Mechanisms of eukaryotic transcription activation

真核转录激活机制

• 批准号: 7279212
• 负责人: Steven M Hahn
• 金额: $ 47.55万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 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 2009-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7279212
• 关键词: Binding; Biochemical; Biochemical Genetics; Biological Assay; C-terminal; Chromatin; Class; Complex; Development; Disease; Disruption; End Point; Gene Expression Regulation; Genes; Genetic Transcription; Hydroxyl Radical; In Vitro; Individual; Lead; Length; Malignant Neoplasms; Maps; Methods; Modeling; Molecular; Molecular Genetics; Mutagenesis; Mutation; N-terminal; Pathway interactions; Peptides; Proteins; RNA Polymerase II; Ribosomal Proteins; SAGA; Signal Transduction; Site; Staging; Structure; Tertiary Protein Structure; Testing; Transcription Coactivator; Transcriptional Activation; Transcriptional Regulation; Work; base; crosslink; in vivo; promoter; protein protein interaction; transcription factor

DESCRIPTION (provided by applicant): Activation of transcription is the ultimate endpoint for many signal transaction and developmental pathways, and understanding the mechanism of activation is a key to understanding the action of these pathways. From previous studies, it is clear that disruption of normal gene regulation by mutations in gene-specific transcription activators can lead to cancer and other diseases. The broad long-term objectives of this proposal are to determine the mechanisms used by gene-specific transcription factors to activate transcription by RNA Polymerase II (Pol II). The proposed work will provide a basis for understanding gene regulation in normal and diseased states at the molecular level. The specific aims of this work will utilize biochemical, structural, and molecular genetic methods to examine the direct targets of several transcription activators and the mechanism whereby contact with these targets stimulates transcription. Using a newly developed method for mapping protein-protein contacts within large complexes, we will identify direct targets of activation domains in several model activators in the presence or absence of chromatin. Biochemical and molecular genetic studies will be conducted to test the relevance of these interactions in transcription. We will extend these studies to examine the targets of a different activator class acting at TATA-less promoters. After mapping these activator-target interactions by hydroxyl radical cleavage and direct protein-protein interaction assays, we will determine the structure of the activation domains in combination with their relevant targets. Finally, by blocking assembly of the Preinitiation Complex at intermediate stages, we will examine the mechanism by which activator contact with 1 of these targets, the SAGA coactivator complex, stimulates transcription. Our proposed work will illuminate important mechanisms and principles of transcriptional regulation.

描述（由申请人提供）：转录的激活是许多信号交易和发育途径的最终终点，了解激活的机理是理解这些途径的作用的关键。从以前的研究中可以明显看出，基因特异性转录激活剂中突变对正常基因调节的破坏会导致癌症和其他疾病。该提案的广泛长期目标是确定基因特异性转录因子用RNA聚合酶II（POL II）激活转录的机制。拟议的工作将为理解分子水平正常和患病状态的基因调节提供基础。这项工作的具体目的将利用生化，结构和分子遗传学方法来检查几个转录激活剂的直接靶标，以及与这些靶标接触的机制刺激转录。使用新开发的方法在大型复合物中绘制蛋白质 - 蛋白质接触，在存在或不存在染色质的情况下，我们将在几种模型激活剂中识别激活域的直接靶标。将进行生化和分子遗传研究，以测试这些相互作用在转录中的相关性。我们将扩展这些研究，以检查在无塔塔启动子上作用的不同激活剂类的靶标。通过羟基自由基裂解和直接蛋白质 - 蛋白质相互作用测定绘制这些激活剂 - 目标相互作用后，我们将确定激活域的结构与其相关靶标。最后，通过阻断中间阶段的预原始络合物的组装，我们将检查激活剂与其中1个靶标接触的机制，即传奇共振剂复合物，刺激转录。我们提出的工作将阐明重要机制和转录调节原则。