From neural border to migratory stage: A comprehensive single cell roadmap of the timing and regulatory logic driving cranial and vagal neural crest emergence

Neural crest cells exemplify cellular diversification from a multipotent progenitor population. However, the full sequence of molecular choices orchestrating the emergence of neural crest heterogeneity from the embryonic ectoderm remains elusive. Gene-regulatory-networks (GRN) govern early development and cell specification towards definitive neural crest. Here, we combine ultra-dense single cell transcriptomes with machine-learning and large-scale experimental validation to provide a comprehensive GRN underlying neural crest fate diversification from induction to early migration stages. During gastrulation, a transient neural border zone state precedes choice between neural crest and placodes following a "dual convergence model". Transcription factor connectome and bifurcation analyses demonstrate the early emergence of neural crest fates at neural plate stage, alongside an unbiased multipotent lineage persisting until after epithelial-mesenchymal transition. We decipher the circuits driving cranial and vagal neural crest formation and provide a broadly applicable strategy for investigating SC transcriptomes in vertebrate GRNs in development, evolution and disease.