Effects of sex chromosomes on the developmental potential of the mouse oocyte

性染色体对小鼠卵母细胞发育潜力的影响

• 批准号: RGPIN-2018-04464
• 负责人: TaketoHosotani, Teruko
• 金额: $ 2.62万
• 依托单位: McGill University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=686607
• 关键词: Effects; sex; chromosomes; developmental; potential

My long-term objective is to delineate how sex chromosomes affect the developmental potential of mouse oocytes. In mammalian development, germ cells undergo sexual differentiation according to their gonadal environment, testis or ovary, which is determined by the presence or absence of the Y-linked Sry gene. Therefore, spermatogenesis and oogenesis take place in the presence of XY and XX chromosomes, respectively. The gonadal sex also dictates the phenotypic sex by secretion of sex steroid hormones. When the gonadal sex is reversed, however, the germ cell sex becomes discordant with the chromosomal sex. Both XX males and XY females in humans are infertile, while XY females are fertile in certain rodent species. In Mus musculus, XY females are variably fertile depending on the cause of sex-reversal and genetic background. The B6.YTIR mouse, which develops into an infertile female with an intact Y chromosome, provides an excellent model for studying the effects of sex chromosomes on female fertility. We previously reported that the cause of infertility in the B6.YTIR female is intrinsic to its oocytes; they can reach the Metaphase II (MII) but fail to complete the second meiotic division and rarely develop beyond the zytote-stage after fertilization. This developmental incompetence can be attributed to their defective ooplasm; when XY oocyte nuclei have been transferred into enucleated XX oocytes, the reconstructed oocytes generate healthy offspring. Thus, the Y chromosome in the oocyte makes the ooplasm defective, which leaves the first meiotic division largely intact but disrupts the second meiotic division. In the next 5 years, we will investigate the molecular mechanisms underlying the second meiotic defects in the XY oocyte and investigate the involvement of Y-linked gene(s). (1) To characterize the nature of ooplasmic defects, whether it lies in a deficiency in critical components or the presence of detrimental components, we will transfer a small volume of cytoplasm from an XX oocyte into an XY oocyte or vice versa, followed by maturation in vitro, and examine the second meiotic division after parthenogenic activation (or fertilization). (2) To clarify the molecular mechanism underlying the failure in the second meiotic cytokinesis, we will compare the activity of ARP2/3 pathways, which are essential for maintaining the MII-spindle near the cytokinesis furrow, in XX and XY oocytes before and after activation. (3) To evaluate the role of Y-encoded transcription factor ZFY2 in the ooplasmic defects, we will knock-down its transcripts in the XY oocyte by siRNAs expression in vitro or by shRNAs expression from a transgene in vivo. The results of proposed studies will shed light on the mechanism coordinating the sister-chromatid separation and the second polar body extrusion, and evolution of the Y chromosome associated with the loss of fertility in Mus musculus XY females.

我的长期目标是描绘性染色体如何影响小鼠卵母细胞的发育潜力。在哺乳动物的发育中，生殖细胞根据其性腺环境、睾丸或卵巢进行性别分化，这是由Y连锁Sry基因的存在或不存在决定的。因此，精子发生和卵子发生分别在存在XY和XX染色体的情况下发生。性腺的性交也通过分泌性类固醇激素来决定表型性行为。然而，当性腺性别颠倒时，生殖细胞的性别与染色体的性别变得不一致。在人类中，XX雄性和XY雌性都是不育的，而XY雌性在某些啮齿动物物种中是可生育的。在小鼠中，XY雌鼠具有不同的生育能力，这取决于性别逆转的原因和遗传背景。B6.YTIR小鼠发育成Y染色体完整的不育雌性小鼠，为研究性染色体对雌性生育能力的影响提供了极好的模型。我们以前报道过B6雌性不育的原因是其卵母细胞所固有的，它们可以达到中期II期(MII)，但无法完成第二次减数分裂，很少在受精后发育到合子阶段。这种发育不全可以归因于他们的卵质缺陷；当XY卵母细胞核被转移到去核的XX卵母细胞中时，重建的卵母细胞产生健康的后代。因此，卵母细胞中的Y染色体使卵质有缺陷，这使得第一次减数分裂基本保持不变，但扰乱了第二次减数分裂。在接下来的5年里，我们将研究XY卵母细胞第二次减数分裂缺陷的分子机制，并探讨Y连锁基因的参与(S)。(1)为了确定卵质缺陷的性质，无论是关键成分的缺乏还是有害成分的存在，我们将一小部分细胞质从XX卵母细胞移植到XY卵母细胞，反之亦然，然后进行体外成熟，并观察孤雌激活(或受精)后的第二次减数分裂。(2)为了阐明第二次减数分裂失败的分子机制，我们比较了XX和XY卵母细胞激活前后Arp2/3通路的活性，Arp2/3通路是维持胞质分裂沟附近的微纺锤体所必需的。(3)为了探讨Y编码的转录因子ZFY2在卵浆缺陷中的作用，我们在体外通过siRNAs表达或通过体内转基因shRNAs表达来敲除其在XY卵母细胞中的转录。这些研究结果将有助于阐明姐妹染色单体分离和第二极体排出的协调机制，以及Y染色体的进化与XY雌性小鼠生育能力丧失的关系。