REGULATION OF PROTEIN SYNTHESIS DURING EARLY DEVELOPMENT

早期发育过程中蛋白质合成的调节

• 批准号: 3314744
• 负责人: Matthew M. Winkler
• 金额: $ 12.68万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-04-01 至 1986-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3314744
• 关键词: acidity /alkalinity; calcium; cell free system; developmental genetics; early embryonic stage; egg /ovum; fertilization; genetic translation; messenger RNA; microelectrodes; nonmammalian vertebrate embryology; radiotracer; saltwater environment

The broad objective of this project is to elucidate the mechanism by which protein synthesis is regulated in the sea urchin egg. This area has been the subject of intensive study, both because of it's intrinsic importance as a key developmental event and also because it represents a model system for understanding a type of translational regulation which probably occurs in most cell types. The major thrust of this project will be two fold. It will attempt to determine the regulatory pathways by which the known ionic changes at fertilization signal the protein synthetic machinery to activate and to determine the actual components of the protein synthetic machinery which are acted upon to increase the rate of protein synthesis (i.e. masked mRNA, ribosomes, initiation factors, etc.). There will be extensive use of cell free translation systems in which the effects of signals such as changes in pH and calcium may be studied at the level of discrete steps in the process of initiation and elongation. Various parameters of translation will be compared in cell free systems prepared from both unfertilized and fertilized eggs to identify changes at levels other than ionic controls. Both an mRNA dependent nuclease treated translation system and a partially fractionated translation system will be developed to allow a more refined analysis of the protein synthetic machinery. The role of masking of mRNAs by protein repressors will be evaluated both in vivo and in vitro, using a series of novel assays. We will test the hypothesis that the increase in histone synthesis after fertilization is regulated by ionic controls both by direct measurements with ion specific microelectrodes and by manipulation of ionic conditions both in vivo and in vitro. In addition the effects of the calcium activated changes in arginine phophate and inorganic phosphate will be determined on the specific steps in protein synthesis. This part of the project will also use a novel method for measuring adenylate charge in a single intracellular compartment. The calcium regulated activation of protein synthesis may represent the first regulatory pathway in which we can with confidence identify the initial intracellular signal, the pathway by which the signal is mediated and the mechanism by which the protein synthetic machinery responds to that signal.

该项目的总体目标是阐明其机制 海胆卵中的蛋白质合成受到调节。 该区域已被 深入研究的主题，因为它的内在重要性 作为一个关键的发展事件，也因为它代表了一个模型系统 用于理解可能发生的一种翻译调控 在大多数细胞类型中。 该项目的主要目标有两个。 它将尝试确定已知的监管途径 受精时的离子变化向蛋白质合成机制发出信号 激活并确定蛋白质合成的实际成分 提高蛋白质合成速率的机制 （即掩蔽的 mRNA、核糖体、起始因子等）。 将有 广泛使用无细胞翻译系统，其中 pH 和钙变化等信号可以在以下水平进行研究 引发和延伸过程中的离散步骤。 各种各样的 将在准备好的无细胞系统中比较翻译参数 从未受精卵和受精卵中识别水平变化 除了离子控制之外。 均经过 mRNA 依赖性核酸酶处理 翻译系统和部分分割翻译系统将是 开发用于对蛋白质合成进行更精细的分析 机械。 蛋白质阻遏物掩蔽 mRNA 的作用将是 使用一系列新颖的测定法在体内和体外进行评估。 我们 将检验以下假设：组蛋白合成增加 受精是通过直接测量的离子控制来调节的 使用离子特异性微电极并通过控制离子条件 体内和体外。 另外还有钙的作用 精氨酸磷酸盐和无机磷酸盐的激活变化将 确定蛋白质合成的具体步骤。 这部分的 项目还将使用一种新方法来测量腺苷酸电荷 单个细胞内区室。 钙调节的激活 蛋白质合成可能代表我们的第一个调控途径 可以自信地识别初始细胞内信号、通路 信号的介导方式以及蛋白质的机制 合成机械响应该信号。