Protein interaction is critical molecular regulatory activity underlining cellular functions and precise cell fate choices. Using TWIST1 BioID-proximity-labelling and network propagation analyses, we discovered and characterized a TWIST-chromatin regulatory module (TWIST1-CRM) in the neural crest cells (NCC). Combinatorial perturbation of core members of TWIST1-CRM: TWIST1, CHD7, CHD8, and WHSC1 in cell models and mouse embryos revealed that loss of the function of the regulatory module resulted in abnormal differentiation of NCCs and compromised craniofacial tissue patterning. Following NCC delamination, low level of TWIST1-CRM activity is instrumental to stabilize the early NCC signatures and migratory potential by repressing the neural stem cell programs. High level of TWIST1 module activity at later phases commits the cells to the ectomesenchyme. Our study further revealed the functional interdependency of TWIST1 and potential neurocristopathy factors in NCC development.