{"title":"WTAP 通过 m6A 修饰抑制 CES2 的表达，从而促进溃疡性结肠炎的进展","authors":"Xiaoran Xie , Sha Cheng , Xiong Chen , Xia Wang","doi":"10.1016/j.jaut.2024.103295","DOIUrl":null,"url":null,"abstract":"<div><h3>Objective</h3><p>This study will explore the function of WTAP, the critical segment of m<sup>6</sup>A methyltransferase complex, in UC and its regulation on immune response.</p></div><div><h3>Methods</h3><p>The expression levels of key proteins were detected in colon tissues which were derived from UC patients and mice. Macrophage polarization and CD4<sup>+</sup> T cell infiltration were detected by flow cytometry and IF staining. ELISA assay was utilized to analyze the level of the inflammatory cytokines. m<sup>6</sup>A-RIP-PCR, actinomycin D test, and RIP assays were utilized to detect the m<sup>6</sup>A level, stability, and bound proteins of CES2 mRNA. A dual luciferase reporter assay was conducted to confirm the transcriptional interactions between genes. A co-culture system of intestinal epithelium-like organs was constructed to detect the primary mouse intestinal epithelial cells (PMIEC) differentiation. The interaction between proteins was detected via Co-IP assay.</p></div><div><h3>Results</h3><p>The expression of WTAP and CES2 in UC tissues was increased and decreased, respectively. Knockdown of WTAP inhibited the progression of UC in mice by inhibiting M1 macrophage polarization and CD4<sup>+</sup> T cell infiltration. WTAP combined YTHDF2 to promote the m<sup>6</sup>A modification of CES2 mRNA and inhibited its expression. CES2 co-expressed with EPHX2 and overexpression of CES2 promoted the differentiation of PMIEC. The inhibitory effect of WTAP knockdown on the progress of UC was partially abrogated by CES2 knockdown.</p></div><div><h3>Conclusion</h3><p>WTAP/YTHDF2 silences CES2 by promoting its m<sup>6</sup>A modification and then promotes the progression of UC. WTAP could be a promoting therapy target of UC.</p></div>","PeriodicalId":15245,"journal":{"name":"Journal of autoimmunity","volume":"148 ","pages":"Article 103295"},"PeriodicalIF":7.9000,"publicationDate":"2024-08-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"WTAP promotes the progression of ulcerative colitis by silencing the expression of CES2 through m6A modification\",\"authors\":\"Xiaoran Xie , Sha Cheng , Xiong Chen , Xia Wang\",\"doi\":\"10.1016/j.jaut.2024.103295\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><h3>Objective</h3><p>This study will explore the function of WTAP, the critical segment of m<sup>6</sup>A methyltransferase complex, in UC and its regulation on immune response.</p></div><div><h3>Methods</h3><p>The expression levels of key proteins were detected in colon tissues which were derived from UC patients and mice. Macrophage polarization and CD4<sup>+</sup> T cell infiltration were detected by flow cytometry and IF staining. ELISA assay was utilized to analyze the level of the inflammatory cytokines. m<sup>6</sup>A-RIP-PCR, actinomycin D test, and RIP assays were utilized to detect the m<sup>6</sup>A level, stability, and bound proteins of CES2 mRNA. A dual luciferase reporter assay was conducted to confirm the transcriptional interactions between genes. A co-culture system of intestinal epithelium-like organs was constructed to detect the primary mouse intestinal epithelial cells (PMIEC) differentiation. The interaction between proteins was detected via Co-IP assay.</p></div><div><h3>Results</h3><p>The expression of WTAP and CES2 in UC tissues was increased and decreased, respectively. Knockdown of WTAP inhibited the progression of UC in mice by inhibiting M1 macrophage polarization and CD4<sup>+</sup> T cell infiltration. WTAP combined YTHDF2 to promote the m<sup>6</sup>A modification of CES2 mRNA and inhibited its expression. CES2 co-expressed with EPHX2 and overexpression of CES2 promoted the differentiation of PMIEC. The inhibitory effect of WTAP knockdown on the progress of UC was partially abrogated by CES2 knockdown.</p></div><div><h3>Conclusion</h3><p>WTAP/YTHDF2 silences CES2 by promoting its m<sup>6</sup>A modification and then promotes the progression of UC. WTAP could be a promoting therapy target of UC.</p></div>\",\"PeriodicalId\":15245,\"journal\":{\"name\":\"Journal of autoimmunity\",\"volume\":\"148 \",\"pages\":\"Article 103295\"},\"PeriodicalIF\":7.9000,\"publicationDate\":\"2024-08-13\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of autoimmunity\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S089684112400129X\",\"RegionNum\":1,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"IMMUNOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of autoimmunity","FirstCategoryId":"3","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S089684112400129X","RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"IMMUNOLOGY","Score":null,"Total":0}
引用次数: 0
摘要
方法 在 UC 患者和小鼠的结肠组织中检测关键蛋白的表达水平。采用流式细胞术和 IF 染色法检测巨噬细胞极化和 CD4+ T 细胞浸润。m6A-RIP-PCR、放线菌素 D 试验和 RIP 试验用于检测 CES2 mRNA 的 m6A 水平、稳定性和结合蛋白。为了证实基因之间的转录相互作用，还进行了双荧光素酶报告实验。构建了肠上皮样器官共培养系统，以检测小鼠原代肠上皮细胞（PMIEC）的分化情况。结果WTAP和CES2在UC组织中的表达分别增加和减少。敲除 WTAP 可抑制 M1 巨噬细胞极化和 CD4+ T 细胞浸润，从而抑制小鼠 UC 的恶化。WTAP与YTHDF2结合可促进CES2 mRNA的m6A修饰并抑制其表达。CES2与EPHX2共表达，过表达CES2可促进PMIEC的分化。结论 WTAP/YTHDF2通过促进CES2的m6A修饰沉默CES2，进而促进UC的进展。WTAP可能是促进UC治疗的靶点。
WTAP promotes the progression of ulcerative colitis by silencing the expression of CES2 through m6A modification
Objective
This study will explore the function of WTAP, the critical segment of m6A methyltransferase complex, in UC and its regulation on immune response.
Methods
The expression levels of key proteins were detected in colon tissues which were derived from UC patients and mice. Macrophage polarization and CD4+ T cell infiltration were detected by flow cytometry and IF staining. ELISA assay was utilized to analyze the level of the inflammatory cytokines. m6A-RIP-PCR, actinomycin D test, and RIP assays were utilized to detect the m6A level, stability, and bound proteins of CES2 mRNA. A dual luciferase reporter assay was conducted to confirm the transcriptional interactions between genes. A co-culture system of intestinal epithelium-like organs was constructed to detect the primary mouse intestinal epithelial cells (PMIEC) differentiation. The interaction between proteins was detected via Co-IP assay.
Results
The expression of WTAP and CES2 in UC tissues was increased and decreased, respectively. Knockdown of WTAP inhibited the progression of UC in mice by inhibiting M1 macrophage polarization and CD4+ T cell infiltration. WTAP combined YTHDF2 to promote the m6A modification of CES2 mRNA and inhibited its expression. CES2 co-expressed with EPHX2 and overexpression of CES2 promoted the differentiation of PMIEC. The inhibitory effect of WTAP knockdown on the progress of UC was partially abrogated by CES2 knockdown.
Conclusion
WTAP/YTHDF2 silences CES2 by promoting its m6A modification and then promotes the progression of UC. WTAP could be a promoting therapy target of UC.
期刊介绍：
The Journal of Autoimmunity serves as the primary publication for research on various facets of autoimmunity. These include topics such as the mechanism of self-recognition, regulation of autoimmune responses, experimental autoimmune diseases, diagnostic tests for autoantibodies, as well as the epidemiology, pathophysiology, and treatment of autoimmune diseases. While the journal covers a wide range of subjects, it emphasizes papers exploring the genetic, molecular biology, and cellular aspects of the field.
The Journal of Translational Autoimmunity, on the other hand, is a subsidiary journal of the Journal of Autoimmunity. It focuses specifically on translating scientific discoveries in autoimmunity into clinical applications and practical solutions. By highlighting research that bridges the gap between basic science and clinical practice, the Journal of Translational Autoimmunity aims to advance the understanding and treatment of autoimmune diseases.
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