TET-mediated DNA oxidations in mucosal innate defense

项目总结 肠道微生物区系可改变肠上皮细胞基因组DNA甲基化模式。多么 在动态平衡和应激状态下，不同类型的IEC的功能受到这种表观遗传变化的影响。 目前情况仍不明朗。DNA甲基化是一种抑制性表观遗传标记，可以被 TET1、TET2和TET3酶的10-11易位家族。TETS是DNA双加氧酶， 将甲基化DNA-5-甲基胞嘧啶(5mC)-氧化成5-羟基-甲基胞嘧啶(5HmC)，5-甲基胞嘧啶(5mC)- 甲酰胞嘧啶(5fC)和5-羧基胞嘧啶(5caC)。5fC和5caC都可以通过基于DNA的切除来切除 导致未修饰胞嘧啶的修复因素。Tet酶最近被认为是一种新的危险因素 炎症性肠病(IBD)患者，但Tet介导的DNA氧化在体内平衡和体内的作用 对环境应激源的反应是未知的。初步数据显示，人类IECS显示出一种海拔 在侵袭性病原体感染时5hmC DNA氧化，而缺乏Tet3的小鼠IECs 回肠内皮细胞5hmC丰度。ScRNA-Seq表明小鼠Tet3是体内含量最丰富的Tet酶 IECs，尤其是Paneth细胞。Tet3DIEC小鼠成熟的Paneth细胞减少，对 由肠道病原体或破坏屏障的化学物质引起的炎症。该项目检验了一个中心假说 TET3介导的DNA氧化在诱导IEC分化和促进抗菌作用中的双重作用 通过5hmC诱导的容许性染色质反应，同时通过5fC和5caC诱导的抑制分化 转录暂停。Aim 1将使用全基因组方法来鉴定肠道干细胞(ISC)和Paneth 细胞特异性TET3和DNA氧化基因调控网络在动态平衡和对不同 细胞应激源，如病原体和化学物质。目标2将描述ISC和Paneth细胞的特异性 TET3介导的DNA氧化调节粘膜炎症反应。这个MPI项目，利用 表观遗传学和肠道生物学方面的互补专业知识，以解决DNA氧化如何调节 表观基因组对应激源的反应，以帮助化解炎症。Tet介导的DNA氧化的想法 可能是一种不可或缺的粘膜天然免疫成分，以应对感染期间的氧化应激 炎症是新奇的。阐明特定IEC类型中的Tet功能可能对人类产生重大影响 胃肠粘膜免疫学与疾病。如果Tet酶确实被这项研究发现为 炎症反应的关键介质和调节因子，正如文献和我们的初步数据所预测的那样， 结果可能与转化医学高度相关。

PROJECT SUMMARY DNA methylation pattern in the genome of intestinal epithelial cells (IECs) can be altered by gut microbiota. How the functions of various IEC types are affected by such epigenetic changes under homeostatic and stress conditions remain unclear. DNA methylation is a repressive epigenetic mark that can be actively reversed by the Ten-Eleven Translocation family of enzymes TET1, TET2 and TET3. TETs are DNA dioxygenases that successively oxidize methylated DNA - 5-methylcytosine (5mC) - into 5-hydroxy-methylcytosine (5hmC), 5- formylcytosine (5fC), and 5-carboxylcytosine (5caC). Both 5fC and 5caC can be excised by DNA based excision repair factors leading to unmodified cytosines. TET enzymes were recently implicated as new risk factors in inflammatory bowel disease (IBD) patients, but the role of TET-mediated DNA oxidation in homeostasis and in response to environmental stressors are unknown. Preliminary data show that human IECs display an elevation of 5hmC DNA oxidation upon infection by invasive pathogen, and mouse IECs lacking Tet3 had reduction of 5hmC abundance in ileal IECs. scRNA-Seq suggests that mouse Tet3 is the most abundant TET enzyme in IECs, especially in Paneth cells. Tet3DIEC mice had reduced mature Paneth cells, increased susceptibilities to inflammation caused by enteric pathogen or barrier-disrupting chemical. The project tests a central hypothesis that TET3-mediated DNA oxidations play dual roles in guiding IEC differentiation and promoting anti-microbial response via 5hmC-induced permissive chromatin while restraining differentiation via 5fC- and 5caC-induced transcriptional pausing. Aim 1 will use genome wide approaches to identify intestinal stem cell (ISC) and Paneth cell specific TET3 and DNA oxidation gene regulatory networks under homeostasis and in responding to distinct cellular stressors such as pathogen and chemical. Aim 2 will characterize how ISC and Paneth cell specific TET3-mediated DNA oxidation regulate mucosal inflammatory response. This MPI project, utilizing complementary expertise in epigenetics and intestinal biology to address how DNA oxidations regulate the epigenome in response to stressors to help resolve inflammation. The idea that TET-mediated DNA oxidations may be an integral mucosal innate immune component to cope with oxidative stresses during infection and inflammation is novel. Elucidating TET functions in specific IEC types may exert major impact on human gastrointestinal mucosal immunology and diseases. If TET enzymes are indeed uncovered by this research as key mediator and regulator of inflammatory responses, as predicted by literature and our preliminary data, the outcome can be of high relevance to translational medicine.