Defining adaptive processes in a model bacterial species by integrating proteomic and metabolomic analytical strategies

BBSRC相关性：该项目将KCL和ASTA联系起来，与BBSRC的战略政策领域“生物科学的系统方法”和“生物科学的技术开发”直接相关（见http：//www.bbsrc.ac.uk/funding/grants/priorities/）。正如在下面的影响部分，在BBSRC战略政策领域的抗菌素耐药性和农业的好处也是预期outcomes.AIM：利用尖端技术来调查复杂的监管机构，促进铜绿假单胞菌对它的物种内diversity.Background的传播：新的DNA测序技术的到来，导致了爆炸的人和许多环境的微生物组的数据。虽然这些信息提供了一个全面的物种目录，其中许多目前无法培养，但这些数据既没有深入了解微生物的功能，也没有深入研究如何传播到新栖息地的问题。微生物的栖息地是多种多样的。然而，一些关键物种可以超越它们原来的“边界”，开拓新的生态位。尽管几个研究中心试图绘制通常高度耐抗生素的物种从环境到人类的传播途径，但目前尚不清楚使微生物离开其正常栖息地并殖民新地点的驱动力。本研究将探讨铜绿假单胞菌作为一种模型，由于极端的代谢多样性和“无处不在”的新网站，它显示殖民的适应性潜力的关键监管机构。本种和密切相关的分类群也广泛用于许多工业过程，包括生物修复。此外，由于铜绿假单胞菌在染色体上携带抗生素耐药基因的能力，它是目前最大的治疗挑战之一，并对环境造成巨大威胁。该项目背后的基本假设是，适应过程的表型可塑性选择性地使微生物能够离开其正常栖息地并殖民新的地点，这是由过多的关键调控因子驱动的，信号分子，在铜绿假单胞菌的情况下占其基因组的近10%。然而，目前的证据表明，适应过程的表型可塑性在于蛋白质组和代谢组水平。一旦转录，许多涉及的蛋白质/代谢物可以被修饰（翻译后修饰- PTM）以促进它们的转变。因此，绘制这些过程不仅涉及这些复杂生物分子的检测和定量，还涉及其修饰的蛋白质形式。直到最近，蛋白质和PTM的高通量检测还在生物科学的范围之外。如今，随着质谱（MS）和核磁共振光谱的新形式，这样的分析是可能的，但技术和技能一直局限于相对较少的专业实验室。这个工业案例奖学金将建立在监管团队对铜绿假单胞菌的工作经验，从学术界和（KDB，AJM）和工业（OB，HNS）研究与其种群结构相关的代谢组和蛋白质组。将选择来自所选谱系的代表性菌株进行比较深入分析，以阐明调控标记，其中所选谱系对水、污水、土壤等几个生态位具有超适应性。这些将使用每个鉴定的标志物的RNA表达分析进行验证，以证实其分布和表达水平。该项目将与学生一起开发，通过四年期间的重叠期将培训和实验工作结合起来，如下所述。

BBSRC RELEVANCE: This project, linking KCL and ASTA, is of direct relevance to BBSRC Strategic Policy areas "Systems approaches to the biosciences" and "Technology development for the biosciences" see http://www.bbsrc.ac.uk/funding/grants/priorities/). As in the Impact section below, benefits in BBSRC Strategic Policy areas relating to antimicrobial resistance and agriculture are also expected outcomes.AIM: To utilise cutting-edge technologies to investigate the complex regulators that facilitate transmission of Pseudomonas aeruginosa against its intra-species diversity.BACKGROUND: The arrival of novel DNA sequencing technologies have led to an explosion of data on the microbiome of man and numerous environments. While such information provides a comprehensive catalogue of species, many of which cannot be cultured at present, the data give no insight into the function of the microbe nor the incisive question of how transmission to a new habitat occurs. The habitats that microbes colonise are diverse. A few key species can however transcend their original "boundaries" to colonise new niches. Despite attempts by several research centres to map routes of transmission of often highly antibiotic resistant species from the environment to man, the driving forces that enable a microbe to leave its normal habitat and colonise a new site are not currently known. This study will investigate key regulators of the adaptive potential of Pseudomonas aeruginosa as a model due to the extreme metabolic versatility and "ubiquitous" colonisation of new sites that it displays. This species and closely related taxa are moreover used widely in many industrial processes including bioremediation. In addition, due to its remarkable capacity to harbour antibiotic resistance genes chromosomally, P. aeruginosa is currently one of our greatest therapeutic challenges and poses an immense environmental threat.HYPOTHESIS: The underlying hypothesis behind this project is that the phenotypic plasticity for adaptive processes that selectively enables a microbe to leave its normal habitat and colonise a new site is driven by a plethora of key regulators and signaling molecules, which in the case of P. aeruginosa accounts for nearly 10% of its genome. Current evidence indicates however that the phenotypic plasticity for adaptive processes lies at the level of the proteome and metabolome. Once transcribed, many of the proteins/metabolites involved may be modified (post-translational modification - PTM) to facilitate their transition. Mapping these processes therefore involves not only detection and quantitation of these complex biomolecules but their modified proteoforms. Until recently, high throughput detection of proteins and PTMs were outside the reach of bioscience. Today, with novel forms of Mass spectrometry (MS) and NMR spectroscopy, such analyses are possible but the technologies and skills have been limited to relatively few specialist laboratories.PLAN: This Industrial CASE Studentship will build upon the experience of the supervisory team's work on P. aeruginosa from both academia (KDB, AJM) and industry (OB, HNS) to investigate the metabolome and proteome in relation to its population structure. Representative strains from selected lineages where hyper-adaptation to several ecological niches such as water, sewage, soil etc, will be selected for comparative in-depth analysis to elucidate regulatory markers. These will be validated using RNA expression analysis of each identified marker to corroborate their distribution and expression level. The project will be developed with the student combining training and experimental work through overlapping periods during the four years as outlined below.