叶酸缺乏对核糖体rRNA基因转录介导细胞命运决定因子路径在神经管畸形中的分子机制研究

The mechanism of folic acid metabolic disorder causing neural tube defects (NTDs) is not clear. “rRNA gene transcription - cell fate determination factors - developmental signals” integration pathway affects cell differentiation in organs forming. It is not known whether cell fate determination factors affect the differentiation of cells by folate metabolic environment in early embryo development. In samples from NTDs with low folate level, aberrant UBF level was observed along with up-regulation of rRNA gene transcription in our previous studies. To understand the effects of these synergistic molecules on the key neural tube closure pathways and the differentiation of neuroectoderm, we will use the cell model and animal model in folate deficiency to investigate the mechanism of rRNA gene transcriptional regulation and the differentiation of neural stem cells with the technique of CHIP-seq, RNA-seq and Co-IP.The effects of folic acid supplementation on the above-mentioned integration pathway and the intervention mechanism of cell differentiation will be revealed by DNase-seq. Furthermore, the validation of the function of rRNA gene transcription mediated by UBF interaction with cell fate determination factors will be made in human NTDs samples combined with Nanostring technology. This project attempts to analyse NTDs etiology from the rRNA transcription perspective to improve reproductive health.

叶酸代谢紊乱致神经管畸形(NTDs)发生机制不清楚。rRNA转录-细胞命运决定因子-发育信号通路这一整合路径影响器官形成细胞分化走向。胚胎早期细胞命运决定因子在叶酸代谢变化下是否影响细胞分化走向而影响神经管闭合并不清楚。我们已证实低叶酸NTDs标本上游结合因子（UBF）表达增加同时rRNA基因转录改变，现拟通过叶酸缺乏细胞及动物模型明确叶酸对UBF协同细胞命运决定因子介导rRNA基因转录调控机制及神经干细胞分化去向；采用CHIP-seq、RNA-seq、Co-IP探讨以上分子协同作用对神经外胚层分化及神经管闭合关键通路活性影响；采用DNase-seq探讨叶酸增补对上述整合路径的影响及细胞分化走向的干预机制；结合Nanostring验证NTDs标本中UBF协同细胞命运决定因子调控核糖体基因转录。本项目拟从叶酸影响rRNA转录-细胞命运决定因子-神经管闭合通路路径探讨NTDs病因，促进生殖健康

神经管畸形（Neural Tube Defects, NTDs）是一类常见出生缺陷疾病，中国是NTDs的高发国家，叶酸增补可预防NTDs但其机制不明。核糖体rRNA—细胞命运决定因子—发育信号通路这一整合路径影响器官形成细胞分化走向。胚胎早期细胞命运决定在叶酸代谢障碍下是否会影响细胞分化走向进而影响神经管闭合并不清楚。.本项目通过叶酸缺乏的细胞、小鼠、临床NTDs标本三个层面研究叶酸代谢障碍导致同型半胱氨酸增多，进而导致组蛋白H3K79Hcy修饰富集，调控下游细胞命运决定相关基因Smarc4、Cecr2、Dnmnt3b转录表达下降，参与NTDs的发生。本研究首次揭示了组蛋白H3K79Hcy参与细胞命运决定在胚胎神经发育中的分子机制，为叶酸代谢紊乱导致的同型半胱氨酸修饰参与NTDs及其他疾病的发生提供了重要理论基础, 研究结果发表于Nature Communcation。.基于叶酸代谢紊乱的研究思路，本项目还发现叶酸缺乏下细胞命运决定因子Gcm1异常高表达后激活经典Wnt/β-catenin信号通路导致NTDs的发生，这一发现揭示细胞命运决定因子—发育信号通路在叶酸缺乏导致NTDs中的作用机制，结果发表于Cell Death & Disease。.最后本项目在前期甲氨蝶呤处理细胞导致核糖体rRNA基因转录改变的基础上，从基因组不稳定性及表观染色质改变探讨叶酸代谢障碍导致的NTDs易感基因表观特征的改变，并据此筛出7个新的NTDs易感基因，这为叶酸障碍导致的NTDs发生机制提供新的研究方向, 相关结果目前投稿于Ebio Medicine。.综上，本项目从核糖体rRNA—细胞命运决定因子—发育信号通路组成的分子路径探讨叶酸缺乏导致NTDs的分子机制，推动叶酸等营养素代谢变化对表观遗传修饰和基因调控影响规律的科学认知，也为降低出生缺陷，保障生殖健康提供新的理论依据。.