Transcription factor-mediated intestinal metaplasia and the role of a shadow enhancer

Epigenetic analysis on mouse gastric organoids overexpressing either CDX2, HNF4A or control GFP. Epigenetic analyses include ATAC-seq, Cut&Run. Mouse gastric and intestinal stem cell ATAC-seq and mouse gastric antrum and intestinal villus ChIP-seq. Overall design: We used lentiviral transduction to generate isogenic mouse gastric organoid lines expressing intestinal TFs CDX2 or HNF4A. For comparison we expressed a neutral product (GFP) in gastric organoids and CDX2 in squamous esophageal organoids. In these perturbed organoids we assessed chromatin accessibility using the assay for transposase-accessible chromatin (ATAC-seq), specific TF occupancy using CUT&RUN, and transcriptional changes using RNA-seq. We used CRISPR-Cas9 cutting to delete a novel Cdx2 enhancer in the mouse germline and dCas9-conjugated transcriptional repressor KRAB to further assess this cis-element in gastric organoids.