MOLECULAR ANALYSIS OF THE SEA URCHIN SPERM RECEPTOR

海胆精子受体的分子分析

• 批准号: 2203027
• 负责人: KATHLEEN R FOLTZ
• 金额: $ 10.15万
• 依托单位: UNIVERSITY OF CALIFORNIA SANTA BARBARA
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-08-01 至 1998-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2203027
• 关键词: Asteroidea; affinity chromatography; alternatives to animals in research; antireceptor antibody; cell membrane; complementary DNA; crosslink; cytoplasm; cytoskeletal proteins; egg /ovum; immunofluorescence technique; immunoprecipitation; microinjections; monoclonal antibody; peptides; phosphorylation; protein tyrosine kinase; receptor binding; receptor expression; recombinant proteins; sea urchins; sperm

Fertilization is the result of a set of well-choreographed events between the sperm and the egg. Successful gamete interactions result in the activation of the egg and initiate developmental pathways. Unsuccessful interactions, as a result of either sperm or egg dysfunction, are disastrous. Although much is known about the steps preceding binding and sperm-egg fusion, the mechanism of the binding event and the resulting signal transduction pathways that occur within the egg have been elusive. The recent identification and cloning of the sea urchin egg plasma membrane receptor for sperm now makes it possible to investigate these questions and to determine how the receptor functions at egg activation. This proposal describes experiments designed to address this structure/function question in the sea urchin, a well-characterized model system for fertilization and early development. The general question centers on the role that the receptor plays in transducing a signal in response to sperm binding, be it direct or indirect. Our preliminary data indicate that the receptor interacts with an egg cortical protein tyrosine kinase and may serve to organize the actin cytoskeletal rearrangements that occur at fertilization. The specific aims are: (i) To define the topology of the receptor in the membrane in order to establish which domains may interact with other egg proteins; (ii) To identify egg proteins that interact with the receptor; (iii) To test the functional role(s) of the cytoplasmic domain by injection of specific antibodies and peptides; and (iv) To initiate mutational analyses of the receptor, employing a heterologous expression system. These experiments will provide for a greater understanding of how sperm binding triggers the changes that occur in the egg, adding to our general knowledge of the basic processes of fertilization so important to our understanding of fertility and infertility.

受精是一系列精心设计的事件的结果 精子和卵子。成功的配子相互作用会导致 激活卵子并启动发育途径。不成功 由于精子或卵子功能障碍而产生的相互作用 灾难性的。尽管我们对绑定和 精子-卵子融合，结合事件的机制和导致的 卵子内的信号转导途径一直难以捉摸。 海胆卵浆的最新鉴定和克隆 精子的膜受体现在使研究这些成为可能 问题，并确定受体在卵子激活时如何发挥作用。 这份提案描述了为解决这一问题而设计的实验 海胆的结构/功能问题--一个很有特色的模型 受精和早期发育的系统。一般问题是 集中在受体在信号转导中所起的作用 对精子结合的反应，无论是直接的还是间接的。我们的初步数据 表明该受体与鸡蛋皮质蛋白酪氨酸相互作用 并可能用于组织肌动蛋白的细胞骨架重排。 在受精时发生。具体目的是：(I)界定 膜上受体的拓扑结构，以确定 结构域可能与其他鸡蛋蛋白相互作用；(Ii)识别鸡蛋 与受体相互作用的蛋白质；(Iii)测试 注射特异性抗体和抗体在细胞质结构域中的作用(S) 多肽；以及(Iv)启动该受体的突变分析， 使用异源表达系统。这些实验将提供 为了更好地理解精子结合是如何触发这种变化的 发生在鸡蛋中，增加了我们对基本过程的一般知识 受精对我们理解生育能力和 不孕不育。