scholarly journals NAD(P)+-Malic Enzyme Mutants of Sinorhizobium sp. Strain NGR234, but Not Azorhizobium caulinodans ORS571, Maintain Symbiotic N2Fixation Capabilities

2012 ◽  
Vol 78 (8) ◽  
pp. 2803-2812 ◽  
Author(s):  
Ye Zhang ◽  
Toshihiro Aono ◽  
Phillip Poole ◽  
Turlough M. Finan
Keyword(s):  
Malic Enzyme ◽  
Sinorhizobium Meliloti ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Rhizobium Leguminosarum ◽  
Dicarboxylic Acids ◽  
Dry Weight ◽  
Tca Cycle ◽  
Broad Host Range ◽  
Azorhizobium Caulinodans ◽  
Content Type

ABSTRACTC4-dicarboxylic acids appear to be metabolized via the tricarboxylic acid (TCA) cycle in N2-fixing bacteria (bacteroids) within legume nodules. InSinorhizobium melilotibacteroids from alfalfa, NAD+-malic enzyme (DME) is required for N2fixation, and this activity is thought to be required for the anaplerotic synthesis of pyruvate. In contrast, in the pea symbiontRhizobium leguminosarum, pyruvate synthesis occurs via either DME or a pathway catalyzed by phosphoenolpyruvate carboxykinase (PCK) and pyruvate kinase (PYK). Here we report thatdmemutants of the broad-host-rangeSinorhizobiumsp. strain NGR234 formed nodules whose level of N2fixation varied from 27 to 83% (plant dry weight) of the wild-type level, depending on the host plant inoculated. NGR234 bacteroids had significant PCK activity, and while singlepckAand singledmemutants fixed N2at reduced rates, apckA dmedouble mutant had no N2-fixing activity (Fix−). Thus, NGR234 bacteroids appear to synthesize pyruvate from TCA cycle intermediates via DME or PCK pathways. These NGR234 data, together with other reports, suggested that the completely Fix−phenotype ofS. meliloti dmemutants may be specific to the alfalfa-S. melilotisymbiosis. We therefore examined the ME-like genesazc3656andazc0119fromAzorhizobium caulinodans, asazc3656mutants were previously shown to form Fix−nodules on the tropical legumeSesbania rostrata. We found that purified AZC3656 protein is an NAD(P)+-malic enzyme whose activity is inhibited by acetyl-coenzyme A (acetyl-CoA) and stimulated by succinate and fumarate. Thus, whereas DME is required for symbiotic N2fixation inA. caulinodansandS. meliloti, in other rhizobia this activity can be bypassed via another pathway(s).

scholarly journals Pyruvate Is Synthesized by Two Pathways in Pea Bacteroids with Different Efficiencies for Nitrogen Fixation

2010 ◽  
Vol 192 (19) ◽  
pp. 4944-4953 ◽  
Author(s):  
Geraldine Mulley ◽  
Miguel Lopez-Gomez ◽  
Ye Zhang ◽  
Jason Terpolilli ◽  
Jurgen Prell ◽  
...  
Keyword(s):  
Nitrogen Fixation ◽  
Malic Enzyme ◽  
Rhizobium Leguminosarum ◽  
Reduction Rate ◽  
Dicarboxylic Acids ◽  
Dry Weight ◽  
Wild Type ◽  
Type Rate ◽  
Pep Carboxykinase ◽  
Wild Type Rate

ABSTRACT Nitrogen fixation in legume bacteroids is energized by the metabolism of dicarboxylic acids, which requires their oxidation to both oxaloacetate and pyruvate. In alfalfa bacteroids, production of pyruvate requires NAD+ malic enzyme (Dme) but not NADP+ malic enzyme (Tme). However, we show that Rhizobium leguminosarum has two pathways for pyruvate formation from dicarboxylates catalyzed by Dme and by the combined activities of phosphoenolpyruvate (PEP) carboxykinase (PckA) and pyruvate kinase (PykA). Both pathways enable N2 fixation, but the PckA/PykA pathway supports N2 fixation at only 60% of that for Dme. Double mutants of dme and pckA/pykA did not fix N2. Furthermore, dme pykA double mutants did not grow on dicarboxylates, showing that they are the only pathways for the production of pyruvate from dicarboxylates normally expressed. PckA is not expressed in alfalfa bacteroids, resulting in an obligate requirement for Dme for pyruvate formation and N2 fixation. When PckA was expressed from a constitutive nptII promoter in alfalfa dme bacteroids, acetylene was reduced at 30% of the wild-type rate, although this level was insufficient to prevent nitrogen starvation. Dme has N-terminal, malic enzyme (Me), and C-terminal phosphotransacetylase (Pta) domains. Deleting the Pta domain increased the peak acetylene reduction rate in 4-week-old pea plants to 140 to 150% of the wild-type rate, and this was accompanied by increased nodule mass. Plants infected with Pta deletion mutants did not have increased dry weight, demonstrating that there is not a sustained change in nitrogen fixation throughout growth. This indicates a complex relationship between pyruvate synthesis in bacteroids, nitrogen fixation, and plant growth.

scholarly journals Genetic Analysis of Mutations Affecting pckA Regulation in Rhizobium (Sinorhizobium) meliloti

Genetics ◽  
1997 ◽  
Vol 147 (4) ◽  
pp. 1521-1531 ◽  
Author(s):  
Magne Østerås ◽  
Shelley A P O'Brien ◽  
Turlough M Finan
Keyword(s):  
Sinorhizobium Meliloti ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Root Nodules ◽  
Genetic Regulation ◽  
Carbon Sources ◽  
Dicarboxylic Acids ◽  
Phenotypic Analysis ◽  
Gene Encoding ◽  
Type Locus ◽  
Rapid Induction

Abstract The enzyme phosphoenolpyruvate carboxykinase (Pck) catalyzes the first step in the gluconeogenic pathway in most organisms. We are examining the genetic regulation of the gene encoding Pck, pckA, in Rhizobium (now Sinorhizobium) meliloti. This bacterium forms N2-fixing root nodules on alfalfa, and the major energy sources supplied to the bacteria within these nodules are C4-dicarboxylic acids such as malate and succinate. R. meliloti cells growing in glucose minimal medium show very low pckA expression whereas addition of succinate to this medium results in a rapid induction of pckA transcription. We identified spontaneous mutations (rpk) that alter the regulation of pckA expression such that pckA is expressed in media containing the non-inducing carbon sources lactose and glucose. Genetic and phenotypic analysis allowed us to differentiate at least four rpk mutant classes that map to different locations on the R. meliloti chromosome. The wild-type locus corresponding to one of these rpk loci was cloned by complementation, and two Tn5 insertions within the insert DNA that no longer complemented the rpk mutation were identified. The nucleotide sequence of this region revealed that both Tn5 insertions lay within a gene encoding a protein homologous to the Ga1R/LacI family of transcriptional regulators that are involved in metabolism.

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 Lipogenesis and Redox Balance in Nitrogen-Fixing Pea Bacteroids

2016 ◽  
Vol 198 (20) ◽  
pp. 2864-2875 ◽  
Author(s):  
Jason J. Terpolilli ◽  
Shyam K. Masakapalli ◽  
Ramakrishnan Karunakaran ◽  
Isabel U. C. Webb ◽  
Rob Green ◽  
...  
Keyword(s):  
Direct Reduction ◽  
Root Nodules ◽  
Dicarboxylic Acids ◽  
Tca Cycle ◽  
Redox Potentials ◽  
Acetyl Coa ◽  
Content Type ◽  
The Tca Cycle ◽  
Redox Poise ◽  
Phb Synthase

ABSTRACTWithin legume root nodules, rhizobia differentiate into bacteroids that oxidize host-derived dicarboxylic acids, which is assumed to occur via the tricarboxylic acid (TCA) cycle to generate NAD(P)H for reduction of N2. Metabolic flux analysis of laboratory-grownRhizobium leguminosarumshowed that the flux from [13C]succinate was consistent with respiration of an obligate aerobe growing on a TCA cycle intermediate as the sole carbon source. However, the instability of fragile pea bacteroids prevented their steady-state labeling under N2-fixing conditions. Therefore, comparative metabolomic profiling was used to compare free-livingR. leguminosarumwith pea bacteroids. While the TCA cycle was shown to be essential for maximal rates of N2fixation, levels of pyruvate (5.5-fold reduced), acetyl coenzyme A (acetyl-CoA; 50-fold reduced), free coenzyme A (33-fold reduced), and citrate (4.5-fold reduced) were much lower in bacteroids. Instead of completely oxidizing acetyl-CoA, pea bacteroids channel it into both lipid and the lipid-like polymer poly-β-hydroxybutyrate (PHB), the latter via a type III PHB synthase that is active only in bacteroids. Lipogenesis may be a fundamental requirement of the redox poise of electron donation to N2in all legume nodules. Direct reduction by NAD(P)H of the likely electron donors for nitrogenase, such as ferredoxin, is inconsistent with their redox potentials. Instead, bacteroids must balance the production of NAD(P)H from oxidation of acetyl-CoA in the TCA cycle with its storage in PHB and lipids.IMPORTANCEBiological nitrogen fixation by symbiotic bacteria (rhizobia) in legume root nodules is an energy-expensive process. Within legume root nodules, rhizobia differentiate into bacteroids that oxidize host-derived dicarboxylic acids, which is assumed to occur via the TCA cycle to generate NAD(P)H for reduction of N2. However, direct reduction of the likely electron donors for nitrogenase, such as ferredoxin, is inconsistent with their redox potentials. Instead, bacteroids must balance oxidation of plant-derived dicarboxylates in the TCA cycle with lipid synthesis. Pea bacteroids channel acetyl-CoA into both lipid and the lipid-like polymer poly-β-hydroxybutyrate, the latter via a type II PHB synthase. Lipogenesis is likely to be a fundamental requirement of the redox poise of electron donation to N2in all legume nodules.

scholarly journals Insertion of Transposon Tn5tac1 in the Sinorhizobium meliloti Malate Dehydrogenase (mdh) Gene Results in Conditional Polar Effects on Downstream TCA Cycle Genes

2004 ◽  
Vol 17 (12) ◽  
pp. 1318-1327 ◽  
Author(s):  
Sergiy l. Dymov ◽  
David J. J. Meek ◽  
Blaire Steven ◽  
Brian T. Driscoll
Keyword(s):  
Malate Dehydrogenase ◽  
Sinorhizobium Meliloti ◽  
Tricarboxylic Acid Cycle ◽  
Root Nodules ◽  
Carbon Sources ◽  
Dry Weight ◽  
Tca Cycle ◽  
Growth Conditions ◽  
Oxoglutarate Dehydrogenase ◽  
Succinyl Coa Synthetase

To isolate Sinorhizobium meliloti mutants deficient in malate dehydrogenase (MDH) activity, random transposon Tn5tac1 insertion mutants were screened for conditional lethal phenotypes on complex medium. Tn5tac1 has an outward-oriented isopropyl-β-D-thiogalactopyranoside (IPTG)- inducible promoter (Ptac). The insertion in strain Rm30049 was mapped to the mdh gene, which was found to lie directly upstream of the genes encoding succinyl-CoA synthetase (sucCD) and 2-oxoglutarate dehydrogenase (sucAB and lpdA). Rm30049 required IPTG for wild-type growth in complex media, and had a complex growth phenotype in minimal media with different carbon sources. The mdh∷ Tn5tac1 insertion eliminated MDH activity under all growth conditions, and activities of succinyl-CoA synthetase, 2-oxoglutarate dehydrogenase, and succinate dehydrogenase were affected by the addition of IPTG. Reverse-transcriptase polymerase chain reaction (RT-PCR) studies confirmed that expression from Ptac was induced by IPTG and leaky in its absence. Alfalfa plants inoculated with Rm30049 were chlorotic and stunted, with small white root nodules, and had shoot dry weight and percent-N content values similar to those of uninoculated plants. Cosmid clone pDS15 restored MDH activity to Rm30049, complemented both the mutant growth and symbiotic phenotypes, and was found to carry six complete (sdhB, mdh, sucCDAB) and two partial (lpdA, sdhA) tricarboxylic acid cycle genes.

scholarly journals The Sinorhizobium melilotintrXGene Is Involved in Succinoglycan Production, Motility, and Symbiotic Nodulation on Alfalfa

2013 ◽  
Vol 79 (23) ◽  
pp. 7150-7159 ◽  
Author(s):  
Dong Wang ◽  
Haiying Xue ◽  
Yiwen Wang ◽  
Ruochun Yin ◽  
Fang Xie ◽  
...  
Keyword(s):  
Sinorhizobium Meliloti ◽  
Nitrogen Starvation ◽  
Response Regulator ◽  
Insertion Mutant ◽  
Sesbania Rostrata ◽  
Symbiotic Relationship ◽  
Azorhizobium Caulinodans ◽  
Free Living ◽  
Content Type ◽  
Regulatory Systems

ABSTRACTRhizobia establish a symbiotic relationship with their host legumes to induce the formation of nitrogen-fixing nodules. This process is regulated by many rhizobium regulators, including some two-component regulatory systems (TCSs). NtrY/NtrX, a TCS that was first identified inAzorhizobium caulinodans, is required for free-living nitrogen metabolism and symbiotic nodulation onSesbania rostrata. However, its functions in a typical rhizobium such asSinorhizobium melilotiremain unclear. Here we found that theS. melilotiresponse regulator NtrX but not the histidine kinase NtrY is involved in the regulation of exopolysaccharide production, motility, and symbiosis with alfalfa. A plasmid insertion mutant ofntrXformed mucous colonies, which overproduced succinoglycan, an exopolysaccharide, by upregulating its biosynthesis genes. This mutant also exhibited motility defects due to reduced flagella and decreased expression of flagellins and regulatory genes. The regulation is independent of the known regulatory systems of ExoR/ExoS/ChvI, EmmABC, and ExpR. Alfalfa plants inoculated with thentrXmutant were small and displayed symptoms of nitrogen starvation. Interestingly, the deletion mutant ofntrYshowed a phenotype similar to that of the parent strain. These findings demonstrate that theS. melilotiNtrX is a new regulator of succinoglycan production and motility that is not genetically coupled with NtrY.

scholarly journals Polyamines produced by Sinorhizobium meliloti Rm8530 contribute to symbiotically relevant phenotypes ex planta and to nodulation efficiency on alfalfa

Microbiology ◽  
2020 ◽  
Vol 166 (3) ◽  
pp. 278-287 ◽  
Author(s):  
Victor A. Becerra-Rivera ◽  
Alejandra Arteaga ◽  
Alfonso Leija ◽  
Georgina Hernández ◽  
Michael F. Dunn
Keyword(s):  
Oxidative Stress ◽  
Sinorhizobium Meliloti ◽  
Dry Weight ◽  
Nitrogen Fixing ◽  
Wild Type ◽  
Content Type ◽  
Phenotypic Differences ◽  
Nodule Biomass ◽  
Increased Sensitivity ◽  
Exogenous Spermidine

In nitrogen-fixing rhizobia, emerging evidence shows significant roles for polyamines in growth and abiotic stress resistance. In this work we show that a polyamine-deficient ornithine decarboxylase null mutant (odc2) derived from Sinorhizobium meliloti Rm8530 had significant phenotypic differences from the wild-type, including greatly reduced production of exopolysaccharides (EPS; ostensibly both succinoglycan and galactoglucan), increased sensitivity to oxidative stress and decreased swimming motility. The introduction of the odc2 gene borne on a plasmid into the odc2 mutant restored wild-type phenotypes for EPS production, growth under oxidative stress and swimming. The production of calcofluor-binding EPS (succinoglycan) by the odc2 mutant was also completely or mostly restored in the presence of exogenous spermidine (Spd), norspermidine (NSpd) or spermine (Spm). The odc2 mutant formed about 25 % more biofilm than the wild-type, and its ability to form biofilm was significantly inhibited by exogenous Spd, NSpd or Spm. The odc2 mutant formed a less efficient symbiosis with alfalfa, resulting in plants with significantly less biomass and height, more nodules but less nodule biomass, and 25 % less nitrogen-fixing activity. Exogenously supplied Put was not able to revert these phenotypes and caused a similar increase in plant height and dry weight in uninoculated plants and in those inoculated with the wild-type or odc2 mutant. We discuss ways in which polyamines might affect the phenotypes of the odc2 mutant.

scholarly journals AcheZ-Like Gene inAzorhizobium caulinodansIs a Key Gene in the Control of Chemotaxis and Colonization of the Host Plant

2017 ◽  
Vol 84 (3) ◽  
Author(s):  
Xiaolin Liu ◽  
Wei Liu ◽  
Yu Sun ◽  
Chunlei Xia ◽  
Claudine Elmerich ◽  
...  
Keyword(s):  
Host Plant ◽  
Active Site ◽  
Deletion Mutant ◽  
Sinorhizobium Meliloti ◽  
Response Regulator ◽  
General Situation ◽  
Symbiotic Association ◽  
Azorhizobium Caulinodans ◽  
Content Type ◽  
In The Wild

ABSTRACTChemotaxis can provide bacteria with competitive advantages for survival in complex environments. The CheZ chemotaxis protein is a phosphatase, affecting the flagellar motor inEscherichia coliby dephosphorylating the response regulator phosphorylated CheY protein (CheY∼P) responsible for clockwise rotation. AcheZgene has been found inAzorhizobium caulinodansORS571, in contrast to other rhizobial species studied so far. The CheZ protein in strain ORS571 has a conserved motif similar to that corresponding to the phosphatase active site inE. coli. The construction of acheZdeletion mutant strain and ofcheZmutant strains carrying a mutation in residues of the putative phosphatase active site showed that strain ORS571 participates in chemotaxis and motility, causing a hyperreversal behavior. In addition, the properties of thecheZdeletion mutant revealed that ORS571 CheZ is involved in other physiological processes, since it displayed increased flocculation, biofilm formation, exopolysaccharide (EPS) production, and host root colonization. In particular, it was observed that the expression of severalexpgenes, involved in EPS synthesis, was upregulated in thecheZmutant compared to that in the wild type, suggesting that CheZ negatively controlsexpgene expression through an unknown mechanism. It is proposed that CheZ influences theAzorhizobium-plant association by negatively regulating early colonization via the regulation of EPS production. This report established that CheZ inA. caulinodansplays roles in chemotaxis and the symbiotic association with the host plant.IMPORTANCEChemotaxis allows bacteria to swim toward plant roots and is beneficial to the establishment of various plant-microbe associations. The level of CheY phosphorylation (CheY∼P) is central to the chemotaxis signal transduction. The mechanism of the signal termination of CheY∼P remains poorly characterized amongAlphaproteobacteria, except forSinorhizobium meliloti, which does not contain CheZ but which controls CheY∼P dephosphorylation through a phosphate sink mechanism.Azorhizobium caulinodansORS571, a microsymbiont ofSesbania rostrata, has an orphancheZgene besides twocheYgenes similar to those inS. meliloti. In addition to controlling the chemotaxis response, the CheZ-like protein in strain ORS571 is playing a role by decreasing bacterial adhesion to the host plant, in contrast to the general situation where chemotaxis-associated proteins promote adhesion. In this study, we identified a CheZ-like protein amongAlphaproteobacteriafunctioning in chemotaxis and theA. caulinodans-S. rostratasymbiosis.

scholarly journals Role of O2 in the Growth of Rhizobium leguminosarum bv. viciae 3841 on Glucose and Succinate

2016 ◽  
Vol 199 (1) ◽  
Author(s):  
Rachel M. Wheatley ◽  
Vinoy K. Ramachandran ◽  
Barney A. Geddes ◽  
Benjamin J. Perry ◽  
Chris K. Yost ◽  
...  
Keyword(s):  
Rhizobium Leguminosarum ◽  
Regulatory Protein ◽  
Carbon Sources ◽  
Cytosine Deaminase ◽  
Optimal Growth ◽  
Tca Cycle ◽  
Cell Physiology ◽  
Type I ◽  
Content Type ◽  
Glutamate Synthesis

ABSTRACT Insertion sequencing (INSeq) analysis of Rhizobium leguminosarum bv. viciae 3841 (Rlv3841) grown on glucose or succinate at both 21% and 1% O2 was used to understand how O2 concentration alters metabolism. Two transcriptional regulators were required for growth on glucose (pRL120207 [eryD] and RL0547 [phoB]), five were required on succinate (pRL100388, RL1641, RL1642, RL3427, and RL4524 [ecfL]), and three were required on 1% O2 (pRL110072, RL0545 [phoU], and RL4042). A novel toxin-antitoxin system was identified that could be important for generation of new plasmidless rhizobial strains. Rlv3841 appears to use the methylglyoxal pathway alongside the Entner-Doudoroff (ED) pathway and tricarboxylic acid (TCA) cycle for optimal growth on glucose. Surprisingly, the ED pathway was required for growth on succinate, suggesting that sugars made by gluconeogenesis must undergo recycling. Altered amino acid metabolism was specifically needed for growth on glucose, including RL2082 (gatB) and pRL120419 (opaA, encoding omega-amino acid:pyruvate transaminase). Growth on succinate specifically required enzymes of nucleobase synthesis, including ribose-phosphate pyrophosphokinase (RL3468 [prs]) and a cytosine deaminase (pRL90208 [codA]). Succinate growth was particularly dependent on cell surface factors, including the PrsD-PrsE type I secretion system and UDP-galactose production. Only RL2393 (glnB, encoding nitrogen regulatory protein PII) was specifically essential for growth on succinate at 1% O2, conditions similar to those experienced by N2-fixing bacteroids. Glutamate synthesis is constitutively activated in glnB mutants, suggesting that consumption of 2-ketoglutarate may increase flux through the TCA cycle, leading to excess reductant that cannot be reoxidized at 1% O2 and cell death. IMPORTANCE Rhizobium leguminosarum, a soil bacterium that forms N2-fixing symbioses with several agriculturally important leguminous plants (including pea, vetch, and lentil), has been widely utilized as a model to study Rhizobium-legume symbioses. Insertion sequencing (INSeq) has been used to identify factors needed for its growth on different carbon sources and O2 levels. Identification of these factors is fundamental to a better understanding of the cell physiology and core metabolism of this bacterium, which adapts to a variety of different carbon sources and O2 tensions during growth in soil and N2 fixation in symbiosis with legumes.

scholarly journals Mutation in the ntrR Gene, a Member of the vap Gene Family, Increases the Symbiotic Efficiency of Sinorhizobium meliloti

2001 ◽  
Vol 14 (7) ◽  
pp. 887-894 ◽  
Author(s):  
Boglárka Oláh ◽  
Erno Kiss ◽  
Zoltán Györgypál ◽  
Judit Borzi ◽  
Gyöngyi Cinege ◽  
...  
Keyword(s):  
Nitrogen Fixation ◽  
Pathogenic Bacteria ◽  
Sinorhizobium Meliloti ◽  
Root Nodules ◽  
Nifh Gene ◽  
Dry Weight ◽  
Nodule Occupancy ◽  
Gene Encoding ◽  
Symbiotic Efficiency ◽  
Host Interactions

In specific plant organs, namely the root nodules of alfalfa, fixed nitrogen (ammonia) produced by the symbiotic partner Sinorhizobium meliloti supports the growth of the host plant in nitrogen-depleted environment. Here, we report that a derivative of S. meliloti carrying a mutation in the chromosomal ntrR gene induced nodules with enhanced nitrogen fixation capacity, resulting in an increased dry weight and nitrogen content of alfalfa. The efficient nitrogen fixation is a result of the higher expression level of the nifH gene, encoding one of the subunits of the nitrogenase enzyme, and nifA, the transcriptional regulator of the nif operon. The ntrR gene, controlled negatively by its own product and positively by the symbiotic regulator syrM, is expressed in the same zone of nodules as the nif genes. As a result of the nitrogen-tolerant phenotype of the strain, the beneficial effect of the mutation on efficiency is not abolished in the presence of the exogenous nitrogen source. The ntrR mutant is highly competitive in nodule occupancy compared with the wild-type strain. Sequence analysis of the mutant region revealed a new cluster of genes, termed the “ntrPR operon,” which is highly homologous to a group of vap-related genes of various pathogenic bacteria that are presumably implicated in bacterium-host interactions. On the basis of its favorable properties, the strain is a good candidate for future agricultural utilization.

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