母源因子NKX3-1调控人类植入前胚胎基因组激活的分子机制研究

胚胎基因组激活（EGA）是囊胚发育的关键，但具体机制尚未明确。我们基于课题组已发表的人类植入前胚胎转录组测序数据发现minor EGA 基因的转录因子预测显著富集于NKX3-1，并且NKX3-1过表达显著上调EGA基因。因此，我们提出母源因子NKX3-1是植入前胚胎EGA激活的关键因子的假说。本项目拟在前期基础上，首先利用已构建的Nkx3-1基因敲除鼠，系统评估Nkx3-1基因敲除对早期胚胎发育的影响。其次通过敲除和野生型鼠卵子及植入前胚胎的转录组测序分析，明确NKX3-1对植入前胚胎EGA激活的重要作用。最后利用NKX3-1稳定过表达hESCs，免疫共沉淀质谱分析筛选NKX3-1调控EGA激活的关键互作蛋白，并在细胞和胚胎水平对其进行验证，初步揭示NKX3-1调控EGA激活的分子机制。本项目的完成将有助于深入理解EGA激活的母源调控机制，为辅助生殖临床上胚胎发育阻滞的溯源分析提供新思路。

Embryonic genome activation (EGA) is the key factor of blastocyst development, but its mechanism has not been known well. We screened the novel critical activator genes of embryonic genome based on our previously published transcriptome sequencing data in human preimplantation embryos. We performed the prediction of transcription factors involved in minor EGA and identified NKX3-1 as the most enriched gene which may obviously upregulate expression of EGA genes when NKX3-1 gets over expressed. Therefore, we suppose that maternal factor NKX3-1 may play an important role in EGA. Based on the previous results, this project is planning to systematically evaluate the effect of Nkx3-1 deficiency on embryonic development using the constructed Nkx3-1 knockout mice. Next, we will determine the important role of NKX3-1 in EGA by transcriptome sequencing analyzing on oocytes and early embryos from both Nkx3-1 knockout mice and wild-type counterparts. Finally, with the established NKX3-1 stably over-expressed human embryonic stem cell line, screening of interacting proteins associated with NKX3-1 initiating EGA transcription will be achieved by co-immunoprecipitation and mass spectrometry analysis. The completion of the project is of great help to understand maternal regulation mechanisms of EGA in depth, and provides new ideas and important scientific basis for the origin study of embryo developmental block in assisted reproduction field.