MECHANISMS OF TRANSCRIPTIONAL REGULATION IN EUKARYOTES

真核生物转录调控机制

• 批准号: 2180745
• 负责人: James T. Kadonaga
• 金额: $ 19.88万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-12-01 至 1997-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2180745
• 关键词: DNA directed RNA polymerase; Drosophilidae; HeLa cells; developmental genetics; embryology; eukaryote; gel mobility shift assay; gene expression; genetic promoter element; genetic regulatory element; genetic transcription; laboratory rabbit; nucleic acid sequence; transcription factor

The objective of the proposed study is to elucidate the mechanisms of basal and regulated transcription by RNA polymerase II. At present, most of the basal transcription factors have been purified and clone, and thus, the next several years should be a productive period for the analysis of the mechanism of the reconstituted with purified, recombinant TFII, TBP, and TFIIF30 along with purified RNA polymerase II from Drosophila, calf thymus, or HeLa cells. This minimal transcription system appears to constitute the central catalytic core of the RNA polymerase II transcriptional machinery. Beginning with this purified, minimal system, it will be possible to examine, in greater detail, the roles of TFIIB, TBP, and TFIIF30 as well as to identify the function of other basal transcription factors such as TFIIA, TFIIE, TFIIH, and TFIIJ. In addition, recent findings in our lab and others indicate that there are variations in the mechanism of basal examination of both the cis-acting elements in the promoters as well as the protein factors that are responsible for the promoter-specific effects that have been observed. These studies should lead to a better understanding of the fundamental transcription process and should eventually be applied to the molecular analysis of diseases involving the regulation of gene expression, such as AIDS and some forms of cancer. Identification and characterization of factors that are required for basal transcription. We have found that TFIIB, TBP, TFIIF30, and polymerase can accurately transcribe only a subset of promoters. It is thus, important to identify the factors that are required for transcription of the remaining genes. Examination of the molecular basis for distinct transcriptional properties of different promoters. We have been able to classify promoters into different functional categories based on their behaviour in various assays. The aim of these studies is to identify and to characterize the cis-acting sequences and the protein factors that are responsible for the promoter-specific effects. Kinetics and mechanism of transcription with a purified transcription system. We will carry out a systematic kinetic analysis of the assembly of the initiation complex with purified components. This work should provide new insights into the pathway leading to the assembly of a productive initiation complex. Analysis of transcriptional activation with the purified transcription system. A primary goal of these experiments is to determine the minimal components that are required to observe activation of transcription by promoter-and enhancer-binding factors.

拟议研究的目的是阐明 RNA聚合酶II的基础和调节转录。 目前，大多数 基础转录因子已被纯化和克隆，并且 因此，接下来的几年应该是 分析用纯化的重组重组重构的机制 TFII，TBP和TFIIF30以及纯化的RNA聚合酶II 果蝇，小牛胸腺或HeLa细胞。这个最小的转录系统 似乎构成了RNA聚合酶的中心催化核心 II转录机械。 从这个纯化的，最小的开始 系统，可以更详细地检查 TFIIB，TBP和TFIIF30，以及识别其他的功能 基础转录因子，例如TFIIA，TFIIE，TFIIH和TFIIJ。 在 此外，我们实验室的最新发现表明 顺式作用的基础检查机制的变化 启动子中的元素以及蛋白质因子 负责已观察到的启动子特异性效应。 这些研究应该可以更好地了解基本 转录过程，最终应应用于分子 分析涉及基因表达调节的疾病 作为艾滋病和某些形式的癌症。 识别和表征对因素所需的因素 基础转录。 我们发现TFIIB，TBP，TFIIF30和 聚合酶只能准确转录启动子的子集。 这是 因此，重要的是要确定 其余基因的转录。 检查分子基础的不同转录 不同启动子的特性。 我们已经能够分类 根据其行为，促进者分为不同的功能类别 在各种测定中。 这些研究的目的是识别和 表征顺式作用序列和蛋白质因子 负责启动子特异性效应。 纯化转录的动力学和转录机理 系统。 我们将对组件进行系统的动力学分析 带有纯化成分的启动复合物。 这项工作应该 向通道的通道提供新的见解 生产性启动综合体。 用纯化的转录分析转录激活 系统。 这些实验的主要目标是确定最小 通过观察转录激活所需的组件 启动子和增强子结合因素。