Regulation of Bacillus subtilis tRNA synthetase genes

描述（由申请人提供）：枯草芽孢杆菌和相关生物体中参与氨基酸代谢的许多基因受T盒机制（一种新型转录终止控制系统）调控。该家族中每个基因的表达依赖于特定的不带电荷的tRNA与新生RNA转录物的结合，导致终止信号的通读，否则会减弱下游基因的表达。tRNA的识别需要tRNA反密码子与前导RNA中的单个密码子配对，以及tRNA的受体末端与竞争终止子螺旋形成的抗终止子元件配对;这些配对对于抗终止是必要的，但不是充分的。B的tRNA依赖性抗终止。枯草杆菌glyQS基因可以在纯化的系统中复制，并且tRNAGly与glyQS前导RNA的结合导致整个RNA的结构变化。T盒RNA直接和特异性识别其同源效应分子以调节基因表达的能力将这些RNA置于调节RNA的核糖开关家族中。下一个项目期将致力于详细分析tRNA依赖性抗终止的分子机制，以及T盒RNA对tRNA的特异性识别。我们将专注于RNA元素不同于已知的碱基配对区域，和glyQS模型RNA之间的功能差异，它代表了一个自然的缺失变体，和更常见的复杂的T盒RNA，含有额外的结构元件，其功能是未知的。我们还将研究在转录终止水平上操作的T盒元件（主要在厚壁菌门中发现）和在翻译起始水平上操作的T盒元件（主要在放线菌中发现，包括分枝杆菌属）之间的机制差异。T盒机制的进一步表征对于开发靶向该机制的新型抗微生物剂的长期目标至关重要，该机制在许多革兰氏阳性病原体中发现。近年来，RNA作为调控分子的作用越来越受到重视，T盒系统代表了RNA介导的基因调控的独特分子机制，以及tRNA在细胞中的新功能。公共卫生相关性：RNA介导的调节最近已成为所有生物体的核心参与者。本研究旨在分析T盒系统，这是一种独特的调节机制，其中RNA转录物直接感测特定的tRNA分子来控制基因表达。T盒机制被广泛用于革兰氏阳性细菌，包括重要的病原体，以调节多个必需基因，这项工作的长期目标是鉴定新的抗菌剂，抑制该系统，从而防止生长。

DESCRIPTION (provided by applicant): Many genes involved in amino acid metabolism in Bacillus subtilis and related organisms are regulated by the T box mechanism, a novel transcription termination control system. Expression of each gene in this family depends on binding of a specific uncharged tRNA to the nascent RNA transcript, resulting in readthrough of a termination signal that otherwise attenuates downstream gene expression. tRNA recognition requires pairing of the tRNA anticodon with a single codon, the "Specifier Sequence," in the leader RNA, as well as pairing of the acceptor end of the tRNA to an antiterminator element that competes with formation of the terminator helix; these pairing are necessary but not sufficient for antitermination. tRNAGly-dependent antitermination of the B. subtilis glyQS gene can be reproduced in a purified system, and binding of tRNAGly to glyQS leader RNA results in structural changes throughout the RNA. The ability of T box RNAs to directly and specifically recognize their cognate effector molecule to regulate gene expression places these RNAs in the riboswitch family of regulatory RNAs. The next project period will be directed toward detailed analysis of the molecular mechanism of tRNA-dependent antitermination, and specific tRNA recognition by T box RNAs. We will focus on RNA elements distinct from the known regions of base-pairing, and functional differences between the glyQS model RNA, which represents a natural deletion variant, and the more common complex T box RNAs that contain additional structural elements, the function of which is unknown. We will also investigate the mechanistic difference between T box elements that operate at the level of transcription termination (found primarily in Firmicutes), and those that operate at the level of translation initiation (found primarily in Actinomycetes, including Mycobacterium sp.). Further characterization of the T box mechanism is essential to the long-term goal of developing novel antimicrobial agents that target this mechanism, which is found in many Gram-positive pathogens. The role of RNA as a regulatory molecule has been increasingly appreciated in recent years, and the T box system represents a unique molecular mechanism for RNA-mediated gene regulation, and a new function for tRNA in the cell. PUBLIC HEALTH RELEVANCE: RNA-mediated regulation has recently emerged as a central player in all organisms. This study is directed toward analysis of the T box system, a unique regulatory mechanism in which RNA transcripts directly sense a specific tRNA molecule to control gene expression. The T box mechanism is widely used in Gram-positive bacteria, including important pathogens, to regulate multiple essential genes, and a long-term goal of this work is the identification of novel antimicrobial agents that inactivate this system, therefore preventing growth.