组蛋白去甲基化酶KDM2B在天然免疫应答及炎症性疾病中的调控作用及表观机制研究

Various molecules are known to be involved in the tight regulation of innate immune responses triggered by pathogens and endogenous ligands to maintain the immunological balance. The levels of histone methylation on the promoters of many genes are alerted to regulate and control gene expression in immune response, thus the histone methyltransferase and demethylase are involved in innate and adaptive immunity. However, little attention was paid to the roles of histone demethylase in innate immunity and inflammatory diseases. In the preliminary experiments of this case, we found that the expression of histone H3K4 and H3K36 demethylase KDM2B was markedly increased in macrophages upon stimulation with TLR4 ligand LPS or infection with SeV virus. Furthermore, the production of IL-6 and IFN-b was increased in macrophages from KDM2B knock out mice treated with LPS、poly(I:C) or SeV, suggested that KDM2B is involved in the negative regulation of TLR and virus-triggered innate immune response. The primary aim of this study is to further investigate the epigenetic regulatory function of KDM2B in bacterial and virus-triggered innate immunity. Since the inflammation cytokine IL-6 is involved in the generation of collagen II-induced mouse arthritis (CIA model), thus we will also observe the roles of KDM2B in CIA, and explore the target genes and the underlying molecular mechanisms through epigenetic approaches, which will shed new light on the treatment of infectious diseases and inflammatory autoimmune diseases.

病原体或内源性配体通过模式识别受体触发的天然免疫应答受到很多分子的严密调控。组蛋白甲基修饰酶通过控制基因的组蛋白甲基化状态以改变基因的表达水平，参与免疫应答反应，然而组蛋白去甲基化酶在天然免疫应答与炎症性疾病中的功能研究较少。我们预实验研究发现小鼠巨噬细胞中组蛋白H3K4和H3K36去甲基化酶KDM2B的表达在TLR4配体LPS及RNA病毒SeV刺激后上调。KDM2B基因敲除小鼠巨噬细胞中LPS、poly(I:C)或SeV诱导的IL-6和IFN-b产生显著增加，提示KDM2B可能负向调控TLR及病毒触发的天然免疫应答。本研究拟在体内外探讨KDM2B对细菌和病毒诱导的天然免疫反应的表观调控功能，鉴于IL-6在胶原诱导的小鼠关节炎（CIA模型）中起重要作用，我们也将在CIA模型中观察KDM2B在炎症性自身免疫病中的作用，深入探讨表观学调控机制，为感染性疾病和炎症性自身免疫病的治疗提供新的思路。

本项目研究了组蛋白去甲基化酶KDM2B及其他免疫分子在天然免疫与炎症性疾病中的功能。研究发现KDM2B基因敲除小鼠的腹腔巨噬细胞中TLR配体LPS或Poly(I:C)诱导的IL-6产生较野生型小鼠巨噬细胞显著降低，但TLR配体或病毒诱导的TNF-a和IFN-b无明显差异。小鼠腹腔注射LPS或Poly(I:C)构建内毒素休克模型发现KDM2B缺陷小鼠血清中IL-6水平较对照小鼠显著降低，而TNF-a、IFN-b和IL-1b无明显差异，肺部炎症损伤显著减轻。DSS诱导的肠炎模型中，KDM2B缺陷小鼠体重下降较对照小鼠减少，血清中产生的IL-6水平较低，肠道炎症损伤程度更轻。KDM2B缺陷并不影响巨噬细胞中LPS诱导的MAPK、NF-kB和IRF3通路的活化。机制研究发现在TLR活化的巨噬细胞中，KDM2B可以选择性结合到IL-6基因启动子区，但并不能与TNF-a和IFN-b的基因启动子区结合，且不影响这三种细胞因子基因启动子区组蛋白H3K4me3和H3K36me2修饰的水平。但KDM2B在核内与染色质重塑复合物的核心催化亚基BRG1相互作用，通过BRG1对IL-6启动子区染色质重塑使其开放程度增加，进一步招募RNA Polymerase II，从而选择性促进了IL-6的转录表达。本部分研究发现了KDM2B在炎症反应中发挥非甲基化酶活性依赖的新的调控功能，发现了新的选择性调控IL-6转录表达的染色质修饰机理。另一部分研究发现了鞘糖脂iGb3能通过活化细胞内CD1d逆向信号增强巨噬细胞中TLR诱导的天然免疫炎症反应。第三部分研究发现Dot1l能够选择性地结合到IL-6和IFN-b启动子区，催化H3K79me2/3修饰，增强其转录表达。本项目已发表标注该基金资助的SCI论文4篇，分别为Cell Mol Immunol（2019 Jun 13. doi: 10.1038/s41423-019-0251-z，共同通讯作者）、Cell Research（2019;29:42-53，共同通讯作者）、J Exp Med（2019;216:772-785，共同第一作者）、Cell Mol Immunol（2018 Oct 1. doi: 10.1038/s41423-018-0170-4，共同第一作者）。项目负责人作为主要参与人员获得学术奖励2项。本项目已经顺利完成研究目标。