THE MEMBRANE BLOCK TO POLYSPERMY IN MAMMALIAN EGGS

描述（由申请方提供）：多精受精，即一个卵子被一个以上的精子受精，被认为是人类至少5%的自发性流产的原因。为了抑制额外精子的受精，卵子开发了称为多精受精阻断的预防机制。在卵细胞外被膜水平的阻断在许多不同的动物物种中已经得到很好的表征，并且在卵质膜水平的阻断在一些非哺乳动物物种中被理解。然而，几乎没有什么是已知的膜块多精受精在哺乳动物的鸡蛋，尽管数据可以追溯到50-90年，提供证据证明它的存在。我们最近的数据表明，精子诱导的Ca 2+信号和卵细胞肌动蛋白细胞骨架是受精后变化的两个组成部分，这些变化将卵膜从支持受精的形式转变为阻止受精的形式。本项目的广泛，长期目标是阐明多精受精的膜阻断机制，从受精相关信号启动膜阻滞建立到卵膜和皮质的变化，阻止额外的受精。为了朝着这一目标取得进展，并在我们最近发表的初步数据的基础上，我们在本提案中的研究将解决以下具体目标。具体目标1将确定精子和精子诱导的Ca 2+信号传导诱导膜阻滞建立的机制。这一目标将确定哪些精子成分参与膜阻滞的建立，以及精子诱导的Ca 2+信号传导如何影响膜阻滞的特征。特异性目标2将通过鉴定参与建立多精受精膜阻断的Ca 2+依赖性效应分子来表征Ca 2+后途径的下一步。最后，具体目标3将集中在后来的步骤，最终在膜阻断多精受精，检查具体的变化，在卵膜和皮层。这一目标将测试的假设，皮质颗粒胞吐，胞吞作用，受精后皮质成分和膜顺序的变化，有助于下调受精后精子的卵膜容受性。这些调查精子诱导的信号，钙，受精后膜和皮质动力学将提供重要的见解哺乳动物膜块多精受精的细胞和分子机制，推进我们的知识，这一基本问题在生殖和发育生物学。

DESCRIPTION (provided by applicant): Polyspermy, or fertilization of an egg by more than 1 sperm, is believed to be the cause of at least 5% of spontaneous pregnancy loss in humans. To inhibit fertilization by additional sperm, eggs have developed preventative mechanisms known as blocks to polyspermy. The block at the level of the egg extra cellular coat has been well characterized in many different animal species, and the block at the level of the egg plasma membrane is understood in some non-mammalian species. However, virtually nothing is known about the membrane block to polyspermy in mammalian eggs, despite data dating back 50-90 years that provide evidence for its existence. Our recent data demonstrates that sperm-induced Ca2+ signaling and the egg actin cytoskeleton are 2 components involved in this post-fertilization change that transforms the egg membrane from a form that supports fertilization to 1 that prevents it. The broad, long-term goal of this project is elucidating the mechanism of the membrane block to polyspermy, from the fertilization-associated signaling that initiates membrane block establishment to the changes in the egg membrane and cortex that prevent additional fertilization. To make progress toward this goal and to build on our recent published and preliminary data, our studies in this proposal will address the following specific aims. Specific Aim 1 will determine the mechanisms by which sperm and sperm-induced Ca2+ signaling induce the establishment of the membrane block. This aim will identify what sperm component(s) is involved in membrane block establishment and how sperm-induced Ca2+ signaling affects characteristics of the membrane block. Specific Aim 2 will characterize the next step in the pathway following Ca2+ by identifying the Ca2+-dependent effector molecules that are involved in the establishment of the membrane block to polyspermy. Finally, Specific Aim 3 will focus on the later steps of the pathway culminating in the membrane block to polyspermy, examining specific changes in the egg membrane and cortex. This aim will test the hypotheses that cortical granule exocytosis, endocytosis, and post-fertilization changes in cortex composition and membrane order contribute to the down-regulation of egg membrane receptivity to sperm that follows fertilization. These inquiries into sperm-induced signaling, calcium, and post-fertilization membrane and cortical dynamics will provide important insights into the cellular and molecular mechanisms underlying the mammalian membrane block to polyspermy, advancing our knowledge of this fundamental question in reproductive and developmental biology.