Transcription Factor IIIA

转录因子IIIA

• 批准号: 6519505
• 负责人: DAVID R SETZER
• 金额: $ 2.23万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-08-01 至 2002-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6519505
• 关键词: DNA binding protein; DNA directed RNA polymerase; DNA footprinting; Drosophilidae; RNA binding protein; RNA biosynthesis; Schizosaccharomyces pombe; Xenopus; chemical binding; developmental genetics; gel mobility shift assay; gene interaction; genetic transcription; genetically modified animals; polymerase chain reaction; protein protein interaction; protein structure function; ribosomal RNA; ribosomal proteins; site directed mutagenesis; species difference; thermodynamics; transcription factor; yeast two hybrid system; DNA binding protein; DNA directed RNA polymerase; DNA footprinting; Drosophilidae; RNA binding protein; RNA biosynthesis; Schizosaccharomyces pombe; Xenopus; chemical binding; developmental genetics; gel mobility shift assay; gene interaction; genetic transcription; genetically modified animals; polymerase chain reaction; protein protein interaction; protein structure function; ribosomal RNA; ribosomal proteins; site directed mutagenesis; species difference; thermodynamics; transcription factor; yeast two hybrid system

The proposed studies address specific hypotheses concerning the molecular and physical chemical mechanisms of sequence-specific DNA recognition by a transcriptional regulatory protein, transcription factor IIIA (TFIIIA). In addition, approaches for analyzing the interactions of TFIIIA with other proteins that form part of the transcription complex are proposed. As with other central cellular processes, such as protein synthesis and processing of pre-mRNAs, transcription occurs in the context of a large macromolecular complex that is assembled on a nucleic acid substrate containing specific signals which direct the production of a functional complex. Being able to describe the structures of the constituent components of these complexes, the processes (such as binding, dissociation, conformational change, and catalysis) leading to their assembly, function, and disassembly, and the relationships between structure and function will be necessary if a fundamental understanding of these central cellular functions is to be achieved. The proposed studies are motivated by this overall goal. TFIIIA is responsible for nucleating the formation of a large macromolecular complex on 5S rRNA genes that is responsible for synthesis of 5S rRNA. Recent structural studies have suggested a novel mechanism of sequence- specific DNA binding involving a dynamic DNA-protein interface in the TFIIIA-5S rRNA gene complex. The validity of this model will be tested by experiments proposed in this application. Previous studies have also suggested the existence of a large energetic strain in the TFIIIA-5S rRNA gene complex, resulting in a "compensatory" mode of binding, and specific experimental tests of a mechanism for producing such energetic strain are proposed. A powerful genetic assay for analyzing the Xenopus TFIIIA-5S rRNA gene interaction will be used to identify mutant forms of TFIIIA that specifically recognize variant 5S rRNA genes, and these will be used both to better understand the specific contacts that occur between the DNA and the protein in the TFIIIA-5S rRNA gene complex as well as reagents for analyzing the function of mutant forms of TFIIIA in vivo. Finally, methods and reagents for analyzing the interaction of TFIIIA-5S rRNA gene complexes with other components of the transcriptional machinery will be developed and used to study these higher order interactions.

拟议的研究介绍了有关通过转录调节蛋白（转录因子IIIA（TFIIIA））识别序列特异性DNA识别分子和物理化学机制的特定假设。此外，提出了分析TFIIIA与其他构成转录复合物一部分的蛋白质相互作用的方法。与其他中央细胞过程一样，例如蛋白质的合成和前MRNA的加工，转录发生在大型大分子复合物的背景下，该复合物组装在核酸基板上，该核酸底物包含指导特定信号的特定信号，该信号指导了功能复合物的产生。能够描述这些复合物的组成成分的结构，如果要实现对这些中心细胞功能的基本理解，则过程（例如结合，分离，构象变化和催化），导致它们的组装，功能和拆卸，以及结构和功能之间的关系。拟议的研究是由这一总体目标激励的。 TFIIIA负责在5S rRNA基因上形成大型大分子复合物，该基因负责5S rRNA的合成。最近的结构研究提出了一种新型的序列特异性DNA结合机制，涉及TFIIIA-5S rRNA基因复合物中的动态DNA-蛋白界面。该模型的有效性将通过本应用程序中提出的实验测试。先前的研究还表明，在TFIIIA-5S rRNA基因复合物中存在大量的能量菌株，从而导致了“补偿性”结合方式，并提出了用于产生这种能量菌株的机制的特定实验测试。用于分析爪蟾TFIIIA-5S rRNA基因相互作用的强大遗传测定将用于识别特定识别变体5S rRNA基因的TFIIIA的突变形式，这些形式将用于更好地理解TFIIIIA-5S RRNA基因中DNA和蛋白质之间发生的特定接触，以及分析的综合体以及分析的综合群体的综合群体。体内Tfiiia。最后，将开发并使用用于分析TFIIIA-5S rRNA基因复合物与转录机械其他组件的相互作用的方法和试剂。