Production of mouse androgenetic embryos using spindle perturbation
Production of mouse androgenetic embryos using spindle perturbation
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DOI：
10.1038/s41598-020-63010-x
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发表时间：
2020-04
影响因子：
4.6
通讯作者：
Takaya Totsuka;Miho Ohsugi
中科院分区：
文献类型：
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作者：
Takaya Totsuka;Miho Ohsugi
为了研究哺乳动物发育过程中母本和父本基因组之间的功能差异，通常使用只有一个亲本基因组的胚胎。雄激素胚胎是通过在受精之前或之后通过需要专门技能的技术去除母体染色体而产生的，并且与细胞损伤的高风险相关。在这里，我们开发了一种新的方法，用于生产雄核发育小鼠胚胎没有侵入性去核过程。我们发现，在体外受精过程中，在存在低剂量的诺考达唑，微管不稳定的药物，整个卵母细胞染色体被挤出到第二极体，导致雄激素胚胎的生产。我们进一步证明，低剂量的诺考达唑减少纺锤体的大小，防止染色体分离，但不损害卵母细胞减数分裂恢复。这导致在染色体周围形成突起，胞质分裂蛋白调节因子1（PRC1）积累到染色体周围的微管，并在突起的颈部区域组装收缩环。我们的方法使用内在的细胞动力学机制，以排除母体染色质的受精卵，并可能适用于其他哺乳动物。
To study the functional differences between maternal and paternal genomes in mammalian development, embryos with only one parental genome are often used. Androgenetic embryos are produced by the removal of maternal chromosomes before or after fertilization by techniques that require specialized skills and are associated with high risk of cellular damage. Here, we developed a novel method for producing androgenetic mouse embryos without the invasive enucleation process. We found that duringin vitrofertilization in the presence of low-dose nocodazole, a microtubule destabilizing drug, whole oocyte chromosomes were extruded into the second polar body resulting in the production of androgenetic embryos. We further demonstrated that low-dose nocodazole decreased the spindle size and prevented chromosome segregation but did not compromise oocyte meiotic resumption. This led to the formation of a protrusion around the chromosomes, accumulation of protein regulator of cytokinesis 1 (PRC1) to the microtubules around the chromosomes, and assembly of a contractile ring at the neck region of the protrusion. Our method uses the intrinsic cytokinetic mechanism to exclude maternal chromatin from zygotes and may be applicable to other mammals.