滋养细胞凋亡中NSPcl沉默基因HOX转录调控模式的建立及表观遗传学调控机制

Trophoblast excessive apoptosis is the key link of Hypertension disorder complicating pregnancy (HDCP), but the exact mechanisms of which are still unclear. Our research group previously observed that DNA methylation is one of a significant pathogenesis in HDCP, while the ubiquitination of histone and another modifications of histone act in concert to participate in gene transcription which regulated by DNA methylation. Furthermore, the results of our pre-experiment also suggested that NSPc1, HOXA7 and HOXB13 are involved in the trophoblast apoptosis, but the transcriptional regulatory pattern and mechanism of H2A ubiquitination in NSPc1-mediated HOX silencing and the molecular are not clear. Therefore, the purpose in this study is to copy the model of HDCP, filter and determine the key subtypes of HOX in HDCP which caused by trophoblast apoptosis; To determine the HOXA7, B13 are key target genes regulated by NSPcl, elucidate the mechanisms of NSPcl which specifically binds to HOX to regulate its expression; To explore the mechanisms of EZH2 which promotes H2A ubiquitination through NSPcl/RING2 compounds; To reveal that EZH2 recruit DNMTs and HDACs is the key link of regulating gene transcription; To establish transcriptional regulatory pattern that NSPc1 regulate HOX through EZH2, DNMTs and HDACs; To clarify the molecular mechanisms that NSPcl siliencing HOX, look for the critical target, then provide theoretical basis for the prevention and treatment of HDCP.

滋养细胞凋亡致胎盘功能障碍是妊娠期高血压疾病(HDCP)的主要病理特征，但机制未清。课题组前期观察到DNA甲基化是滋养细胞凋亡的重要机制，但不解的是：为何同条件下不同基因DNA高低甲基化并存？组蛋白修饰是表观遗传学修饰的重要方式，与DNA甲基化能够建立有序的“串扰”，构成复杂的调控网络，已成为基因转录不可或缺的环节，同时预实验也观察到NSPc1沉默同源异形基因(HOX)参与了滋养细胞凋亡，故本课题拟在HDCP动物模型的基础上，分别过表达和沉默NSPc1并转染细胞，探讨其在滋养细胞凋亡中的作用，筛选并确定HOXA7，B13是受NSPcl调控的关键基因，并明确其相互作用的结构域；采用多种阻断(阻断剂、基因敲除等)为主的策略，揭示NSPc1调控表观遗传学因子EZH2/DNMT1/HDAC1介导HOX表达的机制，建立NSPc1沉默HOX转录调控模式，探寻关键靶点，为防治HDCP提供新策略。

背景：滋养细胞凋亡在子痫前期（PE）发病机制中起着重要作用，而DNA甲基化作为一种重要的表观遗传修饰方式，常常与基因沉默有关，课题组前期也观察到DNA甲基化是滋养细胞凋亡的重要机制，但其在PE中的具体调控机制仍不清楚。近年来，DNA甲基化与多梳组蛋白(PcG)介导的基因沉默之间错综复杂的相互作用已成为研究的热点。其中，NSPc1是Polycomb家族蛋白(PcG)的一个新基因，预实验也观察到NSPc1沉默同源异形基因(HOX)参与滋养细胞凋亡。.内容：本课题采用现代分子生物学技术，从整体和细胞水平观察NSPc1在滋养细胞凋亡中的具体作用，明确HOXA11是其调控的关键靶基因，探讨滋养细胞凋亡中NSPc1与DNA甲基化相互作用沉默HOX基因转录的分子机制，建立NSPc1沉默HOX转录调控模式，探寻关键靶点，为防治PE提供新策略。.结果：分别在PE胎盘组织和缺氧诱导滋养细胞中检测到凋亡水平升高，NSPc1表达增加，而将NSPc1干扰后滋养细胞凋亡水平降低，反之则升高，并且NSPc1可以作为转录失活剂，抑制HOXA11的表达；同时，甲基转移酶抑制滋养细胞后HOXA11启动子区甲基化水平降低且HOXA11表达增加。.该项目共发表SCI一篇，中文期刊论文7篇。.意义：该课题的实施有利于为PE发病机制的研究提供新的理论依据，寻找致病环节，确定关键靶点，为PE的防治提供新的思路。