Heterogeneity in Colicin E2 expression of Escherichia coli

大肠杆菌大肠杆菌素 E2 表达的异质性

• 批准号: 217673565
• 负责人: Professor Dr. Erwin Frey
• 金额: --
• 依托单位: Center for NanoScience (CeNS)
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 2012
• 资助国家: 德国
• 起止时间: 2011-12-31 至 2018-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/217673565?language=en
• 关键词: Heterogeneity; Colicin; E2; expression; Escherichia

Bacteria possess several mechanisms enabling them to respond to changing and unfavorable environmental conditions or to outcompete other bacteria. One particular mechanism is the production and the release of toxins such as bacteriocins. Current research focuses on production, release and uptake of these toxins by bacteria. However, little is known about the quantitative aspects of the underlying gene regulatory processes. In the first funding period, we quantitatively studied the expression dynamics of the Colicin E2 operon in E. coli on a single cell level using fluorescence time-lapse microscopy. We were able to show that heterogeneous timing is characteristic for Colicin E2 expression at low concentrations of the exogeneous stressor Mitomycin C (MitC), while high concentrations of the exogenous stressor lead to a synchronized stress response of all cells. In the next funding period, we aim to understand the molecular mechanisms underlying Colicin gene expression during SOS response. Extending our previous analysis we are in particular interested how gene expression responds to time-varying environmental conditions. Our approach will be modular: Upon studying cea and cel expression in a broad set of mutant strains we will address the regulatory mechanisms in the various modules of the Colicin E2 system separately and in combination. Due to the complexity of the system it will be essential to closely integrate experimental and theoretical analysis in order to disentangle the relative role of and the interplay between the regulatory elements at the transcriptional and the post-transcriptional level. The experimental analysis will provide large ensembles of time traces for fluorescence reporters of the cea and cel genes for the different mutant strains whose statistical properties will be compared in detail with the results obtained from stochastic simulations of corresponding mathematical models. This will allow us to arrive at a detailed molecular understanding in a bottom-up fashion. These analyses will then lead to a molecular understanding of how the different modules regulating Colicin E2 expression are activated in dependence on given environmental conditions. In particular, this should give us a deeper insight into what causes the observed heterogeneous timing at low stress levels, and how the transition towards a synchronized response of the whole cell population is regulated. Finally, we want to study how phenotypic heterogeneity is passed on within growing bacterial microcolonies. Here, we want to understand if either a Colicin-dependent feedback acting on neighboring Colicin producing cells, or cell density (quorum sensing) affect the fraction of cells expressing Colicin E2. These experiments shall enable us to better understand the biological significance of heterogeneous Colicin E2 expression and how it is sustained within growing microcolonies.

细菌拥有几种机制，使它们能够对不断变化和不利的环境条件做出反应，或与其他细菌竞争。一种特殊的机制是细菌素等毒素的产生和释放。目前的研究重点是细菌对这些毒素的产生、释放和吸收。然而，人们对潜在的基因调控过程的量化方面知之甚少。在第一个资助期，我们利用荧光时移显微镜在单细胞水平上定量研究了Colicin E2操纵子在大肠杆菌中的表达动态。我们能够证明，在低浓度的外源应激源丝裂霉素C(MitC)下，Colicin E2的表达具有不同的时间特征，而高浓度的外源应激源导致所有细胞的同步应激反应。在下一个资金阶段，我们的目标是了解SOS响应过程中Colicin基因表达的分子机制。扩展我们之前的分析，我们特别感兴趣的是基因表达如何对时间变化的环境条件做出反应。我们的方法将是模块化的：在研究了CEA和CEL在广泛的突变菌株中的表达后，我们将分别和结合地研究Colicin E2系统的各个模块中的调控机制。由于该系统的复杂性，有必要密切结合实验和理论分析，以理清转录和转录后水平上调控元件的相对作用和相互作用。实验分析将为不同突变菌株的CEA和CEL基因的荧光报告器提供大量的时间轨迹，其统计特性将与相应数学模型的随机模拟结果进行详细比较。这将使我们能够以自下而上的方式达到详细的分子理解。然后，这些分析将导致对调节Colicin E2表达的不同模块如何根据给定的环境条件而被激活的分子理解。特别是，这应该让我们更深入地了解是什么导致了在低应激水平下观察到的不同时序，以及如何调控整个细胞群体向同步反应的转变。最后，我们想要研究表型异质性是如何在不断增长的细菌微菌落中传递的。在这里，我们想要了解作用于邻近产生Colicin的细胞上的Colicin依赖的反馈，或者细胞密度(群体感应)是否影响Colicin E2表达的细胞比例。这些实验将使我们能够更好地理解异质性Colicin E2表达的生物学意义，以及它是如何在不断增长的微克隆中保持的。