Do we need Y chromosome for successful reproduction?

哺乳动物的生殖通常需要雌性提供卵母细胞，雄性提供精子来实现受精。我们超越了这一传统，提出了一个假设，即任何性别的成年个体都可以被诱导产生异性的配子，这些配子在辅助受精中起作用。这一假设的前提是，我们最近发表的研究表明，具有有限或没有Y染色体基因的基因工程雄性小鼠可以通过辅助受精（ART）成功繁殖，以及细胞重编程和分化领域的重大进展。我们的实验室已经证明，在小鼠中，只有两个Y染色体基因，睾丸决定因素Sry和精子发生驱动因素Eif2s3y，足以使雄性产生具有ART功能的单倍体雄性配子。我们随后证明，这两个基因的功能可以被编码在其他染色体上的同源基因所取代，并且缺乏所有Y染色体基因的单个X染色体（XO）小鼠在辅助受精后可以产生雄性配子并产生健康的后代。我们的合作者Mitinori saiitou的实验室已经证明，从诱导多能干细胞（iPSC）分化为原始生殖细胞样细胞（PGCLC）可以获得雄性和雌性配子。在这个提案中，我们将我们的发现和专业知识结合起来，提出三个问题：成年雌鼠能否产生辅助受精的雄性配子？目标2。成年雄鼠能否产生辅助受精的雌性配子？目标3。任何一种性别的成年老鼠都能产生孤代后代吗？为了解决这些问题，我们将从给定性别的成年小鼠中培养体细胞系，重新编程为iPSC，鉴定失去一条性染色体并成为XO的克隆，并分化为PGCLC。为了产生雄性配子，我们将转基因地添加精子发生驱动因子，并将PGCLC移植到新生儿雄性的睾丸中。为了产生雌性配子，我们将在体外重建卵巢并将其移植到受体雌性的卵巢囊下。我们将使用辅助生殖技术来测试这些衍生的雄性和雌性配子的功能。为了产生孤代后代，将使用来自同一个体的雄性和雌性配子进行受精。这个项目的发现将影响我们对性别特异性差异的理解，特别是关于性染色体和X、Y基因对生殖细胞发育的影响。如果成功，我们还将为该领域提供一个原理证明，即从一个个体中可以获得两性后代，这将对物种保护产生影响。

Mammalian reproduction conventionally requires oocytes provided by a female and sperm provided by a male to achieve fertilization. We go beyond this convention and propose the hypothesis that an adult individual of any sex can be induced to produce gametes of the opposite sex, and that these gametes are functional in assisted fertilization. The premise for this hypothesis comes from our recently published studies demonstrating that genetically engineered male mice with limited or no Y chromosome genes can successfully reproduce by assisted fertilization (ART), and from significant advancements in field of cell reprogramming and differentiation. Our laboratory has shown that in the mouse only two Y chromosome genes, testis determinant Sry and spermatogenesis driver Eif2s3y, are sufficient for a male to produce haploid male gametes functional in ART. We have subsequently demonstrated that the function of these two genes could be replaced by that of their homologues encoded on other chromosomes, and that a mouse with a single X chromosome (XO) lacking all Y chromosome genes can produce male gametes and sire healthy offspring after assisted fertilization. The laboratory of our collaborator, Mitinori Saitou, has shown that both male and female gametes can be obtained from induced pluripotent stem cells (iPSC) differentiated into primordial germ cell like-cells (PGCLC). In this proposal, we marry our findings and expertise and ask 3 questions: Aim 1. Can an adult female mouse produce male gametes functional in assisted fertilization? Aim 2. Can an adult male mouse produce female gametes functional in assisted fertilization? Aim 3. Can an adult mouse of either sex sire uniparental offspring? To address these questions we will develop somatic cell lines from an adult mouse of a given sex, reprogram to iPSC, identify the clones that have lost one sex chromosome and became XO, and differentiate into PGCLC. To produce male gametes we will transgenically add a spermatogenesis driver and transplant PGCLC to testes from neonatal males. To produce female gametes we will reconstitute ovaries in vitro and transplant them under ovarian bursa of recipient females. We will test the function of such derived male and female gametes using assisted reproduction. To produce uniparental offspring, male and female gametes derived from the same individual will be used for fertilization. The findings from this project will impact on our understanding of sex specific differences, especially pertaining to effects of sex chromosomes and X and Y genes on germline development. If successful, we will also provide the field with a proof-of-principle that offspring of both sexes can be obtained from a single individual, which will impact on species preservation.