Mechanism and targets of Sen1-dependent RNA polymerase II termination

Sen1依赖性RNA聚合酶II终止的机制和靶标

• 批准号: 8429463
• 负责人: DAVID A BROW
• 金额: $ 26.39万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-15 至 2015-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8429463
• 关键词: Active Sites; Alleles; Amino Acids; Amyotrophic Lateral Sclerosis; Ataxia; Bacteria; Base Pairing; Biochemical; Biological Assay; Biological Models; Cellular biology; Code; Collection; Data Set; Defect; Disease; Enzymes; Eukaryota; Foundations; Functional RNA; Fungal Genome; Gene Expression; Gene Expression Profile; Gene Expression Regulation; Gene Targeting; Genes; Genetic; Genetic Suppression; Genetic Transcription; Genome; Goals; Guanosine Triphosphate; Health; Homeostasis; Homologous Gene; Human; Human Biology; Investigation; Lead; Maps; Modeling; Molecular; Molecular Conformation; Muscle Weakness; Mutation; Nerve Degeneration; Neurons; Pathway interactions; Polyadenylation; Polymerase; Process; Protein Analysis; Proteins; Publishing; Quantitative Microscopy; RNA; RNA Polymerase II; RNA-Binding Proteins; Reading; Regulation; Regulator Genes; Saccharomyces cerevisiae; Signal Transduction; Site; Structure; System; Teenagers; Testing; Transcript; Transcription Initiation Site; Transcriptional Regulation; Work; Yeasts; attenuation; base; disabling disease; genetic selection; helicase; human senataxin protein; insight; mathematical model; motor neuron degeneration; mutant; novel; protein function; public health relevance; termination factor; transcription termination; transcriptome sequencing

DESCRIPTION (provided by applicant): The realization that non-coding transcripts are remarkably abundant and participate in the regulation of protein- coding genes has revolutionized the study of eukaryotic gene expression. Yet little is known about how non- coding RNAs are made and function. We discovered an RNA polymerase II (Pol II) termination pathway that influences the synthesis and function many non-coding RNAs. This pathway requires the conserved helicase Sen1 and a collection of RNA-binding proteins that recognize the termination signal in the nascent transcript. The Sen1 pathway is essential in the model eukaryote S. cerevisiae (brewer's yeast), and mutations in the human Sen1 gene (SETX) cause the degeneration of motor neurons, leading to forms of ataxia and amyotrophic lateral sclerosis (ALS). Remarkably, Sen1-dependent termination regulates some protein-coding genes by transcription attenuation, which was previously thought to be restricted to bacteria. We will determine the molecular mechanism of Sen1-dependent termination using both genetic and biochemical approaches. Our structure/function studies will focus on two key proteins in the pathway, Sen1 and Pol II, in which we have isolated mutations that result in terminator read-through. Regulation of the IMD2 gene will be examined in detail as a paradigm for NTP homeostasis by Sen1-dependent attenuation and alternative start site selection. We will also identify and characterize new examples of Sen1-dependent gene regulation. The cis- and trans-acting determinants for regulation of these genes will be defined using genetic selections, quantitative gene expression assays, and mathematical modeling. The information gained in these studies will serve as a foundation for the investigation of RNA-based transcriptional regulation in other eukaryotes, including humans. Given the emerging evidence for the importance of non-coding transcripts in gene regulation in humans, our results will have broad implications for eukaryotic biology and human health, and will lead to a better understanding of the causes of motor neuron degeneration.

描述（由申请人提供）：非编码转录物非常丰富并且参与蛋白质编码基因的调节的认识已经彻底改变了真核基因表达的研究。然而，人们对非编码 RNA 的制造和功能知之甚少。我们发现了一种 RNA 聚合酶 II (Pol II) 终止途径，它影响许多非编码 RNA 的合成和功能。该途径需要保守的解旋酶 Sen1 和一组识别新生转录物中终止信号的 RNA 结合蛋白。 Sen1 通路在模型真核生物酿酒酵母（啤酒酵母）中至关重要，人类 Sen1 基因 (SETX) 的突变会导致运动神经元退化，从而导致共济失调和肌萎缩侧索硬化症 (ALS)。值得注意的是，Sen1 依赖性终止通过转录减弱来调节一些蛋白质编码基因，此前人们认为这种现象仅限于细菌。我们将使用遗传和生化方法确定 Sen1 依赖性终止的分子机制。我们的结构/功能研究将重点关注该途径中的两个关键蛋白 Sen1 和 Pol II，我们在其中分离出导致终止子通读的突变。 IMD2 基因的调节将作为 NTP 稳态的范例通过 Sen1 依赖性衰减和替代起始位点选择进行详细检查。我们还将鉴定并表征 Sen1 依赖性基因调控的新例子。将使用遗传选择、定量基因表达测定和数学模型来定义调节这些基因的顺式和反式作用决定因素。这些研究中获得的信息将作为研究包括人类在内的其他真核生物中基于 RNA 的转录调控的基础。鉴于非编码转录本在人类基因调控中重要性的新证据，我们的结果将对真核生物学和人类健康产生广泛影响，并将导致更好地了解运动神经元变性的原因。