We currently lack a comprehensive understanding of the mechanisms
underlying neural tube formation and their contributions to neural tube
defects (NTDs). Developing a model to study such a complex morphogenetic
process, especially one that models human-specific aspects, is
critical. Three-dimensional, human embryonic stem cell (hESC)-derived
neural rosettes (NRs) provide a powerful resource for in vitro modeling
of human neural tube formation. Epigenomic maps reveal enhancer elements
unique to NRs relative to 2D systems. A master regulatory network
illustrates that key NR properties are related to their epigenomic
landscapes. We found that folate-associated DNA methylation changes were
enriched within NR regulatory elements near genes involved in neural
tube formation and metabolism. Our comprehensive regulatory maps offer
insights into the mechanisms by which folate may prevent NTDs. Lastly,
our distal regulatory maps provide a better understanding of the
potential role of neurological-disorder-associated SNPs.