组蛋白H3K9甲基化对卵母细胞成熟和早期胚胎发育调控作用的研究

Assisted reproductive technology is currently the main technology for the infertile couples to give a birth to child, but the quality of oocytes by artificial hormones stimulated and subsequent embryo development potential are relatively low. During oocyte maturation and early embryo development there are many epigenetic modifications which alter significant, and the epigenetic alteration plays an important role in regulation of oocyte maturation and early embryo development. Our preliminary experimental results show: overexpressing histone methyltransferase SUV39H1 in oocytes can significantly increase the level of H3K9 methylation, promote the oocyte maturation. This study intends to further analysis the regulatory mechanism of histone H3K9 methylation on oocyte maturation and early embryo development. We first build the expression plasmid of the selected methyltransferase and demethylase, then get mRNA by in vitro transcription, tailing and purification. By using microinjection, the mRNA will be injected into the oocytes of mice and human to detect the impact on H3K9 methylation modification, recruitment of heterochromatin protein HP1, chromatin condensation and germinal vesicle breakdown etc; then ICSI will be performed to detect the early embryo development after fertilization. The results of this research will help to understand the role of epigenetic modifications in the process of reproduction, and will be expected to improve in vitro culture technique of oocytes and embryos.

目前辅助生殖技术是不孕不育夫妇实现生育愿望的主要技术手段，然而人为激素刺激获得的卵母细胞质量及其胚胎发育潜力比较低。卵母细胞发育成熟及早期胚胎发育过程中伴随着诸多表观遗传修饰的变化，这些变化对卵母细胞成熟和早期胚胎发育起着重要的调控作用。我们的前期实验证明：卵母细胞中过表达组蛋白甲基转移酶SUV39H1能够显著增加H3K9甲基化的修饰水平，促进卵母细胞成熟。本研究拟进一步深入分析组蛋白H3K9甲基化对卵母细胞成熟和早期胚胎发育的调控机制。首先构建选定的组蛋白甲基转移酶和去甲基化酶表达质粒，经过体外转录、加尾和纯化获得mRNA。利用显微注射方法将其注入到人和小鼠卵母细胞中，检测是否影响H3K9甲基化修饰、异染色质蛋白HP1募集、染色质凝聚及生发泡破裂等；利用单精注射技术检测受精后早期胚胎发育情况。本项目的研究结果将有助于理解生殖过程中表观遗传修饰的作用，并有望改进卵母细胞和胚胎体外培养技术。