Collaborative Research: Gene content, gene expression, and physiology in mesopelagic ammonia-oxidizing archaea

合作研究：中层氨氧化古菌的基因含量、基因表达和生理学

• 批准号: 1739144
• 负责人: Alyson Santoro
• 金额: $ 1.03万
• 依托单位: University of California-Santa Barbara
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1739144&HistoricalAwards=false
• 关键词: Collaborative; Research; Gene; content; gene

Intellectual Merit. How organisms respond to their physical and chemical and environment is a central question in marine ecology. For microbes living in the mesopelagic - the ocean's "twilight zone" - an efficient response is particularly important to capitalize on the intermittent delivery of organic and inorganic compounds sinking from the surface ocean. These organisms must have a suite of metabolic and regulatory strategies used to cope with environmental variability, but these strategies are largely unknown. Understanding when and why metabolic genes are expressed is critical to our understanding of nutrient remineralization in the ocean. Marine group 1 (MG1) archaea are ubiquitous, abundant microbes in the meso- and bathypelagic and promising model organisms for investigating these questions. MG1 archaea are chemolithoautotrophs that oxidize ammonia for energy and fix carbon for biomass, and as such, play a central role in the ocean's coupled carbon and nitrogen cycles. Though MG1 have historically eluded cultivation, recent efforts have been successful at bringing representative MG1 archaea from the open ocean into culture and demonstrating their importance in the production of the greenhouse gas nitrous oxide. This project takes advantage of unique MG1 cultures and the recently sequenced draft genome of one of the organisms - strain CN25 - to investigate the physiological and transcriptional responses of MG1 archaea to variations in their chemical environment, specifically:1. Comparative transcriptomics of CN25 cells grown under a range of energy availability and nitrosative stress will identify select genes that can be used to diagnose the physiological state of natural populations2. Improvements in the genomic and transcriptomic knowledge of MG1 archaea will facilitate a thorough reinterpretation of existing metagenomic and metatranscriptomic datasets, as well as provide a better contextual understanding in future studiesThe investigators will conduct comparative transcriptomics of CN25 cells harvested in mid-exponential growth and stationary phase versus starved cells. Transcriptomes of cells grown at high nitrate concentrations and low pO2 with those grown in standard conditions will be characterized. A strand-specific, high-density RNAseq approach will be used to examine the expression of putative ORFs, polycistronic operons, and small RNAs, which, in addition to gene expression profiling, has the ancillary benefit of improving genome annotation. Finally, the investigators will sequence the genomes of two additional MG1 strains isolated from the open ocean, as well as single cells from environmental surveys, and leverage the combination with the CN25 genome to reanalyze available metagenomic and metatranscriptomic datasets. The results will define the transcriptional response of a model mesopelagic microbe to a range of chemical environments, and show how the physicochemical environment induces changes in gene expression and gene content that result in greenhouse gas production. This work will rapidly generate new knowledge of how some of the most ubiquitous, yet heretofore elusive, microorganisms respond to geochemical variability and shape our evolving understanding of the marine nitrogen cycle.Broader Impacts. The scientific and societal impact of the project will be to elucidate the mechanisms of greenhouse gas production in a model marine organism that is of broad interest to biological and chemical oceanographers. Transcriptome sequencing will improve the assembly of the CN25 genome, the first genome of an MG1 archaeon from the open ocean. Both the genome and transcriptomes will be important references for researchers using metagenomics, metatranscriptomics, and metaproteomics in the ocean, as these techniques are reliant on a knowledgebase composed of both DNA sequence and physiology. Thus, the results add value to both existing and future studies. The proposed research will advance education, teaching, and training for the next generation of marine scientists by providing support for two early-career investigators, one postdoctoral researcher, and a secondary school teacher.

智力优点。生物体如何对其物理、化学和环境做出反应是海洋生态学的核心问题。对于生活在中层（海洋的“暮光区”）的微生物来说，有效的反应对于利用从表层海洋下沉的有机和无机化合物的间歇性输送尤其重要。这些生物体必须具有一套用于应对环境变化的代谢和调节策略，但这些策略在很大程度上是未知的。了解代谢基因何时以及为何表达对于我们了解海洋营养物再矿化至关重要。海洋第 1 类 (MG1) 古细菌是中海和深海中普遍存在且丰富的微生物，是研究这些问题的有前景的模式生物。 MG1 古细菌是化能自养生物，可氧化氨以获取能源并固定碳以获取生物质，因此在海洋碳氮耦合循环中发挥着核心作用。尽管 MG1 历来未能进行培养，但最近的努力已成功地将具有代表性的 MG1 古菌从公海带入养殖，并证明了它们在产生温室气体一氧化二氮方面的重要性。该项目利用独特的 MG1 培养物和最近测序的其中一种生物体 - 菌株 CN25 的基因组草案，来研究 MG1 古细菌对其化学环境变化的生理和转录反应，具体来说：1。在一系列能量可用性和亚硝化胁迫下生长的 CN25 细胞的比较转录组学将鉴定可用于诊断自然群体生理状态的精选基因2。 MG1古菌基因组和转录组知识的改进将有助于对现有宏基因组和宏转录组数据集进行彻底的重新解释，并在未来的研究中提供更好的背景理解研究人员将对在指数生长中期和稳定期收获的CN25细胞进行比较转录组学与饥饿的细胞相比。将表征在高硝酸盐浓度和低 pO2 下生长的细胞的转录组以及在标准条件下生长的细胞的转录组。链特异性、高密度 RNAseq 方法将用于检查假定的 ORF、多顺反子操纵子和小 RNA 的表达，除了基因表达谱之外，该方法还具有改善基因组注释的附带好处。最后，研究人员将对从公海分离的另外两种 MG1 菌株以及环境调查中的单细胞进行基因组测序，并利用与 CN25 基因组的组合来重新分析可用的宏基因组和宏转录组数据集。结果将定义中层微生物模型对一系列化学环境的转录反应，并显示物理化学环境如何诱导基因表达和基因内容的变化，从而导致温室气体的产生。这项工作将迅速产生关于一些最普遍但迄今为止难以捉摸的微生物如何响应地球化学变化的新知识，并塑造我们对海洋氮循环不断发展的理解。更广泛的影响。该项目的科学和社会影响将是阐明生物和化学海洋学家广泛感兴趣的模型海洋生物中温室气体产生的机制。转录组测序将改善 CN25 基因组的组装，CN25 基因组是来自公海的 MG1 古菌的第一个基因组。基因组和转录组都将成为研究人员在海洋中使用宏基因组学、宏转录组学和宏蛋白质组学的重要参考，因为这些技术依赖于由 DNA 序列和生理学组成的知识库。因此，结果为现有和未来的研究增加了价值。拟议的研究将为两名早期职业研究人员、一名博士后研究员和一名中学教师提供支持，从而促进下一代海洋科学家的教育、教学和培训。