Decoding regulatory programs underlying placode and dermal condensate cell fate commitment during hair follicle induction via single-cell multi-omics analysis

Abstract

Understanding differences in chromatin state and changes in gene regulatory landscape of placode (Pc) and dermal condensate are crucial for decoding hair follicle (HF) morphogenesis programs. To identify cell-type-specific chromatin accessibility patterns in the developing HF, we integrated chromatin accessibility and transcriptome profiles at single-cell resolution during the murine HF induction stage. We applied unbiased analyses to identify seven major HF cell types and reclustered dermal (Der) and epithelium (Epi) subtypes to trace their cell fate specification. Our analysis showed that gene regulation in Der and Epi lineages is largely determined by cis-regulatory elements that direct gene expression in response to specific developmental cues. The chromatin accessibility of Twist2, Enpp2, Dkk1, and Sox2 varied from fibroblasts (Fb) to pre-DC lineage, while that of Edar, Lhx2, and Wnt10b varied from Epi to Pc lineage. Cell-type-specific enrichment of transcription factor binding motifs implicated Twist2 and Nfatc4 as key regulators in Fb to pre-DC fate specification, and Fos, Bach1, and Klf1 in Epi to Pc niche fate specification. Additionally, alignment of cell-type-specific peaks to super-enhancer databases identified key regulatory elements in both lineages. We identified and validated the critical cis-regulatory elements in pre-DC and Pc fate specifications through embryonic dorsal skin culture in vitro, suggesting that these elements may regulate critical genes essential for HF induction. Overall, our results provide a foundation for a comprehensive analysis of gene regulatory programs that initiate HF development, offering insights into the molecular mechanism of HF morphogenesis and clinical treatments of alopecia by skin grafts.