom Hirschsprung Mouse Implicates Hox Genes in Enteric Neuron Trajectory Allocation

Enteric nervous system (ENS) development requires migration, proliferation, and neuronal diversification of progenitors to achieve normal gastrointestinal motility. Sox10 deficits disrupt the ratios of mature enteric neuron subtypes. How Sox10 deficiency alters the proportions of neuronal subtypes is still being determined. Sox10's prominent expression in neural crest-derived enteric progenitors and lack of this gene in enteric neurons led us to investigate how Sox10 effects in early enteric progenitors could alter neuronal allocation. To explore this further, enteric progenitors, developing neurons, and enteric glia were isolated from Sox10+/+ and Sox10Dom/+ littermates at 15.5 days post-coitus based on dual Sox10-H2BVenus and Phox2b-H2BCerulean transgene expression for single-cell RNA sequencing (scRNA-seq). The data was processed in R to identify cell-specific markers, differentially expressed genes, biological processes associated with gene ontology, cell fate trajectories, and gene regulatory network activity between genotypes. Hybridization chain reaction (HCR) validated expression changes detected in scRNA-seq. Overall design: Myenteric neurons and glia were isolated from Phox2b-CFP, Sox10-YFP fetal intestine expressing Sox10Dom/+ Or Sox10+/+. The whole fetal intestine was dissociated, and single cells were isolated via FACS for single-cell encapsulation using 10X encapsulation methods. DNA libraries were generated and processed for next-generation sequencing.