REGULATION OF TRANSLATION DURING EARLY DEVELOPMENT

早期发展过程中的翻译监管

• 批准号: 3315903
• 负责人: JOHN W HERSHEY
• 金额: $ 5.56万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-04-01 至 1987-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3315903
• 关键词: cytoskeleton; early embryonic stage; eukaryote; fertilization; gel electrophoresis; gene expression; genetic translation; immature animal; immunochemistry; messenger RNA; nonmammalian vertebrate embryology; saltwater environment

This proposal addresses the question of translational regulation as a fundamental element in the control of gene expression during early development. Specifically, we are asking how protein synthesis is activated following fertilization and how it is regulated during embryogenesis. The system chosen for this study is the sea urchin egg and developing embryo. The general approach proposed is to purify and characterize the key components of translation, and develop tools which can monitor changes in the levels, forms and activities of these components, to ultimately discover the mechanisms which govern their interactions. First, we shall purify and characterize the initiation factors of protein synthesis from sea urchin eggs and embryos and produce antibodies against the purified factors. Using immunochemical and gel electrophoretic techniques, we will determine levels and molecular forms (e.g., phosphorylation) of the factors in unfertilized eggs and embryos. Second, sea urchin egg cell-free lysates active in protein synthesis will be prepared following published procedures. We will also construct an assay system entirely of highly purified sea urchin components, and use these two assays for detailed analysis of the role of the purified translational components of the initiation pathway in activation of protein synthesis. Third, we will examine changes in ribosomal proteins during fertilization and early development, and establish how changes are linked to the translational activation at fertilization. The role of cytoskeletal changes during early development in modulating the rate of protein synthesis will also be studied. Finally, we shall purify and characterize the mRNA and proteins of bulk and specific messenger rebonucleoprotein particles using immunological and gel electrophoretic techniques and try to identify a masking factor. The tools developed and the information obtained from these experiments will then allow us to elucidate the molecular mechanisms of mobilization of mRNP into polysomes. These studies will elucidate not only important controlling mechanisms during early development, but also important general mechanisms of translational control in all eukaryotic cells.

该提案将翻译法规问题作为一种 早期基因表达调控的基本要素 发展。具体地说，我们问的是蛋白质合成是如何 受精后激活及其在受精过程中的调节 胚胎发生。本研究选择的系统是海胆卵和 发育中的胚胎。建议的一般方法是净化和 描述翻译的关键组成部分，并开发能够 监控这些组件的级别、形式和活动的变化，以 最终发现支配它们相互作用的机制。第一, 我们将提纯和鉴定蛋白质的启动因子。 从海胆卵和胚胎中合成并产生抗体 纯化因子。利用免疫化学和凝胶电泳法 技术，我们将确定水平和分子形式(例如， 在未受精卵和胚胎中的因子的磷酸化)。第二, 海胆卵细胞裂解物在蛋白质合成中的活性将 按照已公布的程序编写。我们还将构建一种分析方法 系统完全由高纯度的海胆成分组成，并使用这两种 用于详细分析纯化的翻译的作用的分析方法 激活蛋白质合成的起始途径的组成部分。 第三，我们将检测受精过程中核糖体蛋白的变化。 和早期开发，并确定变化如何与 受精时的翻译激活。细胞骨架的作用 早期发育过程中调节蛋白质速率的变化 合成也将被研究。最后，我们将提纯和描述 大宗信使和特异性信使核糖核蛋白的mRNA和蛋白质 粒子使用免疫学和凝胶电泳技术，并试图 确定掩蔽因素。开发的工具和信息 从这些实验中获得的结果将使我们能够阐明 将mRNP动员到多聚体的分子机制。这些研究 不仅将阐明早期的重要控制机制 发展，但也是翻译控制的重要一般机制 在所有真核细胞中。