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: 7421021
• 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 2010-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7421021
• 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（申请人提供）：细菌亮氨酸反应调节蛋白（Lrp）属于一类广泛分布于原核生物中的保守DNA结合蛋白，近年来一直是许多研究的焦点。大肠杆菌的 Lrp 是此类蛋白质中研究最充分的成员，它是一种相对丰富的核相关蛋白，作为全局调节因子，促进细菌适应各种环境压力和营养饥饿。更具体地说，Lrp 似乎在从指数增长期到稳定增长期的过渡过程中发挥着关键作用。此外，Lrp 样蛋白已被证明在毒力机制中发挥重要作用。在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制中的作用。我们的假设是，Lrp 样蛋白在从指数（复制）生长到持久（非复制）生长阶段（即休眠）的转变以及从持久阶段回到指数阶段（即休眠）的转变中发挥重要作用。 ，重新激活）在这种重要的病原体中。我们采用遗传方法，利用我们实验室开发的专门转导技术对上述基因进行敲除突变。将在不同的体外诱导休眠条件下研究这些突变对结核分枝杆菌生长、存活和耐药性的影响。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，并将使用小鼠再激活模型研究它们在再激活中的作用。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它们调节的毒力机制，将在相关条件下使用DNA微阵列和二维凝胶电泳对上述突变体及其同基因亲本进行研究。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，正如之前报道的大肠杆菌 Lrp 一样，使这项研究变得可行。此外，这些突变体中至少有一种在 SCID 小鼠中具有高毒力，并且似乎在全局基因调控方面存在缺陷，这表明它在结核分枝杆菌中充当全局调节因子。