scholarly journals Sinorhizobium fredii HH103 RirA Is Required for Oxidative Stress Resistance and Efficient Symbiosis with Soybean

2019 ◽  
Vol 20 (3) ◽  
pp. 787 ◽  
Author(s):  
Juan Crespo-Rivas ◽  
Pilar Navarro-Gómez ◽  
Cynthia Alias-Villegas ◽  
Jie Shi ◽  
Tao Zhen ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Nitrogen Fixation ◽  
Rhizobium Leguminosarum ◽  
Symbiotic Nitrogen Fixation ◽  
Regulatory Protein ◽  
No Production ◽  
Broad Host Range ◽  
Wild Type ◽  
Iron Deficient ◽  
Soybean Plants

Members of Rhizobiaceae contain a homologue of the iron-responsive regulatory protein RirA. In different bacteria, RirA acts as a repressor of iron uptake systems under iron-replete conditions and contributes to ameliorate cell damage during oxidative stress. In Rhizobium leguminosarum and Sinorhizobium meliloti, mutations in rirA do not impair symbiotic nitrogen fixation. In this study, a rirA mutant of broad host range S. fredii HH103 has been constructed (SVQ780) and its free-living and symbiotic phenotypes evaluated. No production of siderophores could be detected in either the wild-type or SVQ780. The rirA mutant exhibited a growth advantage under iron-deficient conditions and hypersensitivity to hydrogen peroxide in iron-rich medium. Transcription of rirA in HH103 is subject to autoregulation and inactivation of the gene upregulates fbpA, a gene putatively involved in iron transport. The S. fredii rirA mutant was able to nodulate soybean plants, but symbiotic nitrogen fixation was impaired. Nodules induced by the mutant were poorly infected compared to those induced by the wild-type. Genetic complementation reversed the mutant’s hypersensitivity to H2O2, expression of fbpA, and symbiotic deficiency in soybean plants. This is the first report that demonstrates a role for RirA in the Rhizobium-legume symbiosis.

scholarly journals Antioxidant ability of glutaredoxins and their role in symbiotic nitrogen fixation in Rhizobium leguminosarum bv. viciae 3841

2020 ◽  
Author(s):  
Qian Zou ◽  
Yanlin Zhou ◽  
Guojun Cheng ◽  
Yang Peng ◽  
Sha Luo ◽  
...  
Keyword(s):  
Nitrogen Fixation ◽  
Quantitative Proteomics ◽  
Rhizobium Leguminosarum ◽  
Symbiotic Nitrogen Fixation ◽  
Proteome Analysis ◽  
Wild Type ◽  
Triple Mutant ◽  
Antioxidant Proteins ◽  
Mixed Disulfides ◽  
Nodule Symbiosis

Glutaredoxins (Grx) are redoxin family proteins that reduce disulfides and mixed disulfides between glutathione and proteins. Rhizobium leguminosarum bv. Viciae 3841 contains three genes coding for glutaredoxins: RL4289 (grxA) codes for a dithiolic glutaredoxin, RL2615 (grxB) codes for a monothiol glutaredoxin, while RL4261 (grxC) codes for a glutaredoxin-like NrdH protein. We generated mutants interrupted in one, two, or three glutaredoxin genes. These mutants had no obvious differences in growth phenotypes from the wild type RL3841. However, while a mutant of grxC did not affect the antioxidant or symbiotic capacities of R. leguminosarum, grxA-derived or grxB mutants decreased antioxidant and nitrogen fixation capacities. Furthermore, grxA mutants were severely impaired in rhizosphere colonization, and formed smaller nodules with defects of bacteroid differentiation, whereas nodules induced by grxB mutants contained abnormally thick cortices and prematurely senescent bacteroids. The grx triple mutant had the greatest defect in antioxidant and symbiotic capacities of R. leguminosarum and quantitative proteomics revealed it had 56 up-regulated and 81 down-regulated proteins relative to wildtype. Of these proteins, twenty-eight are involved in transporter activity, twenty are related to stress response and virulence, and sixteen are involved in amino acid metabolism. Overall, R. leguminosarum glutaredoxins behave as antioxidant proteins mediating root nodule symbiosis. IMPORTANCE Glutaredoxin catalyzes glutathionylation/deglutathionylation reactions, protects SH-groups from oxidation and restores functionally active thiols. Three glutaredoxins exist in R. leguminosarum and their properties were investigated in free-living bacteria and during nitrogen-fixing symbiosis. All the glutaredoxins were necessary for oxidative stress defense. Dithiol GrxA affects nodulation and nitrogen fixation of bacteroids by altering deglutathionylation reactions, monothiol GrxB is involved in symbiotic nitrogen fixation by regulating Fe-S cluster biogenesis, and GrxC may participate in symbiosis by an unknown mechanism. Proteome analysis provides clues to explain the differences between the grx triple mutant and wild-type nodules.

scholarly journals Succinate Transport Is Not Essential for Symbiotic Nitrogen Fixation by Sinorhizobium meliloti or Rhizobium leguminosarum

2017 ◽  
Vol 84 (1) ◽  
Author(s):  
Michael J. Mitsch ◽  
George C. diCenzo ◽  
Alison Cowie ◽  
Turlough M. Finan
Keyword(s):  
Nitrogen Fixation ◽  
Carbon Source ◽  
Sinorhizobium Meliloti ◽  
Rhizobium Leguminosarum ◽  
Symbiotic Nitrogen Fixation ◽  
Sequencing Analysis ◽  
Wild Type ◽  
Content Type ◽  
Transcriptional Changes ◽  
Symbiotic Properties

ABSTRACTSymbiotic nitrogen fixation (SNF) is an energetically expensive process performed by bacteria during endosymbiotic relationships with plants. The bacteria require the plant to provide a carbon source for the generation of reductant to power SNF. While C4-dicarboxylates (succinate, fumarate, and malate) appear to be the primary, if not sole, carbon source provided to the bacteria, the contribution of each C4-dicarboxylate is not known. We address this issue using genetic and systems-level analyses. Expression of a malate-specific transporter (MaeP) inSinorhizobium melilotiRm1021dctmutants unable to transport C4-dicarboxylates resulted in malate import rates of up to 30% that of the wild type. This was sufficient to support SNF withMedicago sativa, with acetylene reduction rates of up to 50% those of plants inoculated with wild-typeS. meliloti.Rhizobium leguminosarumbv. viciae 3841dctmutants unable to transport C4-dicarboxylates but expressing themaePtransporter had strong symbiotic properties, withPisum sativumplants inoculated with these strains appearing similar to plants inoculated with wild-typeR. leguminosarum. This was despite malate transport rates by the mutant bacteroids being 10% those of the wild type. An RNA-sequencing analysis of the combinedP. sativum-R. leguminosarumnodule transcriptome was performed to identify systems-level adaptations in response to the inability of the bacteria to import succinate or fumarate. Few transcriptional changes, with no obvious pattern, were detected. Overall, these data illustrated that succinate and fumarate are not essential for SNF and that, at least in specific symbioses,l-malate is likely the primary C4-dicarboxylate provided to the bacterium.IMPORTANCESymbiotic nitrogen fixation (SNF) is an economically and ecologically important biological process that allows plants to grow in nitrogen-poor soils without the need to apply nitrogen-based fertilizers. Much research has been dedicated to this topic to understand this process and to eventually manipulate it for agricultural gains. The work presented in this article provides new insights into the metabolic integration of the plant and bacterial partners. It is shown that malate is the only carbon source that needs to be available to the bacterium to support SNF and that, at least in some symbioses, malate, and not other C4-dicarboxylates, is likely the primary carbon provided to the bacterium. This work extends our knowledge of the minimal metabolic capabilities the bacterium requires to successfully perform SNF and may be useful in further studies aiming to optimize this process through synthetic biology approaches. The work describes an engineering approach to investigate a metabolic process that occurs between a eukaryotic host and its prokaryotic endosymbiont.

scholarly journals Oxidative Stress Produced by Paraquat Reduces Nitrogen Fixation in Soybean-Bradyrhizobium diazoefficiens Symbiosis by Decreasing Nodule Functionality

Nitrogen ◽  
2021 ◽  
Vol 2 (1) ◽  
pp. 30-40
Author(s):  
Germán Tortosa ◽  
Sergio Parejo ◽  
Juan J. Cabrera ◽  
Eulogio J. Bedmar ◽  
Socorro Mesa
Keyword(s):  
Oxidative Stress ◽  
Nitrogen Fixation ◽  
Symbiotic Nitrogen Fixation ◽  
Dry Weight ◽  
Fixed Nitrogen ◽  
Biotic And Abiotic Stresses ◽  
Free Living ◽  
Soybean Plants ◽  
Soil Metal ◽  
Free Living Conditions

Soybean (Glycine max.) is one of the most important legumes cultivated worldwide. Its productivity can be altered by some biotic and abiotic stresses like global warming, soil metal pollution or over-application of herbicides like paraquat (1,1’-dimethyl-4,4’-bipyridinium dichloride). In this study, the effect of oxidative stress produced by paraquat addition (0, 20, 50 and 100 µM) during plant growth on symbiotic nitrogen fixation (SNF) and functionality of Bradyrhizobium diazoefficiens-elicited soybean nodules were evaluated. Results showed that the 50 µM was the threshold that B. diazoefficiens can tolerate under free-living conditions. In symbiosis with soybean, the paraquat addition statistically reduced the shoot and root dry weight of soybean plants, and number and development of the nodules. SNF was negatively affected by paraquat, which reduced total nitrogen content and fixed nitrogen close to 50% when 100 µM was added. These effects were due to the impairment of nodule functionality and the increased oxidative status of the nodules, as revealed by the lower leghaemoglobin content and the higher lipid peroxidation in soybean nodules from paraquat-treated plants.

scholarly journals An Alkane Sulfonate Monooxygenase Is Required for Symbiotic Nitrogen Fixation by Bradyrhizobium diazoefficiens (syn. Bradyrhizobium japonicum) USDA110T

2019 ◽  
Vol 85 (24) ◽  
Author(s):  
Justin J. Speck ◽  
Euan K. James ◽  
Masayuki Sugawara ◽  
Michael J. Sadowsky ◽  
Prasad Gyaneshwar
Keyword(s):  
Oxidative Stress ◽  
Nitrogen Fixation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Symbiotic Nitrogen Fixation ◽  
Sulfur Metabolism ◽  
Wild Type ◽  
Content Type ◽  
Organic S ◽  
Symbiotic Associations

ABSTRACT Sulfur (S)-containing molecules play an important role in symbiotic nitrogen fixation and are critical components of nitrogenase and other iron-S proteins. S deficiency inhibits symbiotic nitrogen fixation by rhizobia. However, despite its importance, little is known about the sources of S that rhizobia utilize during symbiosis. We previously showed that Bradyrhizobium diazoefficiens USDA110T can assimilate both inorganic and organic S and that genes involved in organic S utilization are expressed during symbiosis. Here, we show that a B. diazoefficiens USDA110T mutant with a sulfonate monooxygenase (ssuD) insertion is defective in nitrogen fixation. Microscopy analyses revealed that the ΔssuD mutant was defective in root hair infection and that ΔssuD mutant bacteroids showed degradation compared to the wild-type strain. Moreover, the ΔssuD mutant was significantly more sensitive to hydrogen peroxide-mediated oxidative stress than the wild-type strain. Taken together, these results show that the ability of rhizobia to utilize organic S plays an important role in symbiotic nitrogen fixation. Since nodules have been reported to be an important source of reduced S used during symbiosis and nitrogen fixation, further research will be needed to determine the mechanisms involved in the regulation of S assimilation by rhizobia. IMPORTANCE Rhizobia form symbiotic associations with legumes that lead to the formation of nitrogen-fixing nodules. Sulfur-containing molecules play a crucial role in nitrogen fixation; thus, the rhizobia inside nodules require large amounts of sulfur. Rhizobia can assimilate both inorganic (sulfate) and organic (sulfonates) sources of sulfur. However, very little is known about rhizobial sulfur metabolism during symbiosis. In this report, we show that sulfonate utilization by Bradyrhizobium diazoefficiens is important for symbiotic nitrogen fixation in both soybean and cowpea. The symbiotic defect is probably due to increased sensitivity to oxidative stress from sulfur deficiency in the mutant strain defective for sulfonate utilization. The results of this study can be extended to other rhizobium-legume symbioses, as sulfonate utilization genes are widespread in these bacteria.

scholarly journals Regulation and characterization of mutants of fixABCX in Rhizobium leguminosarum.

2021 ◽  
Author(s):  
Isabel Webb ◽  
Jiabao Xu ◽  
Carmen Sanchez-Cañizares ◽  
Ramakrishnan Karunakaran ◽  
Vinoy Ramachandran ◽  
...  
Keyword(s):  
Nitrogen Fixation ◽  
Rhizobium Leguminosarum ◽  
Redox Balance ◽  
Rna Seq ◽  
Wild Type ◽  
Transcription Start Sites ◽  
Sym Plasmid ◽  
Microaerobic Conditions ◽  
Type Infection

Symbiosis between Rhizobium leguminosarum and Pisum sativum requires tight control of redox balance in order to maintain respiration under the microaerobic conditions required for nitrogenase, whilst still producing the eight electrons and sixteen molecules of ATP needed for nitrogen fixation. FixABCX, electron transfer flavoproteins essential for nitrogen fixation, are encoded on the Sym plasmid (pRL10), immediately upstream of nifA, which encodes the general transcriptional regulator of nitrogen fixation. There is a symbiotically-regulated NifA-dependent promoter upstream of fixA (PnifA1), as well as an additional basal constitutive promoter driving background expression of nifA (PnifA2). These were confirmed by 5’-end mapping of transcription start sites using differential (d) RNA-seq. Complementation of polar fixAB and fixX mutants (Fix- strains) confirmed expression of nifA from PnifA1 in symbiosis. Electron microscopy combined with single-cell Raman microspectroscopy characterization of fixAB mutants revealed previously unknown heterogeneity in bacteroid morphology within a single nodule. Two morphotypes of mutant fixAB bacteroids were observed. One was larger than wild-type bacteroids and contained high levels of polyhydroxy-3-butyrate, a complex energy/reductant storage product. A second bacteroid phenotype was morphologically and compositionally different and resembled wild-type infection thread cells. From these two characteristic fixAB mutant bacteroid morphotypes, inferences can be drawn on the metabolism of wild-type nitrogen-fixing bacteroids.

scholarly journals Development of RR soybean in function of glyphosate doses and Bradyrhizobium inoculation

2018 ◽  
Vol 22 (12) ◽  
pp. 854-858
Author(s):  
João W. Bossolani ◽  
Nadia M. Poloni ◽  
Edson Lazarini ◽  
João V. T. Bettiol ◽  
João A. Fischer Filho ◽  
...  
Keyword(s):  
Nitrogen Fixation ◽  
Symbiotic Nitrogen Fixation ◽  
Soybean Seeds ◽  
Leaf Chlorophyll ◽  
Outdoor Experiment ◽  
Glyphosate Herbicide ◽  
Soybean Plants ◽  
Biological Nitrogen ◽  
Different Doses ◽  
Herbicide Glyphosate

ABSTRACT Soybean has traditionally been produced in systems that include the use of herbicides, often in higher than recommended doses. The process of symbiotic nitrogen fixation in legumes can be hampered by these herbicides, both by direct effects on rhizobia and indirect effects on the host plant. An outdoor experiment was performed to evaluate the effects of different doses of a glyphosate herbicide on Bradyrhizobium strains and biological nitrogen fixation in soybean BMX Potência RR plants. Soybean seeds were inoculated with Bradyrhizobium elkanii (SEMIA 5019) and Bradyrhizobium japonicum (SEMIA 5079) strains in a commercial liquid inoculant. The treatments consisted of the absence and presence of Bradyrhizobium genotypes inoculated via seed and four doses of the herbicide glyphosate applied on the leaves (0, 1.0, 2.0, and 4.0 L ha-1 of the commercial product) at the V3 stage. The leaf chlorophyll index of inoculated RR soybean plants did not change on the application of glyphosate and, regardless of inoculation, plants had the capacity to recover from the effects of glyphosate application, without impaired development.

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