Initiation of mRNA decay in Bacillus subtilis

枯草芽孢杆菌中 mRNA 降解的启动

• 批准号: 7921228
• 负责人: DAVID H BECHHOFER
• 金额: $ 3.67万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7921228
• 关键词: 5' -exoribonuclease; Address; Affect; Antibiotics; Bacillus subtilis; Bacteria; Binding; Binding Proteins; Characteristics; Elements; Endoribonucleases; Environment; Escherichia coli; Gene Expression; Gene Expression Profile; Genetic Transcription; Goals; Gram-Positive Bacteria; Half-Life; In Vitro; Individual; Kinetics; Laboratories; Laboratory Research; Lead; Messenger RNA; Modeling; Names; Organism; Phosphodiesterase I; Plasmids; Play; Polyribonucleotide Nucleotidyltransferase; Process; Proteins; RNA; RNA Decay; RNA chemical synthesis; Regulation; Ribonucleases; Role; Site; Small RNA; Structure; Study models; System; Translations; Work; design; endonuclease; endoribonuclease; human tissue; in vitro activity; mutant; overexpression; prevent; research study; ribonuclease E; ribonuclease PH; ribonuclease R; spleen exonuclease; stem

DESCRIPTION (provided by applicant): Project Summary: Our laboratory seeks to understand the control of messenger RNA decay in the gram-positive bacterium, Bacillus subtilis. While much is known about the mechanism and regulation of mRNA synthesis (transcription) and translation of mRNA into protein, little is known about the intermediate step in gene expression-degradation of mRNA. Experiments in this proposal focus on 2 aspects of mRNA decay: the elements (e.g., sequences, structures) of an mRNA that determine its half-life, and the ribonuclease activities that are required for initiation and completion of mRNA decay. The availability of B. subtilis strains that are deficient in 1 or more 3'-to-5' exoribonucleases will make it possible to examine the role of individual ribonucleases in mRNA turnover. The decay of 3 small mRNAs, whose characteristics have been studied to some extent, will be analyzed in detail and will serve as models for the study of mRNA decay generally. Experiments are proposed to clarify: 1) what is the initiation site for decay? 2) which ribonuclease(s) participates in initiation of decay? 3) how does the decay mechanism deal with stable secondary structure? and 4) how do the various 3'-to-5' exoribonucleases bind and degrade mRNA? The role of the 4 known B. subtilis 3'-to-5' exoribonucleases -- PNPase, RNase R, RNase PH, and YhaM-will be assessed in the turnover of model mRNAs, as well as newly-identified mRNAs that are stabilized in a PNPase-deficient mutant strain. The likely participation of 2 B. subtilis endoribonucleases -- RNase J1 and RNase J2 -in mRNA decay will be assessed. An in vitro system will be established that will be useful in probing the characteristics of purified ribonucleases, which will be overexpressed and isolated from E. coli. Relevance: Messenger RNA (mRNA) is the template molecule upon which proteins are synthesized. Bacteria rely on rapid mRNA decay to adapt to changing environments, and the details of this process will be studied in detail in the model microorgansim, Bacillus subtilis. Elucidating the mechanism of mRNA in this bacterium could lead to the design of new antibiotics that inhibit the mRNA decay process and thereby prevent successful bacterial colonization of human tissues.

描述（由申请人提供）：项目摘要：我们的实验室旨在了解革兰氏阳性细菌枯草芽孢杆菌中对信使RNA衰变的控制。尽管对mRNA合成（转录）和将mRNA转化为蛋白质的机理和调节知之甚少，但对mRNA基因表达降解的中间步骤知之甚少。该提案中的实验集中于mRNA衰变的两个方面：确定其半衰期的mRNA的元素（例如序列，结构），以及启动和完成mRNA衰变所需的核糖核酸酶活性。枯草芽孢杆菌菌株的可用性在1或更多3'-to-5'驱虫核中缺乏，将使检查单个核糖珠在mRNA周转率中的作用。将在某种程度上研究其特征的3个小mRNA的衰减将进行详细分析，并将作为研究mRNA衰变的模型。提出了实验来澄清的：1）衰减的启动位点是什么？ 2）哪些核糖核酸酶参与衰减的开始？ 3）衰减机制如何处理稳定的二级结构？ 4）各种3'to-5'驱量如何结合和降解mRNA？在模型mRNA的周转中评估了4种已知的枯草芽孢杆菌3'-to-5'To-5'驱虫核酸酶-PNPase，RNase R，RNase PH和Yham-Will，以及在PNPase缺乏互缺质菌株中稳定的新识别的mRNA。将评估2 B.枯草酵母内切核酸酶-RNase J1和RNase J2 -In mRNA衰变的可能参与。将建立一个体外系统，该系统将有助于探测纯化的核糖核酸的特征，该核果酶将过表达并与大肠杆菌分离。相关性：信使RNA（mRNA）是合成蛋白质的模板分子。细菌依靠快速mRNA衰变来适应不断变化的环境，该过程的细节将在模型的微牙杆菌，枯草芽孢杆菌中详细研究。阐明该细菌中mRNA的机制可能导致设计新抗生素，从而抑制mRNA衰变过程，从而防止成功的细菌定植。