During tissue development, transcription factors bind regulatory DNA regions called enhancers, often located at great distances from the genes they regulate, to control gene expression. The enhancer landscape during embryonic stem cell differentiation has been well characterized. By contrast, little is known about the shared and unique enhancer regulatory mechanisms in different ectodermally derived epithelial cells. Here we use ChIP sequencing (ChIP-seq) to identify domains enriched for the histone marks histone H3 lysine 4 trimethylation, histone H3 lysine 4 monomethylation, and histone H3 lysine 27 acetylation (H3K4me3, H3K4me1, and H3K27ac) and define, for the first time, the super enhancers and typical enhancers active in primary human corneal epithelial cells. We show that regulatory regions are often shared between cell types of the ectodermal lineage and that corneal epithelial super enhancers are already marked as potential regulatory domains in embryonic stem cells. Kruppel-like factor (KLF) motifs were enriched in corneal epithelial enhancers, consistent with the important roles of KLF4 and KLF5 in promoting corneal epithelial differentiation. We now show that the Kruppel family member KLF7 promotes the corneal progenitor cell state; on many genes, KLF7 antagonized the corneal differentiation-promoting KLF4. Furthermore, we found that two SNPs linked previously to corneal diseases, astigmatism, and Stevens-Johnson syndrome fall within corneal epithelial enhancers and alter their activity by disrupting transcription factor motifs that overlap these SNPs. Taken together, our work defines regulatory enhancers in corneal epithelial cells, highlights global gene-regulatory relationships shared among different epithelial cells, identifies a role for KLF7 as a KLF4 antagonist in corneal epithelial cell differentiation, and explains how two SNPs may contribute to corneal diseases.

Highlights d Epidermal cells actively migrate into the interdigital mesenchyme d Active migration of epidermal cells is required for digit separation d Non-adhesive property of periderm cells prevents epithelial fusion between the digits d GRHL3 is required for normal periderm development and function

The migrating keratinocyte wound front is required for skin wound closure. Despite significant advances in wound healing research, we do not fully understand the molecular mechanisms that orchestrate collective keratinocyte migration. Here, we show that, in the wound front, the epidermal transcription factor Grainyhead like-3 (GRHL3) mediates decreased expression of the adherens junction protein E-cadherin; this results in relaxed adhesions between suprabasal keratinocytes, thus promoting collective cell migration and wound closure. Wound fronts from mice lacking GRHL3 in epithelial cells ( Grhl3 -cKO) have lower expression of Fascin-1 (FSCN1), a known negative regulator of E-cadherin. Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) on wounded keratinocytes shows decreased wound-induced chromatin accessibility near the Fscn1 gene in Grhl3 -cKO mice, a region enriched for GRHL3 motifs. These data reveal a wound-induced GRHL3/FSCN1/E-cadherin pathway that regulates keratinocyte-keratinocyte adhesion during wound-front migration; this pathway is activated in acute human wounds and is altered in diabetic wounds in mice, suggesting translational relevance.

No abstract

No abstract

The migration of wound-edge keratinocytes is part of the wound response, crucial for complete wound closure. Despite significant advances, the molecular mechanisms that orchestrate cell-cell adhesion between migrating keratinocytes are not fully characterized. During wound re-epithelization, keratinocytes at the wound edge undergo series of cellular modifications. These cellular modifications require a loosening of cell-cell adhesion for effective migration. Mice lacking the epidermal transcription factor Grainyhead Like-3 (GRHL3) exhibit impaired wound healing and an increased adhesion between keratinocytes at the wound edge. The increased cell-cell adhesion in Krt14-Cre Grhl3fl/fl wounds coincides with high expression of the adherens junction protein E-Cadherin and downregulation of the newly identified wound-response gene Fascin (FSCN1). Gene expression analysis of isolated woundedge keratinocytes shows significant downregulation of Fscn1 mRNA expression in Krt14-Cre Grhl3fl/fl wounds. In addition, ATAC-seq on Krt14-Cre Grhl3fl/fl wound-edge keratinocytes shows loss of wound-specific peaks near Fscn1 gene, in a region that is highly enriched for GRHL3 motifs. Together, these data elucidate a novel wound-specific FSCN1-E-cadherin pathway controlled by GRHL3 that is required for cell-cell loosening between migrating keratinocytes during wound re-epithelization. This pathway is altered in chronic diabetic wounds in mice.

No abstract