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.

该项目的广泛目的是阐明 蛋白质合成受到海胆卵的调节。 这个区域已经 强化研究的主题，既是因为它的内在重要性 作为关键发展事件，也是因为它代表模型系统 了解可能发生的一种翻译法规 在大多数细胞类型中。 该项目的主要推力将是两个折。 它将尝试确定已知的调节途径 受精时的离子变化向蛋白质合成机制表明 激活并确定蛋白质合成的实际成分 作用于增加蛋白质合成速率的机械 （即掩盖的mRNA，核糖体，起始因子等）。 将有 广泛使用无细胞翻译系统，其中的影响 可以研究诸如pH和钙的变化之类的信号。 在开始和伸长过程中的离散步骤。 各种各样的 在准备的无细胞系统中，将比较翻译的参数 从未施肥和受精卵中识别水平的变化 除了离子控制。 两者都依赖于mRNA的核酸酶 翻译系统和部分分解的翻译系统将是 开发以允许对蛋白质合成的更精致的分析 机械。 蛋白质阻遏物对mRNA掩盖的作用将是 使用一系列新颖的测定法，对体内和体外进行了评估。 我们 将检验以下假设 通过直接测量，由离子对照来调节受精 与离子特异性微电极和离子条件的操纵 体内和体外。 此外，钙的影响 精氨酸磷酸盐和无机磷酸盐的活化变化将是 确定蛋白质合成的特定步骤。 这部分 项目还将使用一种新方法来测量在 单细胞内室。 钙调节的激活 蛋白质合成可能代表了我们的第一个调节途径 可以置信处识别初始细胞内信号，即途径 信号介导的信号和蛋白质的机制 合成机械响应该信号。