MECHANISM OF SELECTIVE EXPRESSION OF TRNA GENES

TRNA基因选择性表达机制

• 批准号: 3282020
• 负责人: KAREN U SPRAGUE
• 金额: $ 14.51万
• 依托单位: UNIVERSITY OF OREGON
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-12-01 至 1994-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3282020
• 关键词: Bombycidae; DNA directed RNA polymerase; DNA footprinting; alanine; binding proteins; cell differentiation; cell free system; developmental genetics; endocrine gland /system; eukaryote; gene expression; genetic manipulation; genetic transcription; genome; immunochemistry; microinjections; nucleic acid probes; nucleic acid sequence; radiotracer; transcription factor; transfer RNA

The goal of this work is to discover the mechanism of differential transcription in vitro of two kinds of tRNA genes. These genes encode alanine tRNA in the silkworm, Bombyx mori, and they differentially regulated in this organism. One class (tRNAAlaC genes) encodes alanine tRNA that is common to all silkworm cell types. The other class (tRNAAlaSG genes) encodes a distinct alanine tRNA that is found only the silkgland. The proposed study addresses the general problem of eukaryotic gene control, and the specific problem of transcription by RNA polymerase III. Since RNA polymerase III acts in all cells of all eukaryotes, understanding its function has broad biological and medical significance. The transcription properties displayed by tRNAAlaSG genes in vitro are very different from those displayed by tRNAAlaC genes -- and probably provide the basis for differential regulation in vivo. Therefore, a mechanistic analysis of these in vitro properties is biologically relevant. The interesting features of tRNAAlaSG transcription in vitro are 1.) It is extremely inefficient (relative to tRNAAlaC transcription) under standard conditions, and 2.) It is as efficient as tRNAAlaC transcription under special conditions. These two states could correspond to the inactivity of tRNAAlaSG genes in most silkworm cell types, and their high level of activity (equivalent to tRNAAlaC genes) in the silkgland. The molecular basis of both of these properties will be investigated. Specifically, the proposed experiments will identify the interaction(s) with the transcription machinery, and the step in the transduction cycle that is less efficient for tRNAAlaSG templates. When the defect in tRNAAlaSG transcription has been localized, the mechanism for overcoming it will be analyzed. To determine whether specific stimulation of tRNAAlaSG transcription in vitro is due to a qualitative or a quantitative change in the transcription machinery, the stimulatory component(s) will be resolved from the remainder of the transcription machinery, and tested for its specific activity on the two kinds of tRNAAla genes.

本工作的目的是揭示差异的机制， 两种tRNA基因的体外转录。 这些基因编码 家蚕丙氨酸tRNA的表达， 在这个有机体中。 一类（tRNAAlaC基因）编码丙氨酸 所有家蚕细胞类型共有的tRNA。 另一类（tRNAAlaSG 基因）编码一种独特的丙氨酸tRNA，这种tRNA只在丝腺中发现。 这项研究解决了真核基因的一般问题， 控制，以及RNA聚合酶III转录的具体问题。 由于RNA聚合酶III在所有真核生物的所有细胞中起作用， 其功能具有广泛的生物学和医学意义。 tRNAAlaSG基因在体外表现出的转录特性是非常重要的。 与tRNAAlaC基因所展示的不同， 体内差异调节的基础。 因此，一个机械的 这些体外性质的分析是生物学相关的。 的 tRNAAlaSG体外转录的有趣特征是1.）是 在标准条件下非常低效（相对于tRNAAlaC转录） 条件，2.）它与tRNAAlaC转录一样有效， 特殊条件。 这两种状态可能对应于 tRNAAlaSG基因在大多数家蚕细胞类型中的表达，以及它们的高表达水平， 活性（相当于tRNAAlaC基因）。 这两个属性的分子基础将被调查。 具体而言，拟议的实验将确定相互作用 与转录机制，以及转导周期中的步骤 这对于tRNAAlaSG模板来说效率较低。 当缺陷在 tRNAAlaSG转录已被定位，克服它的机制 将被分析。 为了确定是否特异性刺激tRNAAlaSG 体外转录是由于细胞质的质的或量的变化， 转录机制，刺激成分将被解析， 从其余的转录机器，并测试其 对两种tRNAAla基因的特异活性。