Transcriptome and proteome remodeling by Mycobacterium tuberculosis MazF toxins

项目摘要 结核分枝杆菌（Mtb）已经适应了广泛的攻击，从我们的免疫反应中幸存下来 到抗菌治疗-旨在根除有机体。然而，分子开关， 结核杆菌忍受这些压力，减缓复制或成为潜伏性结核病（TB）感染的休眠状态 不知道。关于压力生存的分子基础的新兴研究通常指向一个主要的 毒素-抗毒素（TA）系统的作用，TA系统是包含编码两种小蛋白的相邻基因的操纵子， 抑制TA蛋白-蛋白复合物中毒素活性的毒素及其同源抗毒素。但几 瓶颈阻碍了对这一挑衅性关联进行严格测试的进展。该提案征集 一个强大的多学科团队，在拟议工作的所有核心组成部分-RNA测序，TA 系统，结核病生物学/生理学和生物信息学/计算生物学。基因组尺度方法 PI实验室开发的5' RNA-seq将用于克服这些障碍，因为它们适用于 结核分枝杆菌迷宫（antitoxin）- MazF（toxin）家族由11个成员组成。5' RNA-seq将促进全面检测 在非应激条件下或暴露于应激后， 与潜伏性结核病感染有关。最后，将研究MazF毒素对Mtb蛋白质组的影响。 总的来说，这些方法将确定触发结核分枝杆菌毒素激活的环境信号， 在这些代谢状态下，MazF毒素靶向RNA的准确快照，并揭示了如何 毒素介导的RNA切割改变Mtb生理学。这些目标与“优先事项1：改善 结核病基础知识”是NIAID结核病研究战略计划的五个组成部分 2018年9月发布。

Project Summary Mycobacterium tuberculosis (Mtb) has adapted to survive a wide range of assaults—from our immune response to antimicrobial therapeutics—intended to eradicate the organism. However, the molecular switches that enable Mtb to endure these stresses, to slow replication or to become dormant as a latent tuberculosis (TB) infection are not known. Emerging studies on the molecular underpinnings of stress survival generally point to a major role for toxin-antitoxin (TA) systems, which are operons comprising adjacent genes encoding two small proteins, a toxin and its cognate antitoxin that inhibits toxin activity in the TA protein-protein complex. However, several bottlenecks have impeded progress toward rigorous testing of this provocative association. This proposal enlists a strong multidisciplinary team with expertise in all core components of the proposed work—RNA-seq, TA systems, Mtb biology/physiology and bioinformatics/computational biology. The genome-scale approach developed in the PI’s laboratory, 5’ RNA-seq, will be used to overcome these obstacles as they apply to the eleven-member MazE (antitoxin) – MazF (toxin) family in Mtb. 5’ RNA-seq will facilitate comprehensive detection of MazF targets in the Mtb transcriptome under unstressed conditions or after exposure to stresses that are relevant to latent TB infection. Finally, the impact of MazF toxins on the Mtb proteome will be investigated. Collectively, these approaches will identify the environmental signals that trigger toxin activation in Mtb, provide an accurate snapshot of RNAs targeted by MazF toxins under these metabolic states, and reveal clues to how toxin-mediated RNA cleavage alters Mtb physiology. These goals align well with “Priority 1: Improve Fundamental Knowledge of TB” of the five components of the NIAID Strategic Plan for Tuberculosis Research released in September 2018.