Biology and Mechanism of the Single Subunit RNA Polymerases

单亚基RNA聚合酶的生物学和机制

• 批准号: 7994857
• 负责人: RUI J. SOUSA
• 金额: $ 29.84万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-30 至 2012-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7994857
• 关键词: Address; Aging; Bacteriophages; Binding; Biological Models; Biology; Cell Nucleus; Complex; DNA; DNA Packaging; DNA-Directed RNA Polymerase; Diabetes Mellitus; Energy Metabolism; Engineering; Enzymes; Eukaryotic Cell; Genes; Genetic; Genetic Transcription; Glucose; Homologous Gene; Macromolecular Complexes; Mitochondria; Molecular; Nuclear; Oncogenes; Polymerase; Polymers; Positioning Attribute; Process; RNA; Reaction; Regulation; Relative (related person); Reporting; Repression; Respiration; Role; Solutions; Structure; System; T7 Lysozyme; T7 RNA polymerase; Transcription Factor 3; Transcription Process; Work; Yeasts; cancer cell; carbohydrate metabolism; human disease; inhibitor/antagonist; model development; polymerization; polypeptide; promoter; public health relevance; research study; single molecule; transcription factor

DESCRIPTION (provided by applicant): The single-polypeptide RNA polymerase of the T7 bacteriophage has been a model system for studying fundamental mechanisms of transcription ever since it was first identified almost 4 decades ago. Work over the past 15 years has revealed that the functions of this RNA polymerase extend beyond transcription of the phage genes, and that homologues of T7 RNA polymerase are widespread, occurring not only in eukaryotic mitochondria but also in mammalian nuclei, where they form the 4th major class of nuclear RNAPs (spRNAPIV). However, when compared to our detailed understanding of the structure and transcriptional mechanisms of T7RNAP, our understanding of the extra-transcriptional functions of this enzyme or of the mechanisms and biology of its mitochondrial and nuclear homologues is limited. To address these gaps in understanding we will: (1) Define the role of T7RNAP and its regulator T7 lysozyme in recruitment and assembly of the T7 DNA packaging machinery and, using both ensemble and single molecule experiments, describe the molecular details of the T7 DNA packaging reaction , (2) Determine crystal structures of yeast mitochondrial RNAP elongation and initiation complexes, and characterize the mechanism of promoter recognition by this RNA polymerase and of its activation by the mitochondrial transcription factor, (3) Identify the genes regulated by nuclear spRNAPIV and the effects of activation of respiration or of catabolite repression on spRNAPIV activity. These studies will advance our understanding of fundamental mechanisms of macromolecular complex assembly and of transcription processes in the mitochondrial and nuclear compartments of eukaryotic cells. PUBLIC HEALTH RELEVANCE RNA polymerases are the central players in the expression of genetic information. Aberrant activity of RNA polymerases leads to human disease. Our work will increase our understanding of how RNA polymerases control genes that are turned on in cancer cells and genes that are involved in cellular carbohydrate and energy metabolism, processes important in aging and diabetes.

描述（由申请人提供）：T7噬菌体的单多肽RNA聚合酶自近40年前首次鉴定以来一直是研究转录基本机制的模型系统。过去15年的工作表明，这种RNA聚合酶的功能超出了噬菌体基因的转录，并且T7 RNA聚合酶的同源物广泛存在，不仅存在于真核线粒体中，而且存在于哺乳动物细胞核中，在那里它们形成第四大类核RNAP（spRNAPIV）。然而，当与我们对T7RNAP的结构和转录机制的详细理解相比时，我们对这种酶的转录外功能或其线粒体和核同源物的机制和生物学的理解是有限的。为了解决这些认识上的差距，我们将：（1）确定T7 RNAP及其调节剂T7溶菌酶在T7 DNA包装机制的募集和组装中的作用，并使用整体和单分子实验描述T7 DNA包装反应的分子细节，（2）确定酵母线粒体RNAP延伸和起始复合物的晶体结构，（3）鉴定spRNAPIV核转录调控基因，以及呼吸激活和代谢抑制对spRNAPIV活性的影响。这些研究将推进我们对真核细胞线粒体和核区室中大分子复合物组装和转录过程的基本机制的理解。公共卫生相关RNA聚合酶是遗传信息表达的主要参与者。RNA聚合酶的异常活性导致人类疾病。我们的工作将增加我们对RNA聚合酶如何控制癌细胞中开启的基因以及参与细胞碳水化合物和能量代谢的基因的理解，这些基因在衰老和糖尿病中很重要。