GENETIC EXPRESSION/DEVELOPMENT/CONTROL/MRNA UTILIZATION

遗传表达/发育/控制/mRNA利用

• 批准号: 3279744
• 负责人: ANTHONY INFANTE
• 金额: $ 11.37万
• 依托单位: WESLEYAN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-02-01 至 1987-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3279744
• 关键词: Escherichia coli; animal population genetics; cell differentiation; developmental genetics; early embryonic stage; gene expression; genetic manipulation; genetic registry /resource /referral center; genetic translation; messenger RNA; molecular cloning; nonmammalian vertebrate embryology; nucleic acid sequence; polysomes; saltwater environment

The objective of this research is to gain further understanding of the molecular mechanisms underlying development and differentiation. The fundamental theme will be the elucidation of the means operating in early embryogenesis for regulating the use of genetic information. Sea urchins will be used as a model embryonic system. Templates for protein synthesis present in the unfertilized egg are located primarily in postribosomal ribonucleoprotein complexes called free mRNP or informosomes, and a large portion of the mRNA synthesized by developing embryos enters the free RNP pool. Because of their cellular location it appears that the translation of these templates is controlled in the cytoplasm. We will systematically characterize both qualitatively and quantitatively specific species of mRNA present in the free RNPs in order to better understand the function of free RNPs and how the expression of the genetic information they contain is controlled. Our main approach will be to compare various properties of the free RNP mRNA to the mRNA actively engaged in protein synthesis, which is present in polysomes. A recombinant DNA library of expressed sequences corresponding to the low complexity-high abundance class of sea urchin poly(A) plus mRNA cloned in E.coli will be employed. We will select from this cDNA library a number of interesting cloned sequences for analysis. This selection involves appropriate DNA-RNA hybridization analyses and hybrid-arrest-translation experiments. The mRNAS isolated from embryos at various stages of development, will be characterized in terms of: concentration, cellular location (polysomes/free RNP distribution), time of synthesis in development, rate of synthesis, turnover kinetics (half-life), and efficiency to be translated in vitro. From these data it will be possible to gain considerable information concerning the metabolism and utilization of a number of specific messenger RNAs in a rapidly developing system and thereby the nature of translational control mechanisms operating in early development.

本研究的目的是为了进一步了解 发育和分化的分子机制。 这 基本主题将是阐明早期运作的手段 胚胎发生用于调节遗传信息的使用。 海胆 将被用作模型胚胎系统。 蛋白质合成模板 存在于未受精卵中的主要位于核糖体后 称为游离 mRNP 或信息体的核糖核蛋白复合物，以及一个大的 发育中胚胎合成的部分 mRNA 进入游离 RNP 水池。 由于它们的细胞位置，似乎翻译 这些模板的控制在细胞质中。 我们将系统地 定性和定量表征特定种类的 mRNA 存在于自由 RNP 中，以便更好地理解自由的功能 RNP 及其所含遗传信息的表达方式 受控。 我们的主要方法是比较 将游离的 RNP mRNA 转移到积极参与蛋白质合成的 mRNA 上，即 存在于多核糖体中。 表达序列的重组 DNA 文库 对应于海胆的低复杂性-高丰度类别 将使用在大肠杆菌中克隆的poly(A) plus mRNA。 我们将从中选择 该 cDNA 文库中有许多有趣的克隆序列可供分析。 这种选择涉及适当的 DNA-RNA 杂交分析和 混合逮捕翻译实验。 从胚胎中分离的 mRNAS 不同的发展阶段，其特点如下： 浓度、细胞位置（多核糖体/游离 RNP 分布）、时间 开发中的合成、合成速率、周转动力学（半衰期）、 和体外转化的效率。 根据这些数据将是 可以获得有关新陈代谢的大量信息 在快速发展的过程中利用许多特定的信使RNA 系统以及由此运行的平移控制机制的性质 在早期发展中。