Integrating Global Responses to Nutrient Limitation in Gram-positive Bacteria

整合全球对革兰氏阳性菌营养限制的反应

• 批准号: 8737911
• 负责人: Shaun R Brinsmade
• 金额: $ 24.21万
• 依托单位: GEORGETOWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8737911
• 关键词: Affect; Amino Acids; Award; Bacillus subtilis; Bacteria; Bacterial Infections; Bacterial Physiology; Behavior; Benign; Binding; Biochemical; Biochemistry; Branched-Chain Amino Acids; Calculi; Cells; Development; Disease; Foundations; Funding; Gene Expression; Gene Expression Profile; Genes; Genetic; Goals; Gram-Positive Bacteria; Guanine Nucleotides; Guanosine Triphosphate; Health; Heart; Human; Human body; In Vitro; Infection; Isoleucine; Knowledge; Laboratories; Lead; Learning; Leucine; Life; Life Style; Maps; Mass Spectrum Analysis; Massive Parallel Sequencing; Measures; Mediating; Mentors; Metabolic; Metabolism; Methods; Mission; Modeling; Monitor; National Institute of General Medical Sciences; Nosocomial Infections; Nutrient; Output; Pathogenesis; Pathway interactions; Phase; Physiological; Physiology; Process; Public Health; Regulation; Regulon; Research; Research Personnel; Resources; Role; Route; Signal Transduction; Soil; Staphylococcus aureus; System; Systems Biology; Techniques; Therapeutic; Time; Training; Universities; Valine; Variant; Virulence; abstracting; antimicrobial; base; biological adaptation to stress; career; career development; design; environmental change; exhaustion; functional genomics; genetic manipulation; in vivo; innovation; medical schools; metabolomics; novel; novel therapeutics; nucleoside triphosphate; pathogen; prevent; programs; promoter; research and development; research study; response; tool; transcription factor

7. Project Summary/Abstract. Expression of bacterial virulence genes often correlates with the exhaus- tion of nutrients, but how the signaling of nutrient availability and the resulting physiological responses are co- ordinated is unclear. Until this gap in knowledge is closed, metabolically diverse bacteria like Staphylococcus aureus will continue to cause perilous hospital-acquired infections. The applicant's long-term goal is to lead an independent academic research group studying how bacteria integrate and respond to information provided by intracellular metabolites (the metabolome) to reconfigure metabolism to adapt to environmental changes and cause disease. The objective of this project is to augment existing genetic and biochemical expertise with high- throughput global techniques to analyze gene expression, intracellular metabolites and flux, and, in doing so, titrate the activity of the global regulator CodY and deduce its regulatory hierarchy in S. aureus. At the heart of this project is the hypothesis that fluctuations in the intracellular pools of branched-chain amino acids and GTP result in a spectrum of CodY activities that produce a graded response to nutrient limitation, culminating in metabolic adaptation and the development of virulence. This hypothesis is based on preliminary studies that identified the true intracellular metabolites that control CodY activity in living cells and revealed hierarchical or- ganization for three genes. The rationale for this project is that comprehensive knowledge of the co-regulation of metabolism and virulence is essential if we are to understand the physiological origins of bacterial patho- genesis. During the mentored (K99) phase at Tufts University School of Medicine, massively parallel sequenc- ing, mass spectrometry-based metabolomics and chemostat cultivation will be mastered to map intersecting metabolic and virulence gene expression patterns in S. aureus, while gaining critical scholarly training needed to launch a successful independent academic career with guidance from a mentoring committee composed of experts in bacterial physiology, biochemistry and systems biology. Mastering the cultivation and genetic ma- nipulation of pathogenic S. aureus along with high-throughput methods will enable efforts during the R00 phase to quantify changes in the S. aureus CodY regulon upon induction of physiological stress response sys- tems. The approach is innovative, because continuous bacterial cultures mimic nutrient-limiting bacterial nich- es in the human body and the experiments will place virulence gene expression in the context of the normal behavior of S. aureus under the nutrient-limiting conditions of the host. Furthermore, correlations between global metabolite pools and CodY activity will provide a previously unattainable linkage of the transcriptome to the metabolome. The project is significant because it will increase our understanding of how the genetic pro- grams of metabolic adaptation and virulence gene expression are interrelated and interdependent. A more thorough understanding of these connections may also offer potentially novel therapeutic strategies. The Pathway to Independence Award will provide the time and resources needed to achieve these goals.

7. 项目总结/摘要。细菌毒力基因的表达通常与消耗相关。 营养物质的重刑，但营养物质可用性的信号和由此产生的生理反应是如何共同作用的 协调不明确。在这一知识差距弥合之前，葡萄球菌等代谢多样化的细菌 金黄色葡萄球菌将继续引起危险的医院获得性感染。申请人的长期目标是领导 独立学术研究小组研究细菌如何整合并响应由细菌提供的信息 细胞内代谢物（代谢组）重新配置新陈代谢以适应环境变化和 引起疾病。该项目的目标是通过高水平增强现有的遗传和生化专业知识。 吞吐量全局技术来分析基因表达、细胞内代谢物和通量，并且在此过程中， 滴定全局调节因子 CodY 的活性并推断其在金黄色葡萄球菌中的调节层级。位于 该项目假设细胞内支链氨基酸和 GTP 池的波动 导致一系列 CodY 活性，对营养限制产生分级反应，最终导致 代谢适应和毒力的发展。该假设基于以下初步研究： 确定了控制活细胞中 CodY 活性的真正细胞​​内代谢物，并揭示了分层或- 三个基因的组织。该项目的基本原理是对共同监管的全面了解 如果我们要了解细菌病理学的生理起源，代谢和毒力的研究至关重要 起源。在塔夫茨大学医学院的指导（K99）阶段，大规模并行测序 荷兰国际集团，基于质谱的代谢组学和恒化器培养将被掌握以绘制交叉图 金黄色葡萄球菌的代谢和毒力基因表达模式，同时获得所需的关键学术培训 在由以下人员组成的指导委员会的指导下开展成功的独立学术生涯 细菌生理学、生物化学和系统生物学专家。掌握栽培和遗传技术 致病性金黄色葡萄球菌的消灭以及高通量方法将使 R00 期间的努力成为可能 阶段量化诱导生理应激反应系统后金黄色葡萄球菌 CodY 调节子的变化 项目。该方法具有创新性，因为连续细菌培养模拟了营养限制性细菌生态位。 es 存在于人体中，实验将毒力基因的表达置于正常情况下 金黄色葡萄球菌在宿主营养限制条件下的行为。此外，之间的相关性 全球代谢物库和 CodY 活性将提供以前无法实现的转录组与 代谢组。该项目意义重大，因为它将增加我们对遗传亲缘关系如何的理解。 克代谢适应和毒力基因表达是相互关联和相互依赖的。一个更多 对这些联系的透彻理解也可能提供潜在的新颖的治疗策略。这 独立之路奖将提供实现这些目标所需的时间和资源。