Human-specific transposable elements shaped the evolution of craniofacial development through regulation of neural crest migration

Craniofacial development and neural crest specification are evolutionarily conserved processes, yet subtle modifications to their gene regulatory networks drive species-specific craniofacial diversity. Transposable elements (TEs) are increasingly recognized as contributors to genome evolution, but their role in shaping neural crest regulatory programs remains underexplored. Here, we investigate the domestication of human-specific TEs as transcriptional enhancers during cranial neural crest cell (CNCC) specification, a process critical for vertebrate head development. Using human iPSC-derived CNCCs, we identified ~250 human-specific TEs acting as active enhancers. These TEs were predominantly LTR5Hs and, to a lesser extent, SVA-E/Fs. We demonstrate that these elements have been co-opted through the acquisition of the conserved CNCC coordinator motif, and are bound by the CNCC signature factor TWIST1, and that their co-option appears to be largely exclusive to CNCCs. To assess their functional relevance, we used CRISPR-interference to repress ~75% of all the LTR5Hs and SVAs active in CNCCs, which led to widespread transcriptional changes in genes associated with neural crest migration, a process essential for CNCCs to populate the embryo and form craniofacial structures. Using a cell migration assay, we showed that CNCC migration was significantly impaired by CRISPR-mediated TE repression. Finally, we demonstrate that genes near human-specific TEs are more highly expressed in human CNCCs relative to chimpanzee, and TE repression returns their expression to chimpanzee levels. These findings reveal how human-specific TEs have been co-opted to fine-tune CNCC regulatory networks, potentially contributing to the evolution of lineage-specific craniofacial traits.