Protein interactions and initiation by RNA polymerase I

蛋白质相互作用和 RNA 聚合酶 I 引发

• 批准号: 7112944
• 负责人: LAWRENCE I ROTHBLUM
• 金额: $ 22.25万
• 依托单位: GEISINGER CLINIC
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-01 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7112944
• 关键词: 3T3 cells; DNA directed RNA polymerase; DNA replication origin; SDS polyacrylamide gel electrophoresis; carcinogenesis; cell growth regulation; cell line; cytogenetics; enzyme induction /repression; gene expression; genetic transcription; high performance liquid chromatography; immunoprecipitation; liquid chromatography mass spectrometry; matrix assisted laser desorption ionization; phosphorylation; protein protein interaction; ribosomal DNA; ribosomal RNA; small interfering RNA; thin layer chromatography; transcription factor; western blottings

DESCRIPTION (provided by applicant): Understanding the mechanism of transcription of the genes that code for 45S pre-ribosomal RNA is essential if we are to understand both normal and abnormal growth processes, e.g. wound healing and neoplasia. Ribosome biogenesis, and therefore the expression of the ribosomal RNA genes, is coordinated with the rate of cell growth, and responds to a variety of signals, depending upon the cell type studied. The long-term objective of our research is to determine the mechanism(s) by which ribosomal RNA gene (rDNA) transcription is regulated. At least two trans-acting factors are required for accurate and efficient rDNA transcription, SL-1, and UBF. SL-1 is required for transcription while UBF activates transcription. Recent studies demonstrate that there are several ways in which rDNA transcription is regulated. One is the regulation of the ability of RNA polymerase I to initiate transcription. It has not been clear how many factors control the ability of pol I to initiate transcription. Two of the factors, Rrn3/TIF-IA and TFIC were believed to be the same factor. We demonstrated that they are two different factors. The yeast Rrn3 gene is essential for cell viability, and experiments in mammalian cells demonstrate that mammalian Rrn3 is essential for ribosomal gene transcription. However, there is considerable controversy concerning the role of Rrn3 in transcription, e.g. is it required for the recruitment of pol I to the rDNA promoter? Moreover, our data demonstrate fundamental differences between the mechanisms that regulate Rrn3 function in yeast and mammalian cells. Two of our goals focus on the determination of the role of Rrn3 in rDNA transcription and the mechanism(s) that regulate Rrn3 activity. Our third aim focuses on the role of PAF53, a second polymerase associated factor. Several lines of evidence suggest that PAF53 is an important component of the apparatus that regulates rDNA transcription. Antibodies to PAF53 block rDNA transcription, and the association of RNA polymerase I with PAF53 correlates with the rate of rDNA transcription. For example, PAF53 levels, but not core RNA polymerase I levels are reduced when NIH 3T3 cells are serum starved, while serum starvation causes the dissociation of PAF53 from RNA polymerase I in 3T6 cells. However, the role of PAF53 in rDNA transcription is yet to be defined.

描述（由申请人提供）：了解45S前核糖体RNA的基因转录机理，如果我们要了解正常生长过程和异常生长过程，例如伤口愈合和肿瘤。核糖体生物发生，因此，核糖体RNA基因的表达与细胞生长速率协调，并根据所研究的细胞类型响应多种信号。我们研究的长期目标是确定调节核糖体RNA基因（rDNA）转录的机制。对于准确有效的RDNA转录SL-1和UBF需要至少两个跨作用因子。在UBF激活转录时，SL-1是转录所需的。最近的研究表明，有几种调节rDNA转录的方法。一种是对RNA聚合酶I启动转录的能力的调节。目前尚不清楚有多少因素控制着pol I启动转录的能力。其中两个因素，RRN3/TIF-IA和TFIC是相同的因素。我们证明它们是两个不同的因素。酵母RRN3基因对于细胞活力至关重要，哺乳动物细胞的实验表明哺乳动物RRN3对于核糖体基因转录至关重要。但是，关于RRN3在转录中的作用，例如将pol I招募到rDNA启动子需要吗？此外，我们的数据证明了调节酵母和哺乳动物细胞中RRN3功能的机制之间的基本差异。我们的两个目标集中在确定RRN3在rDNA转录中的作用和调节RRN3活性的机制。我们的第三个目标重点是PAF53（第二个聚合酶相关的因素）的作用。几条证据表明，PAF53是调节rDNA转录的设备的重要组成部分。 PAF53块rDNA转录的抗体，RNA聚合酶I与PAF53的缔合与rDNA转录速率相关。例如，当NIH 3T3细胞饥饿时，PAF53水平，而不是核心RNA聚合酶I水平降低，而血清饥饿会导致PAF53在3T6细胞中从RNA聚合酶I中解离。但是，PAF53在rDNA转录中的作用尚未定义。