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Environmentally relevant responses in different Escherichia coli pathotypes: a functional genomics study of motility and associated regulons

不同大肠杆菌致病型的环境相关反应：运动性和相关调节子的功能基因组学研究

基本信息

批准号：
BB/E01044X/1
负责人：
Charles Penn
金额：
$ 62.71万
依托单位：
University of Birmingham
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2006
资助国家：
英国
起止时间：
2006 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FE01044X%2F1
关键词：
Environmentally relevant responses different Escherichia

项目摘要

In nature bacteria may act as friends, like 'good' bacteria in our digestive systems, or foes, sometimes as life-threatening pathogens. Escherichia coli can be either friend or foe, and is also a 'model' bacterium, much studied to understand bacterial structure and function. This knowledge is based mainly on a harmless strain named K-12, isolated in 1922. Today, K-12 is so well adapted to the laboratory that it cannot even survive in the human digestive tract. But there are other distinctive 'pathotypes' of E. coli that can cause disease, including E. coli O157. Complete genomic DNA sequences have been determined for K-12 and several other pathotypes. In each, about 20% of its genes are unique, a big surprise since it had been expected that E. coli as a species would be genetically quite homogeneous. So, is K-12 a good model representing the species as a whole? To find out, we will analyse and compare the ways K-12 and other pathotypes respond genetically to conditions that induce them to change their behaviour, focusing on their ability to 'swim' in liquids (motility) and related properties that enable them to survive and persist in the environment. New 'functional genomics' techniques, based on knowledge of genome sequences, enable the expression of every gene in the genome to be monitored simultaneously. This shows how each gene is involved in the behaviour of the bacteria under given conditions. It would be tempting to use these techniques just to fill the gaps in our knowledge of how K-12 functions. But this would neglect the potential to analyse different pathotypes, of known genome sequence, in relation to their behaviour in nature. We propose to exploit that potential to the full. Some transcription networks in E. coli, comprising subsets of genes ('regulons') controlled by proteins called transcription factors or TFs, have received much attention due to their involvement in the basic biochemistry and physiology of the cell. This has been echoed in the choice of networks that have been analysed by functional genomics methods. For example, 3 independent such studies of the FNR regulon involved in adaptation to the absence of oxygen have already been published. In contrast, there has been far less experimental investigation of the regulon that controls motility, governed by the TF complex FlhDC. We believe this TF forms a significant node or 'crossing point' in the global cellular transcription network, interacting with, among others, those that mediate responses to the presence of metals and to oxidative stress. The FlhDC regulon also responds to an important and little-explored signal molecule made within the cell, called cyclic di-GMP. This molecule is known to be involved in the ability of E. coli and related species to colonise solid surfaces in a growth mode known as a 'biofilm' in which motility is strongly repressed. We will use functional genomics methods to analyse the genome-wide transcription activity controlled by FlhDC and associated TFs named Fur and SoxS. We will impose conditions in which the TFs would normally be activated, and monitor global changes in gene expression in representative pathotypes. In parallel experiments we will study gene expression in mutants in which the TFs have been deleted. We also propose to use new methods to study TF binding to different regions of DNA, modulating expression of the genes in those regions. This will provide a definitive picture of how the regulons respond to stimuli and what cellular functions they then modulate to allow the cell to adapt to environmental challenges. We will thus gain exciting insights into the diversity of responses mediated by FlhDC and related TFs in different pathotypes of this uniquely important bacterium, and generate for the first time a global comparison of patterns of gene expression and their control among different representatives of the same bacterial species.

在自然界中，细菌可能是我们消化系统中的“好”细菌，也可能是敌人，有时是威胁生命的病原体。大肠杆菌可以是朋友也可以是敌人，也是一种“模型”细菌，为了了解细菌的结构和功能而进行了大量的研究。这一知识主要基于1922年分离出的一种名为K-12的无害菌株。今天，K-12非常适合实验室，甚至不能在人类消化道中存活。但也有其他独特的“病理型”的E。大肠杆菌，包括大肠杆菌。coli O157。已确定K-12和其他几种致病型的完整基因组DNA序列。在每一个基因中，大约20%的基因是独特的，这是一个很大的惊喜，因为人们一直预计E。大肠杆菌作为一个物种在遗传上是非常同质的。那么，K-12是代表整个物种的好模型吗？为了找到答案，我们将分析和比较K-12和其他致病型对诱导它们改变行为的条件的遗传反应方式，重点是它们在液体中“游泳”的能力（运动性）和使它们能够在环境中生存和坚持的相关特性。新的“功能基因组学”技术，基于基因组序列的知识，能够同时监测基因组中每个基因的表达。这显示了每个基因在给定条件下如何参与细菌的行为。使用这些技术来填补我们对K-12功能的知识空白是很诱人的。但这将忽视分析已知基因组序列的不同病理类型与其自然行为之间关系的潜力。我们建议充分利用这一潜力。E.大肠杆菌中的调节子（regulon）由转录因子（transcription factors，TF）控制，由于其参与细胞的基本生物化学和生理学，因此受到了广泛关注。这一点在功能基因组学方法分析的网络选择中得到了回应。例如，已经发表了3项关于FNR调节子参与适应缺氧的独立研究。相比之下，对由TF复合物FlhDC控制运动的调节子的实验研究少得多。我们认为，这种TF在全球细胞转录网络中形成了一个重要的节点或“交叉点”，与介导对金属存在和氧化应激反应的那些相互作用。FlhDC调节子还响应于细胞内产生的一种重要且很少探索的信号分子，称为环状二GMP。已知该分子参与E.大肠杆菌和相关物种以被称为“生物膜”的生长模式在固体表面定殖，其中运动性被强烈抑制。我们将使用功能基因组学的方法来分析全基因组的转录活性控制FlhDC和相关的TF命名为Fur和SoxS。我们将施加条件，在这些条件下，TF通常会被激活，并监测代表性的病理型基因表达的全球变化。在平行实验中，我们将研究基因表达的突变体中的TF已被删除。我们还建议使用新的方法来研究TF与DNA不同区域的结合，调节这些区域中基因的表达。这将提供一个明确的画面，调节子如何响应刺激，然后它们调节什么细胞功能，使细胞适应环境的挑战。因此，我们将获得令人兴奋的见解FlhDC和相关TF介导的反应的多样性，在不同的病理类型的这种独特的重要细菌，并产生第一次全球比较的模式的基因表达和它们的控制之间的不同代表相同的细菌物种。