RNA甲基转移酶NSUN5对先兆子痫发病的影响及机制研究

Preeclampsia (PE) is a human-specific hypertensive disorder of pregnancy，it seriously threats maternal life, but the etiology of PE is still not clear. Numerous data show that the occurrence of PE is related to genetic imprinting, of which about 35% of the PE risk is from the mother's genetic factors. We detected a missense mutation in the NSUN5 gene in patients with severe PE using MassARRAY SNP technique on a clinical blood samples (982 controls / 868 severe PE), and found the amino acid sequence of NSUN5 was altered， we also found that NSUN5 is highly expressed in human decidua and placentas. NSUN5 encodes RNA methyltransferase, whose function is to catalyze the methylation of RNA cytosine (m5C). RNA methylation is a very important epigenetic modification，and m5C modification exists in mRNA, tRNA, rRNA and long non-coding RNA, it has important biological functions. Williams-Beuren syndrome occurs in the absence of the region of NSUN5 gene. Deletion of this gene in yeast increases its stress resistance and longevity. Therefore, the present application intends to explore the impact of NSUN5 mutations on PE and its pathogenesis by m5C RNA methylation sequencing and transcriptome sequencing, RIP, RNA Pulldown, luciferase reporter system, tissue-specific NSUN5 point mutation mice and other technologies and methods.

先兆子痫（preeclampsia，PE），是人类特有的妊娠期高血压疾病，严重威胁孕产妇生命但病因尚不清楚。研究表明，PE的发生和遗传有关，约35%的易感风险来自母亲的遗传因素。我们利用MassARRAY SNP分析了982例对照/868 例重度PE的血液样本,发现重度PE患者NSUN5基因发生了错义突变，并发现NSUN5在人蜕膜组织及胎盘中高表达。NSUN5编码RNA甲基转移酶，催化RNA胞嘧啶发生甲基化(m5C)。RNA甲基化包括m5C是非常重要的表观遗传修饰，存在于mRNA、tRNA、rRNA及长非编码RNA。NSUN5缺失患者发生Williams-Beuren 综合征。酵母中该基因缺失增加其抗逆性和寿命。因此本申请拟通过m5C RNA甲基化和全转录组测序、RIP、 RNA Pulldown、荧光素酶报告系统、组织特异性NSUN5点突变小鼠等探索NSUN5突变对PE的影响及致病机制。

先兆子痫（PE）是一种妊娠期特异性高血压疾病，严重损害孕妇和胎儿健康。然而，其发病机制仍然未明。蜕膜化缺陷被认为与子痫前期有关。越来越多的证据表明单核苷酸多态性（SNPs）与多种疾病相关。NOP2/Sun 5（NSUN5）是一种RNA甲基转移酶。我们通过人类外显子微珠芯片（Human Exome BeadChip assays）分析370例重度PE/482例非PE病例，并利用MassARRAY DNA质谱基因分析（MassARRAY DNA Mass Spectrometry Gene Analysis）498例重度PE/500例非PE患者发现外周血中NSUN5的SNP rs77133388与PE显著相关。然后，我们因此在NSUN5外显子7的rs77133388处制备了C>T单碱基突变小鼠（NSUN5 R295C）。发现怀孕的NSUN5突变小鼠表现出PE症状，如血压升高和蛋白尿,胎儿及胎盘重量减小等。NSUN5 R295C小鼠中，胎盘的蜕膜区显著减少胎盘的蜕膜区显著变小。在人工诱导的蜕膜化后，NSUN5 R295C小鼠蜕膜化受损，多倍体细胞数量显著减少。人工诱导的蜕膜组织的RNA-seq显示，NSUN5 R295C小鼠的Il-11rα水平显著低于野生型对照小鼠。此外，我们发现磷酸化的JAK2、STAT3水平以及cylinD3和P21表达降低。此外，RNA免疫沉淀实验显示NSUN5与IL-11Rα结合。 总而言之，我们的数据表明，NSUN5突变通过IL-11Rα/JAK2/STAT3/CylinD3途径改变母体蜕膜化，从而损害胎盘发育并导致PE的发生。