EMX2基因突变在苗勒管发育异常发病机制中的作用研究

A number of genes have been identified to participate in the organogenesis of the female reproductive tract and consequently to regulate its proper development. Emx2 is a transcription factor necessary for reproductive tract development in mouse. Emx2 mutant female mice lacked oviducts, uterus and the upper part of the vagina. Human EMX2 gene located in chromosome 10q26.1 and shared a homology of 94.18% with mice. So mutations in EMX2 may also cause MA in human..In our previous study, EMX2 gene was sequenced in 192 patients with MA consulting from Hospital for Reproductive Medicine Affiliated to Shandong University. Among the three variants, c.424G>T was a nonsense variant and present only in one patient with MA. The mutation resulted interruption of translation and deletion of HOX domain in the truncated EMX2 protein..Limited by ethics, further studies are difficult to perform in vivo. By cloning wild and mutated EMX2 genes to eukaryotic expression vector, we planned to observe the effect of this variant to synthesis of EMX2 protein. Teneurin 1 is the first identified target gene of EMX2. We further will evaluate the impact of EMX2 mutation to the expression of Teneurin 1 mRNA and protein. It will help to demonstrate the role of EMX2 in the pathology of MA and the possible pathway.

既往研究提示众多基因参与了苗勒管发育异常(MA)的发生。小鼠emx2是生殖系统发育所必需的转录因子，emx2敲除雌性小鼠的输卵管、子宫和阴道上半部分缺如。人类EMX2与小鼠emx2的同源性高达94.18%，故EMX2基因突变也可能导致人类MA。本课题组之前的研究中，对192例MA患者的EMX2基因进行了测序，发现了3个突变位点，其中新发突变c.424G>T只出现于MA患者中。该突变造成蛋白翻译中断，截短的EMX2蛋白丢失了重要的功能域HOX。受伦理学等限制，EMX2的功能实验在体内难以进行。本课题组拟构建EMX2野生型和突变型重组体，使其在真核细胞中表达，来明确该无义突变对EMX2蛋白合成的影响。Teneurin 1是第一个被证实的EMX2的靶基因，观察EMX2突变对Teneurin 1mRNA及蛋白的表达变化的影响，有助于进一步探讨EMX2在MA发生机制中的作用及其可能的作用通路。

苗勒管融合不全源自苗勒管发育异常。动物研究提示，众多基因与苗勒管发育有关，其同源基因在人类中可能也有类似作用。既往研究证实，Emx2基因敲除小鼠的生殖道、性腺或肾脏缺如。Emx2基因在进化中高度保守，因此EMX2基因成为了人类苗勒管发育异常的候选基因。本研究在苗勒管融合不全患者和对照组中进行了EMX2基因的全外显子测序，发现了新发无义突变c.424G>T (p.E142X)，并在此基础上构建了野生型和突变型表达载体，进行功能研究。结果提示该突变导致了EMX2蛋白的截短，野生型pCS2+MT-EMX2质粒可明显上调P63的表达，突变型pCS2+MT-EMX2质粒则无这种上调作用，并且该突变具有显性负效应，可干扰野生型EMX2的功能，可能是导致IMF表型的原因。尽管该突变的频率较低，却是第一个被证实的可导致人类苗勒管融合不全的致病基因。