Creating a transcriptomic landscape of the Enteric Nervous System
Reena Lasrado1*, Jens Kleinjung1, Vassilis Pachnis1
1The Francis Crick Institute, 1 Midland Road, London NW1 1AT, United Kingdom
Development and functional assembly of the nervous system is dependent upon progenitor expansion and differentiation into neurons and glial cells. The Enteric Nervous System (ENS) is essential for digestive function and gut homeostasis. In contrast to the central nervous system, the ENS originates from neural crest cell progenitors (ENCC) and give rise to ganglionic structures that have no obvious spatial or topographic organization. Despite this chaotic distribution, recent progress in understanding cellular mechanisms that regulate development and connectivity of ENS lineages using reporter lines and fate-mapping has been achieved. However, mechanisms that control the cascade of molecular events from progenitors to pertinent neuronal and glial lineages and their contribution to reproducible organized activity remain obscure. To identify transcriptional signatures of neuronal lineage restriction and differentiation, we used a microfluidic platform to perform single-cell RNA sequencing (scRNA-seq) on FACsorted individual cells at two stages (embryonic and adult) that encompass an early critical period of ENS histogenesis and a late homeostatic period of ENS neuronal function. Using an unbiased approach to reduce the dimensionality of the data we observed that the transcriptomic profile at the embryonic stage identified two diverging cellular states. Semi-supervised biplot and hierarchical clustering analysis showed the emergence of two lineages defined by the expression of progenitor/gliogenic (Erbb3, Sox10, Fabp7 and Plp1) or progenitor/neurogenic (Tubb3, Elavl4, Ret and Phox2b) marker genes. Analysis at the adult stage classified cells into distinct groups outlining the diversity of ENS neuronal populations. We aimed to reconstruct the molecular states that define the developmental and cellular hierarchy of ENS progenitors along the neuronal lineage. Together, our single-cell gene expression profiling provides a molecular view of ENS progenitor dynamics at an early critical period of ENS development, identifies potential regulators of enteric neurogenesis and gliogenesis and delineates molecularly distinct neuronal subtypes.