Mechanism Of Epigenetic Regulatory Factors SETD2 And BRG1 In Inflammatory Bowel Disease

Background:Inflammatory bowel disease（IBD）is a chronic and recurrent intestinal inflammatory disease that can be sub-classified into Crohn’s disease（CD）and ulcerative colitis（UC）.In recent years,the incidence of IBD has been increasing quickly,and the continuous intestinal inflammatory response will have a higher risk of developing colorectal cancer（CRC）.The pathogenesis of IBD is multifaceted,involving genetic factors,environmental factors,autoimmune system,and intestinal microbiota.However,the specific pathogenesis of IBD is still unclear.Studies have shown that IBD is not only related to genetic factors,but also related to epigenetic regulation such as histone modification and chromatin remodeling.Epigenetic regulation disorder is important in the onset and pathogenesis of IBD.BRG1 and SETD2 are two common epigenetic factors,both of which is frequently mutated in UC samples with a high risk of developing CRC,indicating that SETD2 and BRG1 may play an important role in the occurrence of IBD.However,the specific mechanism of SETD2 and BRG1 in IBD remains largely undefined.Objective:This study aims to explore the mechanisms of SETD2 and BRG1 in the occurrence of IBD by the analysis of clinical patient samples and mouse models of IBD.Methods:To define the role of BRG1 and SETD2 in IBD,firstly,we used IBD biopsy specimens to analyze the expression of SETD2 and BRG1 by quantitative RT-PCR（RT-q PCR）assays and immunohistochemistry analyses,and combined the GEO datasets of IBD samples to determine the clinical correlation between SETD2/BRG1 and IBD.Then,we respectively established mouse models of epithelium-specific Setd2 or Brg1,and combined with dextran sodium sulfate（DSS）to induce colitis in mice.The mouse phenotype was analyzed by immunohistochemical staining and western blotting.In addition,we also used azoxymethane（AOM）and DSS to induce inflammation-associated CRC in mice,and performed histological staining and morphological analysis of colon tumors.Finally,the RNA-Seq and Ch IP-Seq bioinformatics analysis methods were used to find the target genes regulated by SETD2 and BRG1 respectively.In addition,the molecular mechanisms of SETD2 and BRG1 regulating the occurrence of IBD were determined by combining with the multi-level rescue experiments of molecular cells and animals.Results:In the first part of our study,SETD2 expression became decreased in IBD patients.Setd2^Vil-KO mice showed a more severe inflammation phenotype during DSS induction.Setd2 deficiency caused mouse intestinal barrier damage and increased intestinal epithelial cell apoptosis,which in turn aggravated the intestinal inflammatory response and subsequent tumorigenesis.Mechanistically,we found that Setd2 depletion resulted in excess reactive oxygen species（ROS）by directly down-regulating antioxidant genes（Prdx3,Prdx6,Gclm,and Srxn1）,leading to defects in barrier integrity and subsequently inflammatory damage.Moreover,overexpression of the antioxidant gene Prdx6 in Setd2^Vil-KO intestinal epithelial cells largely alleviated the overproductions of ROS and improved the cellular survival.In the second part of our study,BRG1 expression became decreased in IBD patients.Brg1^IEC-AKO mice could spontaneously develop colitis,and exhibited a more severe inflammation phenotype during DSS induction.Brg1 deficiency caused mouse intestinal barrier damage and increased intestinal epithelial cell apoptosis,which in turn aggravated the intestinal inflammatory response and subsequent tumorigenesis.Moreover,Brg1overexpression protected the mice from DSS-induced colitis and subsequent tumorigenesis.Mechanistically,we found that BRG1 directly governed the transcription of Atg16l1,Ambra1,Atg7 and Wipi2 to regulate the autophagy response.Defective autophagy caused by Brg1 deficiency results in excess ROS,which promoted epithelial cell apoptosis,and led to the defects in barrier integrity and subsequently inflammatory damage.Conclusions:Together,our findings revealed:（1）SETD2 maintains intestinal epithelial homeostasis and inhibits the occurrence of colitis by regulating the oxidative stress response.（2）BRG1,as an autophagy regulator,reduces the level of ROS by regulating the autophagy pathway,thereby inhibiting the occurrence of intestinal inflammation.In summary,our results establish that SETD2 and BRG1 are likely potential therapeutic targets for IBD intervention.Our study can provide an important theoretical basis for the drug development of IBD and inflammation-associated CRC related to SETD2 and BRG1 mutations.