Molecular Mechanisms of Meiotic Maturation in Drosophila

描述（由申请人提供）：减数分裂是一个与配子发育相关的基本过程。卵母细胞在第一次减数分裂的前期停滞以允许生长。减数分裂成熟过程中会发生前期I的释放，但调控这一过程的分子机制还不完全清楚。减数分裂成熟缺陷可导致不育或染色体分离不准确，导致流产和/或出生缺陷。果蝇（Drosophila melanogaster）提供了复杂的遗传和分子工具，以及描述良好的卵巢细胞生物学，使其成为研究减数分裂成熟控制的强大系统。果蝇卵母细胞在胚系包囊形成后不久进入减数分裂，并在其发育的大部分时间内停滞在前期I。减数分裂成熟后，卵母细胞再次停滞在中期I，直至排卵。在之前的资助期间，我们证明了1-Endosulfine（Endos）是减数分裂成熟所必需的。我们的数据表明，Endos促进细胞周期调节因子CyclinA、波罗和Twine/Cdc 25（均已知为后期促进复合物或APC的靶标）的稳定性，并且它还具有通过抑制E3泛素连接酶Elgi促进减数分裂成熟的单独作用。ENSA（人1-内砜）的表达挽救了内减数分裂缺陷，1-内砜在小鼠卵母细胞中表达，表明1-内砜的减数分裂功能保守。我们的总体假设是，Endos是通过两种机制的减数分裂成熟的中央调节器：它抑制APC的活性，允许关键细胞周期调节剂的积累，它拮抗Elgi靶标的泛素化。我们将通过以下具体目的来检验这一假设：1）确定Endos是否是APC的负调节剂; 2）确定E3泛素连接酶Elgi如何调节Endos下游的减数分裂成熟;以及3）阐明Endos作用的分子机制。阐明减数分裂成熟的分子机制可能会导致治疗不孕症的原因，以及新的避孕药。此外，这些知识也可能导致人类卵母细胞体外成熟的技术策略，以补充目前的体外受精方法。 公共卫生相关性：卵母细胞减数分裂成熟失败导致不育。我们先前的工作已经确定了Endos作为减数分裂成熟的主要调节因子，我们建议在果蝇中使用强大的研究工具来了解这种蛋白质的功能。我们的研究可能会导致更多的治疗方法来解决不孕症，以及技术策略，以补充目前的体外受精方法。

DESCRIPTION (provided by applicant): Meiosis is a fundamental process that is coupled to gamete development. Oocytes arrest in prophase of the first meiotic division to allow for growth. Release from prophase I occurs during meiotic maturation, but the molecular mechanisms regulating this process are incompletely understood. Meiotic maturation defects can result in infertility or in inaccurate segregation of chromosomes, leading to miscarriages and/or birth defects. The fruitfly Drosophila melanogaster offers sophisticated genetic and molecular tools, and a well described ovarian cell biology, making it a powerful system to investigate the control of meiotic maturation. Drosophila oocytes enter meiosis shortly after germline cyst formation and arrest in prophase I during most of their development. After meiotic maturation, oocytes arrest again in metaphase I until ovulation. In the prior funding period, we demonstrated that 1-Endosulfine (Endos) is required for meiotic maturation. Our data suggest that Endos promotes the stability of the cell cycle regulators CyclinA, Polo and Twine/Cdc25 (all known to be targets of the Anaphase Promoting Complex, or APC), and that it also has a separate role via inhibition of the E3 ubiquitin ligase Elgi to promote meiotic maturation. Expression of ENSA, the human 1- endosulfine, rescues the endos meiotic defect, and 1-endosulfine is expressed in mouse oocytes, suggesting conservation of the meiotic function of 1-endosulfine. Our overarching hypothesis is that Endos is a central regulator of meiotic maturation via two mechanisms: it inhibits the activity of the APC to allow the accumulation of key cell cycle regulators, and it antagonizes the ubiquitination of Elgi targets. We will test this hypothesis through the following specific aims: 1) to determine if Endos is a negative regulator of the APC; 2) to determine how the E3 ubiquitin ligase Elgi regulates meiotic maturation downstream of Endos; and 3) to elucidate the molecular mechanism of action of Endos. Elucidation of the molecular mechanisms of meiotic maturation could potentially lead to therapies to address this cause of infertility, as well as to new contraceptives. Furthermore, this knowledge could also lead to technological strategies for in vitro maturation of human oocytes to complement current in vitro fertilization methodology. PUBLIC HEALTH RELEVANCE: Failure of the oocyte to undergo meiotic maturation leads to infertility. Our prior work has identified Endos as a major regulator of meiotic maturation, and we propose to use powerful research tools in fruitflies to understand the function of this protein. Our studies could potentially lead to additional therapies to address infertility, as well as to technological strategies to complement current in vitro fertilization methodology.