Distal Enhancer-Insulator Module of GDF6 is Essential for Cochlear Formation [ChIP-Seq]

Inner-ear malformations cause congenital hearing loss, and several genes are known to guide inner-ear development. However, the contribution of noncoding regulatory elements remains largely unclear. This study investigates the function of distal enhancer elements in the transcriptional regulation of GDF6, a gene crucial for cochlear development. Using mouse models with targeted deletions, human iPSC-derived ear organoids (IEO), and CRISPR interference (CRISPRi) techniques, we identified a downstream regulatory interval harboring a developmental enhancer required to maintain GDF6 expression during otic epithelial maturation, vital for cochlear morphogenesis. Deletion of this regulatory region or targeting of CRISPRi-based repressors to these regions resulted in decreased GDF6 expression, failure of otic-epithelium development, and prevention of hair cell-like differentiation, reflecting cochlear aplasia observed in patients with corresponding genomic deletions. These findings highlight the essential role of long-range regulatory elements in auditory development and illustrate how their disruption contributes to human deafness.