Molecular Basis for mRNA Decay in Bacteria

细菌中 mRNA 衰变的分子基础

• 批准号: 9893215
• 负责人: Alexander Serganov
• 金额: $ 5.74万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-21 至 2020-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9893215
• 关键词: Address; Adopted; Affect; Affinity; Bacteria; Binding; Biochemical; Bioinformatics; Biological Assay; Biological Process; Biology; Catalysis; Cell physiology; Cells; Crystallization; DNA Repair; Data; Dinucleoside Polyphosphates; Diphosphates; Enzymes; Escherichia coli; Excision; Foundations; Future; Gene Expression; Gene Expression Regulation; Genetic; Host Defense; In Vitro; Invaded; Mammalian Cell; Mediating; Messenger RNA; Metabolic; Metabolism; Methodology; Methods; Modeling; Molecular; Mutagenesis; Organism; Pathway interactions; Personal Satisfaction; Physiological; Positioning Attribute; Protein Biosynthesis; Proteins; RNA; RNA Binding; RNA Degradation; RNA triphosphatase; Reaction; Regulation; Research Proposals; Resistance; Role; Signal Transduction; Specificity; Stress; Structure; Substrate Specificity; System; Testing; Time; Transcript; Translating; Virulence; X-Ray Crystallography; base; cellular targeting; crosslink; decapping enzyme; design; diadenosine tetraphosphate; environmental change; enzyme activity; experimental study; genome-wide; in vitro activity; in vivo; inhibitor/antagonist; inorganic phosphate; insight; mRNA Decay; mRNA Transcript Degradation; mRNA decapping; member; novel; nudix hydrolase; pathogenic bacteria; protective effect; response; transcriptome sequencing; tripolyphosphate

Summary A central problem in biology is to understand how cells optimize biological processes in response to changing environmental conditions. mRNA decay mechanisms contribute to coordination of activities by limiting the number of times each mRNA can be translated into protein molecules. Recent studies have identified a novel 5'-end-dependent mRNA decay pathway in bacteria triggered by the initial conversion of the triphosphorylated 5' end of primary transcripts to a monophosphate by the Nudix hydrolase RppH. A triphosphate on the 5'-end of bacterial mRNA has a protective effect analogous to that of the cap structure of eukaryotic mRNAs; therefore, pyrophosphate removal by RppH parallels the action of eukaryotic decapping enzymes. Despite its vital importance, 5'-end-dependent mRNA decay is among the least understood mechanisms of gene regulation. This proposal details a set of specific aims that address specificity, catalytic mechanism, and the role of RppH and RppH-associated factors in regulating gene expression in bacteria. The proposal integrates X-ray crystallography, genetics, biochemical methods, focused genome-wide RNA sequencing and bioinformatics. Specific Aim 1 is devoted to the determination of the catalytic mechanism of RppH and understanding the substrate specificity of RppH. This Aim will address molecular principles of RNA `decapping' based on the crystal structures of Escherichia coli RppH bound to model RNAs and structure-guided mutagenesis. Specific Aim 2 will validate a new model for the 5'-end-dependent mRNA decay. This Aim will identify and characterize new factors involved in the pathway and their impact on gene expression landscape. Specific Aim 3 will identify cellular mRNA targets of RppH on a genome-wide scale under various environmental conditions. This Aim will reveal the molecular features of RNAs that are important for RppH recognition and uncover the contribution of RppH to the control of gene expression in bacteria. The results of these studies will provide a comprehensive picture of the critical steps in the 5'-end-dependent pathway for bacterial mRNA decay and thereby address a fundamental gap in our understanding of the regulatory networks controlled by RNA degradation.

概括 生物学的一个中心问题是了解细胞如何优化生物过程以响应变化 环境条件。 mRNA 衰变机制通过限制 每个 mRNA 可以翻译成蛋白质分子的次数。最近的研究发现了一种新颖的 细菌中由三磷酸化的初始转化触发的 5'-末端依赖性 mRNA 衰减途径 Nudix 水解酶 RppH 将初级转录本的 5' 末端转录为单磷酸。 5'端有一个三磷酸 细菌mRNA具有类似于真核mRNA帽结构的保护作用； 因此，RppH 去除焦磷酸盐与真核脱帽酶的作用是平行的。尽管其 至关重要的是，5'端依赖性 mRNA 衰变是人们最不了解的基因机制之一 规定。该提案详细说明了一系列具体目标，涉及特异性、催化机制和 RppH 和 RppH 相关因子在调节细菌基因表达中的作用。该提案整合了 X 射线晶体学、遗传学、生化方法、重点全基因组 RNA 测序和 生物信息学。具体目标 1 致力于确定 RppH 和 了解 RppH 的底物特异性。该目标将解决 RNA“脱帽”的分子原理 基于与模型 RNA 结合的大肠杆菌 RppH 的晶体结构和结构引导 诱变。具体目标 2 将验证 5' 端依赖性 mRNA 衰减的新模型。这个目标将 识别和表征该途径中涉及的新因素及其对基因表达景观的影响。 具体目标 3 将在各种条件下在全基因组范围内识别 RppH 的细胞 mRNA 靶标 环境条件。该目标将揭示对 RppH 重要的 RNA 分子特征 识别并揭示 RppH 对细菌基因表达控制的贡献。结果 这些研究将全面介绍 5' 端依赖性途径中的关键步骤 细菌 mRNA 衰减，从而解决我们对调控网络理解的根本差距 由RNA降解控制。