Villification of the intestinal epithelium is driven by Foxl1

The primitive gut tube of mammals initially forms as a simple cylinder consisting of the endoderm-derived, pseudostratified epithelium and the mesoderm-derived surrounding mesenchyme. During mid-gestation a dramatic transformation occurs in which the epithelium is both restructured into its final cuboidal form and simultaneously folded and refolded to create intestinal villi and intervillus regions, the incipient crypts. Here we show that the mesenchymal winged helix transcription factor Foxl1, itself induced by epithelial hedgehog signaling, controls villification by activating BMP and PDGFRa as well as planar cell polarity genes in epithelial-adjacent telocyte progenitors, both directly and in a feed- forward loop with Foxo3. In the absence of Foxl1-dependent mesenchymal signaling, villus formation is delayed, the separation of epithelial cells into mitotic intervillus and postmitotic villus cells impaired, and the differentiation of secretory progenitors blocked. Thus, Foxl1 orchestrates key events during the epithelial transition of the fetal mammalian gut. Overall design: Single-cell RNA sequencing (scRNA-seq) was performed to investigate the role of the winged helix transcription factor Foxl1 in orchestrating villus formation during mouse fetal gut development. Proximal half of the small intestines were isolated from embryonic day 15.5 (E15.5) Foxl1-CreERT2-tdTomato mice, and single-cell suspensions were prepared using enzymatic digestion followed by FACS enrichment of live cells. Libraries were generated using the 10x Genomics Chromium Single Cell 3' v2 platform, with a target of ~5,000 cells per sample. The goal was to characterize Foxl1-dependent gene regulatory networks in mesenchymal telocyte progenitors and their impact on epithelial-mesenchymal cross talk and developmental signaling pathways. Comparative analyses were performed between control (heterozygous) and Foxl1-null (homozygous) embryos to define how loss of Foxl1 alters transcriptional programs essential for intestinal villification.