Modification of the Egg Extracellular Matrix at Fertilization

受精时卵细胞外基质的修饰

• 批准号: 0315657
• 负责人: Gary Wessel
• 金额: $ 36万
• 依托单位: Brown University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2006-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0315657&HistoricalAwards=false
• 关键词: Modification; Egg; Extracellular; Matrix; Fertilization

0315657WesselSuccess at fertilization requires both sperm-egg fusion and blockage of excess sperm fusions, or polyspermy. Polyspermy is lethal in most animals, and oocytes have evolved mechanisms to minimize its occurrence. The most conserved mechanism throughout phylogeny involves a rapid remodeling of the egg extracellular matrix following fertilization by secretion of cortical granules, which are stimulus-dependent secretory vesicles unique to eggs and oocytes. Cortical granule contents modify the nascent egg extracellular matrix to form both a permanent block to polyspermy and to provide protection for early embryonic development. The general question addressed in this application is: How do eggs so rapidly and efficiently construct their new extracellular matrix? We will use sea urchin eggs for this project because: 1) we can obtain, on average, 106 eggs per female, enabling large-scale biochemical approaches; 2) each of the constituents of this process - the cortical granules, the fertilization envelope, and the vitelline layer, can be isolated en masse in a functional form; 3) cDNA clones have been isolated that represent each major functional protein of this process; and 4) sea urchin eggs undergo one of the most extreme modifications at fertilization. They invest 6% of their protein mass and nearly 20% of their mRNA to this transformation of the nascent extracellular, vitelline layer into an impervious fertilization envelope within 60 seconds.The broader implications of this project are that undergraduate and graduate students will be trained in diverse contemporary methods of cell, molecular, and developmental biology, and that general principles of fertilization and extracellular matrix biology will be integrated in a classically described, tractable system for educational purposes. In addition, project results and other educational material for K-16 will be continually updated on the Educational Resources page of our lab web site (http://www.brown.edu/Research/Wessel_Lab/).Three Specific Research Aims are proposed:1. Identify the vitelline layer proteins essential for fertilization envelope construction. 2. Determine the domains in cortical granule proteins required for construction of the extracellular matrix and block to polyspermy at fertilization. 3. Identify the peroxide-generating mechanism essential for envelope cross-linking.

成功的受精需要精卵融合和阻止多余的精子融合，或多精受精。多精受精对大多数动物来说都是致命的，卵母细胞已经进化出机制来尽量减少其发生。整个胚胎发育过程中最保守的机制涉及受精后通过分泌皮质颗粒快速重塑卵细胞外基质，皮质颗粒是卵和卵母细胞特有的刺激依赖性分泌囊泡。皮质颗粒内容物修饰新生卵细胞外基质，形成对多精受精的永久性阻断，并为早期胚胎发育提供保护。在这个应用程序中解决的一般问题是：鸡蛋是如何如此迅速和有效地构建其新的细胞外基质？我们将使用海胆卵进行这个项目，因为：1）我们可以获得，平均每只雌性106个卵，使大规模的生物化学方法成为可能; 2）这个过程的每一个组成部分-皮层颗粒，受精包膜和卵黄层，可以以功能形式分离出来; 3）cDNA克隆已经被分离出来，代表了这一过程中的每一个主要功能蛋白; 4）海胆卵在受精时经历了最极端的修改之一。他们投入6%的蛋白质质量和近20%的mRNA，在60秒内将新生的细胞外卵黄层转化为不透水的受精膜。该项目的更广泛意义是，本科生和研究生将接受细胞，分子和发育生物学的各种当代方法的培训，受精和细胞外基质生物学的一般原理将被整合到一个经典描述的、易于处理的系统中，用于教育目的。此外，项目结果和其他教育材料的K-16将不断更新的教育资源页面上我们的实验室网站（http：//www.brown.edu/Research/Wessel_Lab/）。 鉴定受精被膜构建所必需的卵黄层蛋白。2. 确定构建细胞外基质所需的皮质颗粒蛋白的结构域 并在受精时阻止多精受精。3. 确定信封交联必不可少的过氧化物生成机制。