Drosophila as a model to dissect the rise and role of calcium in egg activation.

项目摘要 分化的成熟卵母细胞被“激活”成为全能的并开始胚胎。 其发育通过胞质内游离钙离子水平的增加来实现。在哺乳动物中，这是 增加最初是由受精的精子诱导的，它刺激从 内部商店。然而，最近的研究表明，随后的钙振荡 对于哺乳动物卵子的激活是必不可少的，需要外部钙离子从 环境通过尚不清楚的途径和机制进入卵母细胞。 尽管果蝇并不被认为是这些过程的好模型 因为它的卵在受精前激活，我们最近的研究表明，果蝇 卵子的激活也涉及到局部的钙离子内流。在这种涌入之后，伴随着一波 利用类似这样的机制，增加了扫过卵子细胞质的钙离子 哺乳动物卵母细胞使用的。我们发现果蝇体内发生了最初的钙离子内流 在卵母细胞极，通过机械敏感通道-很可能是保守的 TRP系列中的频道。我们建议建立果蝇作为一种新的模型 以Ca~(2+)为基础的卵子激活阶段。我们将使用此模型来识别 介导精子非依赖性钙离子内流及引起钙通道的条件 只能在蛋杆上打开。我们还将确定增加的钙离子是否 调节通常伴随卵子的卵母细胞磷蛋白质组的变化 激活，以及这些细胞是否需要钙信号转导蛋白钙调神经磷酸酶 磷酸蛋白质组的变化。我们希望我们的研究结果可以推广到 哺乳动物卵子激活的机制。此外，含有钙离子的保守蛋白质- 依赖的磷化修饰将成为未来测试的候选对象 评估人类ART条件、卵母细胞质量和辅助诊断不育症 这不能回应当前的艺术方法。

Project Summary A differentiated, mature oocyte is “activated” to become totipotent and to begin embryo development through an increase in its cytoplasmic free Ca2+ levels. In mammals, this increase is initially induced by the fertilizing sperm, which stimulates release of Ca2+ from internal stores. However, recent studies have shown that subsequent Ca2+ oscillations that are essential for mammalian egg activation require influx of external Ca2+ from the environment into the oocyte, through as yet unknown channels and mechanisms. Although Drosophila had not been considered a good model for these processes because its eggs activate prior to fertilization, our recent work has shown that Drosophila egg activation also involves a local Ca2+ influx. This influx is followed by a wave of increased Ca2+ that sweeps through the egg cytoplasm, using mechanisms like those used by mammalian oocytes. We showed that the initial Ca2+ influx in Drosophila occurs at the oocyte poles, through mechanosensitive channels – most likely conserved channels in the TRP family. We propose to establish Drosophila as a new model for the Ca2+-based phase of egg activation. We will use this model to identify the channels that mediate sperm-independent Ca2+ influx and the conditions that cause Ca2+ channels to be open only at the egg poles. We will also determine whether the increased Ca2+ mediates the changes to the oocyte phosphoproteome that normally accompany egg activation, and whether the Ca2+ signal transducer calcineurin is required for these phosphoproteome changes. We expect the results of our studies to be generalizable to mechanisms in mammalian egg activation. Moreover, conserved proteins with Ca2+- dependent phosphomodifications will be candidates for future testing as markers for assessing human ART conditions, oocyte quality and aiding the diagnosis of infertilities that cannot respond to current ART methods.