DNA Uptake By Naturally Competent Archaea

微生物可以通过一种称为转化的过程直接从环境中获得遗传物质。这些生物属于两大类，细菌和古细菌。转化在细菌中得到了很好的研究，但对这在古生菌中的作用知之甚少，尽管古生菌代表了地球上至少三分之一的生物多样性，并在大型工业和市政生物反应器中催化全球生物地球化学循环中的基本反应。为了解决这一缺陷，这一建议将是第一个表征的转变在一个成员的古菌。通过这项工作，将确定进行这一过程所需的必要基因和蛋白质。这些结果可能会进一步努力研究生物从古生菌通过遗传技术。这项研究有可能直接和积极地影响古细菌催化的任何过程的研究。此外，转化在自然种群中很重要，这里产生的结果将有助于更广泛地理解环境中的基因流动。最后，作为本研究的一部分开发的技术将直接有助于下一代科学家的教育，通过研究生培训和加强本科生和研究生课堂上的实践实验。本项目将描述古细菌领域的自然转化过程-由有能力的生物体直接摄取和基因组整合环境中的DNA。这将通过鉴定和表征古细菌DNA摄取机制的组成部分来实现，这些组成部分参与三个不同的转化步骤：DNA的细胞外结合，跨细胞膜的运输和DNA在细胞质中的维持。确定这些步骤中涉及的基因和蛋白质将有助于通过计算机预测生物体吸收环境DNA的能力。该项目将利用转座子诱变，体外生化分析，和表型测定，以表征转化的甲烷球菌maripaludis，模式生物的古菌域的成员的研究。为了进一步促进古细菌转化机制的表征，本提案中的工作将开发和利用厌氧古细菌活细胞荧光成像系统，以分析主管古细菌中的蛋白质定位和蛋白质-蛋白质相互作用。该奖项反映了NSF的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。

Microorganisms can acquire genetic material directly from their environment through a process called transformation. These organisms belong to two broad groups, the bacteria and the archaea. Transformation is well studied in bacteria, but little is known about how this works in archaea, despite the fact that archaea represent at least a third of the biodiversity on Earth and catalyze essential reactions in both global biogeochemical cycles in large-scale industrial and municipal bioreactors. To address this deficiency, this proposal will be the first characterization of transformation in a member of the archaea. Through this work, the necessary genes and proteins needed to carry out this process will be identified. These results may further efforts to study organisms from the archaea through genetic techniques. This research has the potential to impact directly and positively research in any process that archaea catalyze. Additionally, transformation is important in natural populations, and the results generated here will help develop a broader understanding of gene flow in the environment. Finally, the techniques developed as part of this research will directly contribute to the education of the next generation of scientists through graduate student training and enhancing hands-on experimentation in undergraduate and graduate classrooms.This project will characterize the process of natural transformation - the direct uptake and genomic incorporation of DNA from the environment by competent organisms - in the archaeal domain. This will be accomplished by identifying and characterizing components of the archaeal DNA uptake machinery involved in three distinct steps of transformation: extracellular binding of DNA, transport across the cell membrane, and maintenance of DNA in the cell cytoplasm. Identifying the genes and proteins involved in these steps will facilitate in silico predictions of an organism’s ability to take up environmental DNA. This project will leverage transposon mutagenesis, in vitro biochemical analysis, and phenotypic assays to characterize transformation in Methanococcus maripaludis, a model organism for the study of members of the archaeal domain. To further facilitate the characterization of archaeal transformation machinery, the work in this proposal will develop and utilize a system for live cell fluorescent imaging of anaerobic archaea to analyze protein localization and protein-protein interactions in competent archaea.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.