Thesis
Role of FnrN and FixLJ-FixK in regulating nitrogen fixation in the Mesorhizobium ciceri CC1192-Cicer arietinum symbiosis
Honours, Murdoch University
2023
Abstract
Rhizobia fix N2 within legume root nodules in microaerobic conditions, with low O2 a key signal for expression of the nitrogenase enzyme complex. In Mesorhizobium japonicum R7A, which forms a symbiosis with Lotus corniculatus, the novel LacI/GalR-type regulator FixV is essential to N2 fixation and controls symbiosis gene expression, even though FixV lacks an O2-binding heme group or obvious O2 sensing mechanism. However, in M. ciceri CC1192 which is a microsymbiont of Cicer arietinum, deletion of fixV reduced, but did not abolish nitrogenase activity, indicating some other regulator must controlN2 fixation in this strain. Recently, the O2-sensing regulator FnrN was found to control N2 fixation in Rhizobium spp., and this gene is present in CC1192. To investigate whether fnrN is involved in regulating N2 fixation in CC1192, a vector for site-directed marker-exchange mutagenesis was constructed and an ΔfnrN mutant isolated and confirmed by Illumina whole genome sequencing. CC1192 also harbours a second potential O2 sensor-regulator system (fixLJ-fixK) on its 648-kb repABC-type plasmid pMc1192, so a curing vector was constructed and pMc1192 cured separately from both ΔfnrN and ΔfixV mutant backgrounds. In glasshouse trials with C. arietinum, nitrogenase activity of ΔfixV and ΔfixV plasmid-cured strains was reduced to 25% of the wild-type rate irrespective of whether the plasmid was present, indicating that fixLJ-fixK or other pMc1192 encoded genes were not essential to N2 fixation. In contrast, nitrogenase activity was reduced to 11% for ΔfnrN mutant and less than 1% for the plasmid cured strain relative to the wild-type rate, suggesting a synergistic effect may exist between FnrN and the FixLJ-FixK system. These results demonstrate that FnrN plays a key role in the O2 regulation of N2 fixation in the Mesorhizobium ciceri CC1192-Cicer arietinum symbiosis.
Details
- Title
- Role of FnrN and FixLJ-FixK in regulating nitrogen fixation in the Mesorhizobium ciceri CC1192-Cicer arietinum symbiosis
- Authors/Creators
- Marshall Tye
- Contributors
- Jason Terpolilli (Supervisor) - Murdoch University, Centre for Sustainable Farming SystemsGraham O'Hara (Supervisor) - Murdoch University, Centre for Sustainable Farming Systems
- Awarding Institution
- Murdoch University; Honours
- Identifiers
- 991005597069607891
- Murdoch Affiliation
- School of Medical, Molecular and Forensic Sciences
- Resource Type
- Thesis
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