Functional analysis of Mycobacterium tuberculosis VapC toxins

结核分枝杆菌VapC毒素的功能分析

基本信息

批准号：
8719363
负责人：
Melvili Cintron
金额：
$ 3.19万
依托单位：
RBHS-ROBERT WOOD JOHNSON MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-08-01 至 2017-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8719363
关键词：
Address Amino Acid Sequence Antibiotics Anticodon Antitoxins Bacteria Bacterial Toxins Biochemical Biochemical Genetics Cells Characteristics Cleaved cell Complex Consensus Sequence Coupled Data Employee Strikes Environment Enzymes Escherichia coli Family Family member Fellowship Gene Expression Profile Genome Growth Human Immune response Infection Ions Laboratories Learning Literature Messenger RNA Methods Molecular Mono-S Mycobacterium tuberculosis N-terminal Organism Peptide Sequence Determination Phenotype Physiological Physiology Process Property Protein Family RNA RNA Sequences Refractory Regulation Relative (related person)Ribonucleases Ribosomal RNA Role Serine Site Specificity Structure System Techniques Testing Therapeutic Threonine Time Toxin Training Transfer RNA Translations Tuberculosis Virulence Work antimicrobial assault base cell growth cellular targeting inorganic phosphate latent infection member novel public health relevance research study stem transcriptome sequencing

项目摘要

DESCRIPTION (provided by applicant): Mycobacterium tuberculosis (Mtb) has adapted to survive a wide range of assaults-from our immune response to antimicrobial therapeutics-intended to eradicate the organism. This unique ability to persist for long periods of time in its human host as a latent tuberculosis infection dominates the infection landscape relative to active tuberculosis cases. Despite ongoing efforts to understand latent tuberculosis infection, the molecular switches that enable Mtb to slow or stop replication and become dormant are poorly understood. However, the convergence of studies on bacterial toxin-antitoxin (TA) systems/modules demonstrating their general growth-regulating properties, coupled with the identification of a striking abundance of TA systems in the Mtb genome, suggest that their expression may facilitate the non-replicating persistent state of Mtb during latent tuberculosis infection. This proposal seeks to fill the gap in our understanding of the molecular basis of laten tuberculosis infection by investigating the structure and function of the large, uncharacterized family of Mtb VapC toxins through detailed study of three family members. The studies in this fellowship proposal integrate into the broader focus of the Woychik laboratory and address our hypothesis that the VapC toxin family in Mtb represents a new class of enzymes that recognize both RNA sequence and tertiary structure to specifically cleave tRNA and/or rRNA(s) and inactivate the translation machinery. The studies proposed in Aim 1 enlist a specialized genome scale method, 5' RNA-seq, to identify the RNA targets for three Mtb VapC toxins using an Escherichia coli host, those in Aim 2 validate the RNA targets and cleavage sites identified by 5' RNA-seq.

描述（由申请人提供）：结核分枝杆菌（Mtb）已经适应了多种攻击——从我们的免疫反应到抗菌治疗——旨在根除该生物体。与活动性结核病病例相比，这种作为潜伏性结核感染在人类宿主中长期存在的独特能力主导了感染情况。尽管人们不断努力了解潜伏性结核感染，但人们对结核分枝杆菌减缓或停止复制并进入休眠状态的分子开关知之甚少。然而，对细菌毒素-抗毒素（TA）系统/模块的研究的融合证明了它们的一般生长调节特性，再加上在 Mtb 基因组中鉴定出惊人丰富的 TA 系统，表明它们的表达可能促进非复制潜伏性结核感染期间 Mtb 的持续状态。该提案旨在通过对三个家族成员的详细研究，研究 Mtb VapC 毒素大家族的结构和功能，从而填补我们对潜伏性结核感染分子基础的理解空白。本奖学金提案中的研究融入了 Woychik 实验室更广泛的关注点，并解决了我们的假设，即 Mtb 中的 VapC 毒素家族代表了一类新的酶，可识别 RNA 序列和三级结构，以特异性切割 tRNA 和/或 rRNA（s ）并停用翻译机制。目标 1 中提出的研究采用了一种专门的基因组规模方法 5' RNA-seq，以使用大肠杆菌宿主识别三种 Mtb VapC 毒素的 RNA 靶标，目标 2 中的研究验证了 5' RNA 识别的 RNA 靶标和切割位点RNA测序。