SOX9 Activates IRF6 in Neural Crest Cells

IRF6 is among the best-established genetic contributors to orofacial clefting – causative variants are found in at least 70% of people with Van der Woude syndrome, and common variants raise the risk of non-syndromic clefts. Its role has been attributed mainly to the oral epithelium, where TFAP2A drives expression through the enhancer MCS-9.7.

The authors now show that Irf6 protein is present in a subset of neural crest-derived mesenchymal cells in mouse pharyngeal arches, and single-cell RNA-seq data from human and mouse embryonic facial tissue show that most IRF6-positive mesenchymal cells also express SOX9, the main gene mutated in Pierre Robin sequence. Reporter assays, EMSA and ChIP indicated that SOX9 binds and activates MCS-9.7 via a site spanning the cleft-associated variant rs76145088, with the derived allele responding markedly less.

To test the dependence in a relevant cell type, the team used CRISPR-Cas9 to disrupt Sox9 in the murine cranial neural crest cell line O9-1, generating two clones with frameshift mutations. Both lacked Sox9 protein and Irf6 transcripts, while known upstream regulators of Irf6 showed no consistent change, pointing to direct regulation. Notably, the more SOX9-responsive ancestral allele is the risk allele, leading the authors to propose excess mesenchymal IRF6 expression as an alternative disease mechanism.

The study was led by Matthias Weider and Magdalena Wagner (equal contribution), at Universitätsklinikum Erlangen and Friedrich-Alexander-Universität Erlangen-Nürnberg. It was published in Journal of Molecular Medicine on 26 September 2026.