Multiple sensors provide spatiotemporal oxygen regulation of gene expression in a Rhizobium-legume symbiosis

• Published in: PLoS genetics 2021-02, Vol.17 (2), p.e1009099-e1009099
• Main Authors: Rutten, Paul J, Steel, Harrison, Hood, Graham A, Ramachandran, Vinoy K, McMurtry, Lucie, Geddes, Barney, Papachristodoulou, Antonis, Poole, Philip S
• Format: Article
• Language: English
• Subjects: Bacteria; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacteroids; Biology and Life Sciences; Chromosomes; E coli; Fabaceae - genetics; Fabaceae - metabolism; Gene expression; Gene Expression Regulation, Bacterial - genetics; Histidine Kinase - genetics; Histidine Kinase - metabolism; Host plants; Legumes; Mutation; Nitrogen; Nitrogen fixation; Nitrogen Fixation - genetics; Operon - genetics; Oxygen; Oxygen - metabolism; Physical Sciences; Proteins; Research and Analysis Methods; Rhizobium leguminosarum - genetics; Rhizobium leguminosarum - metabolism; Root nodules; Sensors; Symbiosis; Symbiosis - genetics; Transcription factors; Transcription Factors - genetics; Transcription Factors - metabolism
• ISSN: 1553-7404; 1553-7390; 1553-7404
• DOI: 10.1371/JOURNAL.PGEN.1009099

Regulation by oxygen (O2) in rhizobia is essential for their symbioses with plants and involves multiple O2 sensing proteins. Three sensors exist in the pea microsymbiont Rhizobium leguminosarum Rlv3841: hFixL, FnrN and NifA. At low O2 concentrations (1%) hFixL signals via FxkR to induce expression of the FixK transcription factor, which activates transcription of downstream genes. These include fixNOQP, encoding the high-affinity cbb3-type terminal oxidase used in symbiosis. In free-living Rlv3841, the hFixL-FxkR-FixK pathway was active at 1% O2, and confocal microscopy showed hFixL-FxkR-FixK activity in the earliest stages of Rlv3841 differentiation in nodules (zones I and II). Work on Rlv3841 inside and outside nodules showed that the hFixL-FxkR-FixK pathway also induces transcription of fnrN at 1% O2 and in the earliest stages of Rlv3841 differentiation in nodules. We confirmed past findings suggesting a role for FnrN in fixNOQP expression. However, unlike hFixL-FxkR-FixK, Rlv3841 FnrN was only active in the near-anaerobic zones III and IV of pea nodules. Quantification of fixNOQP expression in nodules showed this was driven primarily by FnrN, with minimal direct hFixL-FxkR-FixK induction. Thus, FnrN is key for full symbiotic expression of fixNOQP. Without FnrN, nitrogen fixation was reduced by 85% in Rlv3841, while eliminating hFixL only reduced fixation by 25%. The hFixL-FxkR-FixK pathway effectively primes the O2 response by increasing fnrN expression in early differentiation (zones I-II). In zone III of mature nodules, near-anaerobic conditions activate FnrN, which induces fixNOQP transcription to the level required for wild-type nitrogen fixation activity. Modelling and transcriptional analysis indicates that the different O2 sensitivities of hFixL and FnrN lead to a nuanced spatiotemporal pattern of gene regulation in different nodule zones in response to changing O2 concentration. Multi-sensor O2 regulation is prevalent in rhizobia, suggesting the fine-tuned control this enables is common and maximizes the effectiveness of the symbioses.