The apportionment of zygotic totipotency: Origins, mechanisms and consequences for natural and assisted reproduction in the mouse model

在哺乳动物中，来自第一次合子分裂的两个卵裂球被认为是同样全能的，能够形成两个同卵双胞胎。然而，第一个卵裂面在不同的胚胎之间是不同的，卵质的成分在两个卵裂球之间的分配是不同的，这对同等的潜力提出了质疑。我们认为，卵胞浆内单精子注射--广泛应用于辅助生殖--通过施加卵母细胞受精的部位和卵裂方向，有助于以可控的方式研究潜力。受非洲爪哇和海胆数据的启发，我们假设，当第一个卵裂面沿着动植物轴线运行时，小鼠的卵裂球最相似，而当第一个卵裂面垂直运行时，它们之间的差异最大。卵裂球的转录将与我们最近对传统2-细胞胚胎的荟萃分析相结合，根据我们的假设，Tead1和Cops3的转录模式确实与卵裂球的相似性相关。可能在本研究中发现了额外的基因(S)X。为了达到超越相关性并确定因果关系，我们在受精前直接在蛋白质水平上消耗卵母细胞中的基因产物。我们评估发育后果，并对Tead1-、Cops3-和X基因缺失的胚胎进行蛋白质组学研究，以确认蛋白质缺失的特异性。在项目过程中，我们将回答两个问题：1)第一个卵裂面的方向是否以及如何导致卵裂球之间的功能差异；以及2)哺乳动物卵母细胞中母体蛋白的沉积是否以及如何对潜力和发育起决定性作用。

In mammals, the two blastomeres resulting from the 1st zygotic division are deemed equally totipotent, able to form two monozygotic twins. However, the first cleavage plane differs among embryos, distributing the constituents of the ooplasm differently among the two blastomeres, questioning equal potentiality. We propose that intracytoplasmic sperm injection –widely used in assisted reproduction– helps to investigate potentiality in a controlled manner, by imposing the site of oocyte fertilization and thus cleavage orientation. Motivated by Xenopus and sea urchin data, we hypothesize that the mouse blastomeres are most similar to each other when the 1st cleavage plane runs along the animal-vegetal axis, yet they are most different from each other when it runs perpendicular. The transcriptomes of blastomeres will be combined with those of our recent meta-analysis of conventional 2-cell embryos, where transcription patterns of, e.g., Tead1 and Cops3 do correlate with blastomere similarity as per our hypothesis. Additional gene(s) X may be revealed in the present study. To reach beyond correlation and establish causality, we deplete gene products directly at the protein level in oocytes prior to fertilization. We assess developmental consequences and subject Tead1-, Cops3- and gene X-depleted embryos to proteomics to confirm the specificity of the protein depletion. During the course of the project we shall answer two questions: 1) if and how the orientation of the 1st cleavage plane introduces functional variation among the blastomeres; and 2) if and how maternal protein deposits in mammalian oocytes are decisive for potentiality and development.