N6‐Methyladenosine (m 6 A) Modifies Regenerative Transcripts in the Intestinal Epithelium

Background The intestinal epithelium forms a critical barrier between foreign antigens in the intestinal lumen and immune cells in the underlying mucosa. This barrier is compromised in inflammatory bowel disease (IBD), resulting in inflammation and epithelial cell death. Improved understanding of intestinal epithelial regeneration would identify novel therapeutic targets to enhance epithelial barrier repair in diseases such as IBD. After injury, tissues such as the blood, skeletal muscle, and central nervous system activate a regenerative program that is partially regulated by N6‐methyladeonsine (m 6 A) modification of RNA. However, few data exist regarding the role of m 6 A within the intestinal epithelium. The goal of this study was to profile m 6 A‐modification transcriptome‐wide in the regenerating intestinal epithelium. We hypothesize that after injury, m 6 A promotes regeneration within the intestinal epithelium by modifying transcripts important in epithelial repair . Methods Wildtype mice were given dextran sodium sulfate (DSS) in drinking water to induce intestinal inflammation and epithelial cell loss. Control mice were provided untreated water. DSS was followed by a two‐week washout period to allow for regenerating epithelium to appear adjacent to areas of epithelial ulceration. After DSS washout, regenerating intestinal epithelium was isolated by flow cytometry, RNA was extracted, and m 6 A‐modified transcripts were immunoprecipitated and sequenced (m 6 A‐seq). Results m 6 A‐seq yielded ~5500 total m 6 A‐modified sites within the regenerating intestinal epithelium. This corresponded to 314 m 6 A‐modified mRNAs that were unique to the regenerating as compared to the homeostatic epithelium. These unique transcripts were enriched for pathways involved in RNA transcription and processing, cell‐cell junctions, and Hippo pathway signaling, all of which have previously been implicated in intestinal epithelial regeneration. Conclusions . Our data support the hypothesis that m 6 A modifies many transcripts with known roles in intestinal epithelial regeneration. Ongoing studies are evaluating how m 6 A modification contributes to regulation of these transcripts and whether m 6 A identifies transcripts with previously unknown roles in regeneration. We are also evaluating whether m 6 A is required for optimal regeneration following epithelial injury.