Quantitative studies of metabolic switches in enteric bacteria

描述（由申请人提供）：本研究讨论了肠道细菌对碳源的分层使用，以及细菌从一种碳源切换到另一种碳源时发生的生长转变动力学。这种现象被称为双生长，是雅克·莫诺在65年前发现的。通常认为这是由“分解代谢物阻遏”引起的，这是一种在大肠杆菌中具有分子特征的调节反应。大肠杆菌和广泛传播的微生物。然而，最近的研究表明，分解代谢物抑制是关于协调碳代谢与其他代谢和细胞生长部门，而不是关于优先使用碳。这项研究旨在阐明肠道细菌在环境中无限组合的碳源中进行选择的调控策略（通常首先采用快速代谢的碳），以及使它们能够在首选碳源耗尽时快速转换碳源的动力学机制。研究将采用多种方法进行：传统的生物化学和分子生物学方法，以量化关键信号分子库;定量蛋白质组学，以表征导致生长过渡期间蛋白质组范围重塑的蛋白质合成和周转;合成遗传结构，使人们能够定量探测不断变化的代谢通量和蛋白质负荷对生长过渡的影响;基于微流体的方法，以表征细胞生长，基因表达和信号分子在细胞的水平;和定量现象学的方法，以开发粗粒度的动力学模型，捕捉生理反应，并将它们与潜在的监管机制。拟议的工作本质上是我们实验室近年来开发的非常成功的方法的主要扩展，该方法将基因表达与生长生理学联系起来，但从稳态到动力学领域。这项研究的输出，一个定量预测模型的diauxic增长相关的关键分子相互作用的生理行为，将提供一个原型，用于建模相关的各种各样的其他问题，从细菌对抗生素的反应，分化的动力学发展后，进入稳定期。关于肠道细菌如何选择其碳偏好的具体知识可以在代谢工程应用中被利用，以去除或改变碳消耗的顺序，而关于生长转变的调节策略的知识可能会导致旨在减缓生长恢复的新型抗微生物策略。

DESCRIPTION (provided by applicant): This research addresses the hierarchical usage of carbon sources by enteric bacteria, and the kinetic of growth transition that occurs when bacteria switch from one to another carbon source. This phenomenon, known as diauxic growth, was discovered by Jacques Monod 65 years ago. It was commonly thought to result from "catabolite repression", a regulatory response well characterized molecularly in E. coli and wide spread among microbes. However, recent studies establish that catabolite repression is about coordinating carbon metabolism with other sectors of metabolism and cell growth, and not about prioritizing the use of carbons. This research aims to elucidate the regulatory strategies enteric bacteria employ to choose among an infinite combination of carbon sources in the environment (often taking the fast-metabolizing carbons first), and the kinetic mechanism that enable them to switch carbon source rapidly when the preferred one runs out. The research will be carried out using a combination of approaches: traditional biochemical and molecular biology approaches to quantify the pools of key signaling molecules; quantitative proteomics to characterize protein synthesis and turnover that result in proteome-wide remodeling during growth transitions; synthetic genetic constructs that allow one to quantitatively probe the effect of changing metabolic fluxes and protein loads on growth transitions; microfluidic-based approaches to characterize cell growth, gene expression, and signaling molecules at a cell-by-cell level; and quantitative phenomenological approaches to develop coarse- grained kinetic models that capture the physiological responses and relate them to the underlying regulatory mechanisms. The proposed work is in essence a major extension of the highly successful approach our lab has developed in recent years to relate gene expression to growth physiology, but from the steady state to the kinetic domains. The output of this research, a quantitative predictive model of diauxic growth relating key molecular interactions to physiological behaviors, will provide a prototype for modeling the kinetics of growth transitions relevant to a wide variety of other problems ranging from the response of bacteria to antibiotic, to the kinetics of differentiation an development after entering the stationary phase. The specific knowledge on how enteric bacteria select their carbon preferences may be exploited in metabolic engineering applications to remove or alter the order of carbon consumption, while knowledge on the regulatory strategies of growth transitions may lead to new classes of antimicrobial strategies aimed at slowing down growth recovery.