DNA Uptake By Naturally Competent Archaea

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.