menin调控肝炎恶性转化的信号网络与组蛋白修饰机制

Menin, a main transcription factor, regulates various cell phenotypes through epigenetic modification, such as histone methylation. Recently, we found an interesting interplay bettwen menin and IL-6 pathway, which co-stimulates the malignant transformation of chronic hepatitis. However, the precise mechanisms remain unclear, and the biological significance should be further confirmed. The present study will focus on the identification of crucial molecules and signal pathway in the malignant transformation of chronic liver disease. Firstly, we will use the menin and/or TLR4 knockout mice to generates primary liver cancer model, to elucidate the crucial biological effect of menin and other Histone methyltransferases in promoting malignant transformation of chronic hepatitis, and systematic evaluate the pathogenic characteristic of the loss of menin in liver cancer development. Secondly, we based on data integration analysis of previous obtained microarray and ChIP-on-ChIP results, to screen and identify the menin regulated essential target gene and inflammation pathway network, and verify the vital role in promoting liver carcinogenesis. We use Chromatin Immunoprecipitation and other key approach to in-depth discuss how menin regulates target gene transcription. Thirdly, we will uncover the interplay mechanism between menin and TLR4/IL-6 inflammation pathway, and coordinated effect on liver carcinogenesis. We will clarify Histone 3 lysine 4 methylation mechanism of target gene transcriptional regulation by menin/MLL Histone Methyltransferase complex, exactly identify histone Methyltransferase complex component, and further confirm the key role that menin mediated histone modification enzyme system in the process of HCC developing. Together, these studies will reveal a novel epigenetic mechanism for the pro-oncogene menin in the promoting malignant transformation of chronic hepatitis.

menin是通过组蛋白甲基化等表观遗传学机制广泛参与各种细胞表型调控的关键转录因子。我们发现，menin与IL-6等前炎症分子通路形成有趣的正反馈调控网络，共同促进肝炎的恶性转化。本研究围绕“慢性肝炎恶性转化的关键分子鉴定及其调控网络”这一科学问题，利用menin、TLR4等基因敲除的模式动物，进一步深入证实menin等组蛋白甲基化酶促进肝炎恶性转化的关键生物学作用，系统评价肝癌发生发展的病理学动态特点；从已获得的肝脏全基因组组蛋白甲基化谱及表达谱芯片数据的整合性分析入手，筛选menin调控的关键信号网络，探讨menin调控肝细胞恶性表型和炎症因子转录的H3K4等组蛋白甲基化规律，分析组蛋白甲基化酶组成；鉴定menin及TLR4/IL-6等促炎信号通路间的相互调控机制，以及肝癌发生的多基因协同效应。这将有助于深入揭示关键分子及其调控的信号网络共同控制肝炎非可控性恶性转化的表观遗传学机制。

MLL/menin复合物为代表的具有H3K4甲基转移酶活性，与急性淋巴细胞白血病、前列腺癌、小儿神经胶质瘤等发病密切相关。然而，非可控性肝脏炎症的恶性转化进程中TrxG及PcG的生物学功能及其介导的H3K4me3等组蛋白共价修饰规律目前尚不清楚。本研究通过建立肝细胞损伤性肝脏局部炎症、原发性肝癌等小鼠模型，发现在肝脏炎症至肝癌发病过程中menin及其组蛋白甲基化复合物明显被激活，且与炎症通路的激活呈正相关。Men1基因杂合敲除抑制肝脏局部的炎症反应及原发性肝癌发生：如CCl4、DEN刺激小鼠肝脏及循环血中的IL-6、TNF-α等炎症因子急剧升高；Men1杂合敲除显著降低了DEN诱导的小鼠原发性肝癌发生率。提示，menin复合物是促进肝脏炎症反应及其恶性转化的关键开关之一。进一步利用ChIP-on-Chip、ChIP等技术鉴定了肝癌中H3K4me3、H3K79me2分布规律以及组蛋白甲基转移酶menin/MLL复合物在基因组染色质中的结合规律，并筛选了menin/MLL通过组蛋白甲基化机制调控的靶基因网络。证实了menin结合在Yap1等肝癌关键原癌基因启动子位点，通过H3K4甲基化修饰而激活其转录，Yap1进一步促进IL-6等炎症因子表达，证实了menin-Yap1-IL6网络是肝炎非可控性恶性转化的关键网络之一。深入分析了肝癌染色质的组蛋白甲基化修饰规律，发现menin及EZH2共定位在大量靶基因启动子位点，而H3K4及H3K27组蛋白甲基化修饰则分别发生在独立的靶基因网络，被激活的H3K4则主要修饰Yap1等促癌基因，而H3K27修饰则主要发生在CDKN2A等典型抑癌基因的启动子位点。以上研究证实了正负性组蛋白甲基化修饰共同促进肝癌发生的协同机制。本研究探讨了肝炎非可控性恶性转化过程中TrxG及PcG家族的生物学功能及其调控的正负性组蛋白甲基化重建规律及其靶基因调控网络，鉴定了肝细胞肝癌中关键表观遗传调控因子的遗传学突变规律，为肝炎非可控性恶性转化的早期预测及预后评估提供了遗传学及表观遗传学理论基础。