Abstract BACKGROUND &amp; AIMS Inflammatory bowel disease (IBD) is a complex and relapsing inflammatory disease, and patients with IBD exhibit a higher risk of developing colorectal cancer (CRC). Epithelial barrier disruption is one of the major causes of IBD in which epigenetic modulation is pivotal. However, the epigenetic mechanisms underlying the epithelial barrier integrity regulation remain largely unexplored. Here, we investigated how SETD2, an epigenetic modifier, maintains intestinal epithelial homeostasis and attenuates colonic inﬂammation and tumorigenesis. METHODS GEO public database and IBD tissues were used to investigate the clinical relevance of SETD2 in IBD. To define a role of SETD2 in the colitis, we generated mice with epithelium-specific deletion of Setd2 ( Setd2 Vil-KO mice). Acute colitis was induced by 2% dextran sodium sulfate (DSS), and colitis-associated CRC was induced by injecting azoxymethane (AOM), followed by three cycles of 2% DSS treatments. Colon tissues were collected from mice and analyzed by histology, immunohistochemistry and immunoblots. Organoids were generated from Setd2 Vil-KO and control mice, and were stained with 7-AAD to detect apoptosis. A fluorescent probe, 2′,7′-dichlorodihydroﬂuorescein diacetate (H2DCFDA), was used to detect the levels of ROS in intestinal epithelial cells (IECs) isolated from the two types of mice. RNA-seq and H3K36me3 ChIP-seq analyses were performed to identify the mis-regulated genes modulated by SETD2. Results were validated in functional rescue experiments by N-acetyl-l-cysteine (NAC) treatment and transgenes expression in IECs. RESULTS SETD2 expression became decreased in IBD patients and DSS-treated colitis mice. Setd2 Vil-KO mice displayed abnormal loss of mucus-producing goblet cells and antimicrobial peptide (AMP)-producing Paneth cells, and exhibited pre-mature intestinal inflammation development. Consistent with the reduced SETD2 expression in IBD patients, Setd2 Vil-KO mice showed increased susceptibility to DSS-induced colitis, accompanied by more severe epithelial barrier disruption and markedly increased intestinal permeability that subsequently facilitated inﬂammation-associated CRC. Mechanistically, deletion of Setd2 resulted in excess reactive oxygen species (ROS), which led to cellular apoptosis and defects in barrier integrity. NAC treatment in Setd2 Vil-KO mice rescued epithelial barrier injury and apoptosis. Importantly, Setd2 depletion led to excess ROS by directly down-regulating antioxidant genes that inhibit ROS reaction. Moreover, overexpression of antioxidant PRDX6 in Setd2 Vil-KO IECs largely alleviated the overproductions of ROS and improved the cellular survival. CONCLUSIONS Deficiency of Setd2 specifically in the intestine aggravates epithelial barrier disruption and inflammatory response in colitis via a mechanism dependent on oxidative stress. Thus, our results highlight an epigenetic mechanism by which Setd2 modulates oxidative stress to regulate intestinal epithelial homeostasis. SETD2 might therefore be a pivotal regulator that maintains the homeostasis of the intestinal mucosal barrier.

The Histone Methyltransferase SETD2 Modulates Oxidative Stress to Attenuate Colonic Inflammation and Tumorigenesis in Mice