Role of Lrp-like proteins in MTB Pathogenesis

Lrp 样蛋白在 MTB 发病机制中的作用

• 批准号: 7229564
• 负责人: MORAD HASSANI
• 金额: $ 12.77万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-05-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7229564
• 关键词: Anaerobiosis; Antimycobacterial Agents; Bacillus (bacterium); Back; Biology; Chromosomes; Class; Complement; Condition; DNA Microarray Chip; DNA Microarray format; DNA-Binding Proteins; Data; Defect; Doctor of Medicine; Doctor of Philosophy; Dose; Drug Delivery Systems; Drug Tolerance; Electrophoretic Mobility Shift Assay; Escherichia coli; Exposure to; Face; Family; Fluoroquinolones; Gene Expression Regulation; Gene Targeting; Genes; Genetic; Genome; Goals; Growth; Homologous Gene; Immune response; In Vitro; Infection; Investigation; Kinetics; Knock-out; Laboratories; Mediating; Modeling; Molecular Profiling; Mus; Mutation; Mycobacterium tuberculosis; Novobiocin; Nutrient; Organism; Parents; Pathogenesis; Pharmaceutical Preparations; Phase; Phenotype; Physical condensation; Play; Polymerase Chain Reaction; Predisposition; Prokaryotic Cells; Proteins; Proteomics; Pyrazinamide; Regulation; Reporting; Rifampin; Role; SCID Mice; Specificity; Starvation; Stress; Structure of parenchyma of lung; System; Technology; Testing; Time; Tuberculosis; Two-Dimensional Gel Electrophoresis; Virulence; base; expression vector; in vivo; inhibitor/antagonist; isoniazid; leucine-responsive regulatory protein; member; mouse model; mutant; pathogen; protein function; success

DESCRIPTION (provided by applicant): Bacterial leucine-responsive regulatory protein (Lrp) belongs to a class of conserved DNA-binding proteins widely distributed among prokaryotes and has been the focus of many investigations in recent years. The Lrp of Escherichia coli is the best studied member of this class of proteins, which is a relatively abundant nucleoid-associated protein that functions as a global regulator facilitating bacterial adaptation to a variety of environmental stresses and nutrient starvation. More specifically, Lrp appears to play a key role during transition from exponential to stationary phase of growth. In addition, Lrp-like proteins have been shown to play major roles in virulence mechanisms. Upon searching the Mycobacterium tuberculosis genome we found three Lrp homologs encoded by Rv2324, Rv2779 and Rv3291c. Our goal is to study the role of these Lrp-like proteins in the pathogenesis of M. tuberculosis. Our hypothesis is that Lrp-like proteins play important roles in the transition from exponential (replicative) growth to persistent (non-replicative) phase of growth (i.e., dormancy) and in the transition from the persistent phase back to the exponential phase (i.e., reactivation) in this important pathogen. We have used a genetic approach making knockout mutations in the above genes using specialized transduction technology developed in our laboratory. The effect of these mutations on growth, survival and drug tolerance of M. tuberculosis will be studied under different dormancy-inducing in vitro conditions. The role of the Lrp-like proteins of M. tuberculosis in establishing a persistent infection will be studied using the mouse model of infection and their role in reactivation will be studied using the murine models of reactivation. In addition, to determine the potential roles of these Lrp-like proteins in M. tuberculosis biology and to elucidate the virulence mechanisms regulated by them the above mutants and their isogenic parents will be studied using DNA microarrays and two-dimensional gel electrophoresis under relevant conditions. Preliminary results from our laboratory has indicated that the genes for all three Lrp-like proteins are non-essential in M. tuberculosis, as previously reported for E. coli Lrp, making this investigation feasible. In addition, at least one of these mutants is hypervirulent in SCID mice and appears to be defective ir global gene regulation suggesting that it functions as a global regulator in M. tuberculosis.

描述（由申请人提供）：细菌亮氨酸反应性调节蛋白（LRP）属于一类保守的DNA结合蛋白，在原核生物中广泛分布，并且近年来一直是许多研究的重点。大肠杆菌的LRP是该类别蛋白质的最佳研究成员，该蛋白是一种相对丰富的核苷相关蛋白，可作为全球调节剂，促进细菌适应各种环境应激和营养饥饿。更具体地说，LRP在从指数阶段到固定阶段的过渡期间似乎起着关键作用。此外，LRP样蛋白已被证明在毒力机制中起主要作用。搜索结核分枝杆菌基因组后，我们发现了由RV2324，RV2779和RV3291C编码的三个LRP同源物。我们的目标是研究这些LRP样蛋白在结核分枝杆菌发病机理中的作用。我们的假设是，类似于LRP的蛋白在从指数（复制）生长到持续性（非复制）生长阶段（即休眠）和从持续相位返回指数相（即重新激活）的过渡中起着重要作用。我们使用了在实验室中开发的专门转导技术，使用了一种在上述基因中进行基因敲除突变的遗传方法。这些突变对结核分枝杆菌的生长，存活和药物耐受性的影响将在不同的体外休眠诱导下进行研究。将使用小鼠感染模型研究结核分枝杆菌的LRP样蛋白在建立持续感染中的作用，并将使用鼠类重新激活模型研究其在重新激活中的作用。此外，为了确定这些LRP样蛋白在结核分枝杆菌生物学中的潜在作用，并阐明了由上述突变体调节的毒力机制及其同源性父母，将使用DNA微阵列和二维凝胶电泳在相关条件下进行研究。我们实验室的初步结果表明，所有三种LRP样蛋白的基因在结核分枝杆菌中都是非必需的，如先前报道的大肠杆菌LRP，这使得这项研究可行。此外，这些突变体中的至少一个在SCID小鼠中是高毒性的，并且似乎是IR全球基因调控，这表明它是结核分枝杆菌的全球调节剂。