Collaborative Research: Resolving the production and fate of nitrogenous metabolites in the surface ocean

合作研究：解决海洋表层含氮代谢物的产生和归宿

• 批准号: 2125886
• 负责人: Anitra Ingalls
• 金额: $ 89.81万
• 依托单位: University of Washington
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2125886&HistoricalAwards=false
• 关键词: Collaborative; Research; Resolving; production; fate

Photosynthetic microbes provide food for nearly all other life in the ocean. Their metabolism produces organic molecules called metabolites that can leak out of cells, be intentionally excreted into seawater, or be released during cell death. Once outside the cell, these metabolites are the basis for specific interactions among microbes and determine community structure and activity. Yet, current understanding of metabolites in the ocean is limited by a historical lack of ability to measure them. The work proposed here will expand current knowledge of metabolite structures, concentrations, and production rates using recently developed analytical methods. These methods have already led to the discovery that homarine, a substituted pyridine first found in lobster in 1933, is the most abundant detectable metabolite in microbial communities of the North Pacific Ocean. While homarine is known as a predator deterrent, osmoprotectant, methyl donor, and antibiofouling agent, studies of its role in microbial community dynamics are lacking. The work proposed will clarify the role of homarine in the ocean’s microbial communities. This work will create an open-source metabolite database that will serve the broader field of metabolomics, a growing area in environmental, engineering, and medical sciences. This collaboration will also promote the careers of a graduate student and a postdoctoral researcher as well as an early career professor from an underrepresented group at a primarily undergraduate institution (PUI). Undergraduates from both institutions will contribute to project development and implementation, local cruises on the R.V. Carson, lab work, and dissemination of results. This research will be integrated into a curriculum-based research experience for undergraduates in a 200-level genetics course at the PUI, University of Puget Sound. The proposed work will carry out field studies and laboratory experiments to test the hypothesis that metabolites are quantitatively significant forms of carbon and nitrogen flowing through microbial communities. The identity, quantity, and production rates of metabolites will also be determined. For homarine, the enzymes and organisms responsible for its transformations will be determined. Specific proposed activities will 1) Quantify nitrogenous metabolite pools and their net production rates (particulate and dissolved) in phytoplankton cultures and in marine surface water communities; 2) Isolate homarine consuming heterotrophic bacteria and use mutagenesis techniques, transcriptomics, and stable isotope assisted metabolomics to annotate genes and characterize the biochemical reactions involved in the degradation of homarine; 3) Carry out incubations of stable isotope labeled homarine in phytoplankton cultures, heterotrophic bacterial cultures sensitive to homarine, and natural communities to quantitatively evaluate the effect of homarine on growth, track homarine through metabolic pathways, and determine the kinetics of homarine uptake; 4) Identify homarine consumers and biochemical pathways for homarine use in the environment by mining existing environmental metatranscriptomes for homarine catabolism genes. The combination of these approaches will provide better understanding of the flow of nitrogen containing metabolites through marine microbial ecosystems. Results from this work will be disseminated through peer reviewed open-source publications as well as presentations to the scientific community and the general public.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

光合微生物为海洋中几乎所有其他生物提供食物。它们的新陈代谢产生被称为代谢物的有机分子，这些分子可以从细胞中泄漏出来，被故意排泄到海水中，或者在细胞死亡时被释放出来。一旦在细胞外，这些代谢物是微生物之间特定相互作用的基础，并决定群落结构和活性。然而，目前对海洋中代谢物的了解受到历史上缺乏测量能力的限制。这里提出的工作将扩大目前的知识代谢产物的结构，浓度和生产率使用最近开发的分析方法。这些方法已经导致发现，homarine，1933年首次在龙虾中发现的取代吡啶，是北太平洋微生物群落中最丰富的可检测代谢物。虽然homarine被认为是一种捕食者威慑剂，植物保护剂，甲基供体和抗生物污损剂，但缺乏对其在微生物群落动态中作用的研究。这项工作将阐明homarine在海洋微生物群落中的作用。这项工作将创建一个开源代谢物数据库，将服务于代谢组学的更广泛领域，这是环境，工程和医学科学中一个不断增长的领域。这种合作也将促进研究生和博士后研究人员的职业生涯，以及来自一个代表性不足的群体在主要是本科院校（PUI）的早期职业生涯教授。来自这两个机构的本科生将有助于项目的开发和实施，在R. V.当地巡航卡森，实验室工作和结果的传播。这项研究将被整合到一个基于知识的研究经验，为本科生在200级遗传学课程在普吉特湾大学的PUI。拟议的工作将进行实地研究和实验室实验，以检验代谢物是通过微生物群落流动的碳和氮的定量重要形式的假设。还将测定代谢物的鉴别、数量和产生速率。对于homarine，将确定负责其转化的酶和生物体。具体的拟议活动将：1）量化含氮代谢物库及其净生产率（颗粒和溶解的）在浮游植物培养物和海洋表面水群落中; 2）分离出消耗高铁碱的异养细菌，并使用诱变技术、转录组学和稳定同位素辅助代谢组学来注释基因并表征高铁碱降解中涉及的生化反应; 3）在浮游植物培养物、对高铁血红素敏感的异养细菌培养物和自然群落中进行稳定同位素标记的高铁血红素的孵育，以定量评估高铁血红素对生长的影响，通过代谢途径追踪高铁血红素，并确定高铁血红素吸收的动力学; 4）通过挖掘现有的环境元转录组的homarine catalysts基因，鉴定homarine消费者和环境中homarine使用的生化途径。这些方法的结合将提供更好地了解含氮代谢物通过海洋微生物生态系统的流动。这项工作的结果将通过同行评审的开源出版物以及向科学界和公众的演示来传播。该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的知识价值和更广泛的影响审查标准进行评估来支持。