The developmental hourglass model is applicable to the spinal cord based on single-cell transcriptomes and non-conserved cis-regulatory elements.

The developmental hourglass model predicts that embryonic morphology is most conserved at the mid-embryonic stage and diverges at the early and late stages. To date, this model has been verified by examining the anatomical features or gene expression profiles at the whole embryonic level. Here, by data mining approach utilizing multiple genomic and transcriptomic datasets from different species in combination, and by experimental validation, we demonstrate that the hourglass model is also applicable to a reduced element, the spinal cord. In the middle of spinal cord development, dorsoventrally arrayed neuronal progenitor domains are established, which are conserved among vertebrates. By comparing the publicly available single-cell transcriptome datasets of mice and zebrafish, we found that ventral subpopulations of post-mitotic spinal neurons display divergent molecular profiles. We also detected the non-conservation of cis-regulatory elements located around the progenitor fate determinants, indicating that the cis-regulatory elements contributing to the progenitor specification are evolvable. These results demonstrate that, despite the conservation of the progenitor domains, the processes before and after the progenitor domain specification diverged. This study will be helpful to understand the molecular basis of the developmental hourglass model.