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Nat Microbiol. 2019 Dec 16. doi: 10.1038/s41564-019-0631-2. [Epub ahead of print]

Control of nitrogen fixation in bacteria that associate with cereals.

Author information

1
Synthetic Biology Center, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.
2
Departments of Bacteriology and Agronomy, University of Wisconsin-Madison, Madison, WI, USA.
3
Department of Plant Sciences, University of Oxford, Oxford, UK.
4
Department of Chemistry and Biochemistry, Montana State University, Bozeman, MT, USA.
5
Institute of Biological Chemistry, Washington State University, Pullman, WA, USA.
6
Department of Biology, Colorado State University-Pueblo, Pueblo, CO, USA.
7
Synthetic Biology Center, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA. cavoigt@gmail.com.

Abstract

Legumes obtain nitrogen from air through rhizobia residing in root nodules. Some species of rhizobia can colonize cereals but do not fix nitrogen on them. Disabling native regulation can turn on nitrogenase expression, even in the presence of nitrogenous fertilizer and low oxygen, but continuous nitrogenase production confers an energy burden. Here, we engineer inducible nitrogenase activity in two cereal endophytes (Azorhizobium caulinodans ORS571 and Rhizobium sp. IRBG74) and the well-characterized plant epiphyte Pseudomonas protegens Pf-5, a maize seed inoculant. For each organism, different strategies were taken to eliminate ammonium repression and place nitrogenase expression under the control of agriculturally relevant signals, including root exudates, biocontrol agents and phytohormones. We demonstrate that R. sp. IRBG74 can be engineered to result in nitrogenase activity under free-living conditions by transferring a nif cluster from either Rhodobacter sphaeroides or Klebsiella oxytoca. For P. protegens Pf-5, the transfer of an inducible cluster from Pseudomonas stutzeri and Azotobacter vinelandii yields ammonium tolerance and higher oxygen tolerance of nitrogenase activity than that from K. oxytoca. Collectively, the data from the transfer of 12 nif gene clusters between 15 diverse species (including Escherichia coli and 12 rhizobia) help identify the barriers that must be overcome to engineer a bacterium to deliver a high nitrogen flux to a cereal crop.

PMID:
31844298
DOI:
10.1038/s41564-019-0631-2

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