Ultrahigh sensitivity metabolomics: opening a window into the metabolic evolution of a sentinel invertebrate through the industrial revolution

代谢组学是一项成熟的技术，可以发现生物体如何应对压力的机制。然而，所采用的质谱学方法还不能提供足够的分析灵敏度来测量环境研究中经常遇到的非常小的样品中的代谢物。纳米液质联用技术(NanoLC-MS)在蛋白质组学中得到了广泛的应用，具有很高的灵敏度，但由于诸多技术挑战，在代谢组学中尚未得到应用。在这里，我们建议与Thermo Fisher Science合作，建立和优化新的纳米LC-MS方法，以实现环境毒理学中的变革性研究。水蚤是淡水无脊椎动物，在食物网中起着核心作用，在生态毒理学中得到了广泛的研究。它们可以形成休眠的卵，积累在沉积物中--沉积物可以在许多世纪后孵化成有功能的动物--从而重建对过去环境变化的反应。这被称为复活生态，目前的记录是700岁的水蚤复活。因此，沉积物岩心中休眠蛋的生物化学研究提供了一个独特的机会，可以调查从工业革命之前和整个工业革命到现代集约农业和大量使用杀虫剂的时间范围内污染物对动物健康的影响。然而，水蚤卵重达1毫克，目前的代谢组学技术无法检测到这一重量。基于我们在代谢组学领域长达十年的跟踪记录，我们建议建立和优化灵敏、可重复性和健壮的NanoLC-MS代谢组学技术，然后应用这些技术通过历史上不断变化的污染物水平来发现水蚤适应性的生化基础。因此，我们将发现环境污染和这种前哨淡水物种的健康之间的联系。我们的案例合作伙伴Thermo Fisher Science(TFS)是开发纳米LC-MS的世界领先者，并已将代谢组学确定为优先领域。2013年，TFS与伯明翰大学建立了技术联盟伙伴关系(TAP)，这是欧洲范围内的第一个此类伙伴关系。这项提议建立在现有的五个技师培训中心奖项的基础上，作为其对研究生培训的承诺的一部分。该团队将共同提供专业培训：纳米LC-MS，包括获得当前和预先发布的代谢组学技术的无与伦比的机会(在TFS)；在代谢组学、环境毒理学和分析科学(Viant实验室；全国最大的环境代谢组学组织)。TRANSFERABLE技能将在伯明翰和TFS教授，通过生物科学研究生院的广泛课程，包括商业意识、项目管理和金融培训。这一培训将真正做到多学科，丰富学生体验。此外，首席导师经验丰富，在过去五年中完成了5个博士学位，并有5名在校生，其中一些人是/曾经是NERC iCASE。

METABOLOMICS is a proven technology for discovering mechanisms of how organisms respond to stress. However, the mass spectrometry methods employed do not yet provide sufficient analytical sensitivity to measure metabolites in the very small samples that are often encountered in environmental studies. Nanoscale liquid chromatography-mass spectrometry (nanoLC-MS) is applied extensively in proteomics - offering high sensitivity - but is not yet used in metabolomics due tomany technical challenges. Here we propose to partner with Thermo Fisher Scientific to establish and optimise novel nanoLC-MS methods to enable transformative investigations in environmental toxicology. Daphnia are freshwater invertebrates that play a central role in foodwebs and are widely studied in ecotoxicology. They can form dormant eggs that accumulate in sediments - which can hatch into functional animals many centuries later - allowing reconstruction of responses to past environmental change. This is termed resurrection ecology, and the current record isthe resurrection of 700 yr old Daphnia. The biochemical study of dormant eggs in sediment cores therefore offers a unique opportunity to investigate pollutant impacts on animal fitness from well before and throughout the industrial revolution, up to modern timescales of intensive agriculture and high pesticide usage. Yet Daphnia eggs weigh <1 mg, undetectable by current metabolomics technologies. Building on our decade-long track record in metabolomics, we propose to establish and optimise nanoLC-MS metabolomics technologies that are sensitive, reproducible and robust, and then apply these to discover the biochemical basis of Daphnia fitness through historically changing pollutant levels. Hence we will discover associations betweenenvironmental pollution and the health of this sentinel freshwater species.Our CASE PARTNER, Thermo Fisher Scientific (TFS), is a world leader in the development of nanoLC-MS and has identified metabolomics as a priority area. In 2013, TFS formed a Technology Alliance Partnership (TAP) with the University of Birmingham, the first such partnership within Europe. This proposal builds upon five existing iCASE awards with TFS as part of their commitment to graduate training. Collectively this team will provide SPECIALIST TRAINING: in nanoLC-MS, including unparalleled access to current and pre-released metabolomics technologies (at TFS); in metabolomics, environmental toxicology and analytical sciences (Viant lab; the largest group in environmental metabolomics nationally).TRANSFERABLE SKILLS will be taught at both Birmingham, through the extensive courses in the Biosciences Graduate Research School, and at TFS, including business awareness, project management and financial training. This training will be truly MULTIDISCIPLINARY to enrich the student experience. Furthermore the main supervisor is highly experienced,having completed 5 PhDs in the past five years and with 5 current students, several of which are/were NERC iCASE.