Control of GTP Homeostasis by (p)ppGpp

(p)ppGpp 对 GTP 稳态的控制

• 批准号: 8722778
• 负责人: Jue D. Wang
• 金额: $ 29.35万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8722778
• 关键词: Amino Acids; Anabolism; Antibiotic Resistance; Antibiotic Therapy; Antibiotics; Bacillus anthracis; Bacillus subtilis; Bacteria; Binding; Binding Sites; Biochemistry; Biological Assay; Cell Survival; Cells; DNA-Directed RNA Polymerase; Data; Development; Drug Tolerance; Enterococcus; Enterococcus faecalis; Environment; Enzymes; Escherichia coli; Feedback; Foundations; Funding; Genetic; Genetic Screening; Genetic Transcription; Genomics; Genus staphylococcus; Glycolysis; Gram-Negative Bacteria; Gram-Positive Bacteria; Growth; Guanosine Triphosphate; Homeostasis; Homologous Gene; Human; In Vitro; Kinetics; Lead; Life; Ligase; Link; Listeria; Mediating; Metabolic; Metabolism; Microbe; Modeling; Molecular; Mutation; Nucleotides; Pathway interactions; Physiological; Purines; Pyruvate Kinase; Regulation; Resistance; Role; Signal Transduction; Starvation; Stress; Structure; Testing; Toxic effect; Work; antimicrobial; bactericide; environmental change; enzyme activity; genetic analysis; glucose metabolism; insight; killings; loss of function mutation; metabolomics; mutant; novel; pathogen; prevent; public health relevance; purine; response; stress management

DESCRIPTION (provided by applicant): The nucleotide (p)ppGpp is critical for stress resistance and antibiotic tolerance. In the Gram-negative bacterium Escherichia coli, (p)ppGpp interacts with RNA polymerase to reprogram transcription upon stress. However, in Firmicutes, a major class of Gram- positive bacteria comprised of pathogens (Staphylococcus, Enterococcus, Listeria, Bacillus anthracis) and beneficial microbes, how (p)ppGpp mediates stress resistance remains poorly understood. The central hypothesis of this proposal is that, in Firmicutes, (p)ppGpp regulates GTP levels to adjust pleiotropic aspects of cellular metabolism to the external environment, thus maintaining homeostasis and facilitating antibiotic tolerance. Our recent work with the bacterium Bacillus subtilis, a widely studied, genetically amenable model for Firmicutes, lends strong support for this hypothesis. Integrating genomic and metabolomic approaches with genetics and biochemistry, we identified enzymes in the GTP biosynthesis pathway as major targets for (p)ppGpp action, and unmasked a novel GTP-(p)ppGpp feedback loop that is critical for cellular viability. Our discoveries thus provide a previously unrecognized view of (p)ppGpp synthesis and function in B. subtilis. We aim to build on this foundation to develop a comprehensive model of stress-management via (p)ppGpp-GTP in B. subtilis that our data predicts will be applicable to Firmicutes and beyond. The specific aims are: 1) Characterize how (p)ppGpp regulates its targets Gmk and HprT; 2) Elucidate how (p)ppGpp controls GTP feedback and glycolysis; and 3) Define how (p)ppGpp contributes to antibiotic tolerance.

描述（由申请人提供）：核苷酸 (p)ppGpp 对于应激抗性和抗生素耐受性至关重要。在革兰氏阴性细菌大肠杆菌中，(p)ppGpp 与 RNA 聚合酶相互作用，在应激时重新编程转录。然而，在厚壁菌门（由病原体（葡萄球菌、肠球菌、李斯特菌、炭疽杆菌）和有益微生物组成的一类主要革兰氏阳性菌）中，(p)ppGpp 如何介导应激抵抗仍知之甚少。该提议的中心假设是，在厚壁菌门中，(p)ppGpp 调节 GTP 水平，以调整细胞代谢对外部环境的多效性，从而维持体内平衡并促进抗生素耐受。我们最近对枯草芽孢杆菌的研究为这一假设提供了强有力的支持，枯草芽孢杆菌是一种经过广泛研究的、遗传上适合厚壁菌门的模型。将基因组学和代谢组学方法与遗传学和生物化学相结合，我们确定了 GTP 生物合成途径中的酶作为 (p)ppGpp 作用的主要靶标，并揭示了一个对细胞活力至关重要的新型 GTP-(p)ppGpp 反馈环。因此，我们的发现为枯草芽孢杆菌中的 (p)ppGpp 合成和功能提供了以前未被认识的观点。我们的目标是在此基础上开发一个通过枯草芽孢杆菌 (p)ppGpp-GTP 进行压力管理的综合模型，我们的数据预测该模型将适用于厚壁菌门及其他物种。具体目标是： 1) 描述 (p)ppGpp 如何调节其靶标 Gmk 和 HprT； 2) 阐明(p)ppGpp如何控制GTP反馈和糖酵解； 3) 定义 (p)ppGpp 如何促进抗生素耐受。