Single-cell transcriptomic analysis unveils the diversity within mammalian spinal motor neurons

Spinal motor neurons (MNs) integrate sensory stimuli and brain commands to generate movements. In vertebrates, the molecular identities of the cardinal MN types such as those innervating limb versus trunk muscles have been well elucidated. Yet the identities of finer subtypes within these cell populations that innervate individual muscle groups remain enigmatic. Using single-cell transcriptomics, we have investigated heterogeneity in mouse MNs and discovered many unreported MN subtypes. Among limb-innervating MNs, we reveal a surprisingly diverse neuropeptide code for delineating putative motor pool identities. Additionally, we have uncovered that axial MNs are subdivided into three molecularly distinct subtypes, defined by mediolaterally-biased Satb2, Nr2f2 or Bcl11b expression patterns with different axon guidance signatures. These three subtypes are present in chicken and human embryos, suggesting a conserved axial MN expression pattern across higher vertebrates. Overall, our study provides a comprehensive molecular resource of spinal MN types and paves the way towards deciphering how neuronal subtypes evolved to accommodate vertebrate motor behaviors associated with different lifestyles. Single cell RNA seq of e13.5 mouse spinal motor neurons at rostral (brachial and thoracic vertebral segments, from C4 to T3) and caudal (lumbosacral, L1 to S5) segments. Motor neurons were labeled in GFP and sorted via FACS, and loaded onto 10x Genomics Single Cell 3’ Chip as two samples.