Sinorhizobium meliloti lsrB is involved in alfalfa root nodule development and nitrogen-fixing bacteroid differentiation

During root nodule development several key genes involved in nitrogen fixation and assimilation exhibit enhanced levels of expression. However, little is known about the temporal and spatial distribution patterns of these transcripts. In a systematic study the transcripts for 13 of the essential enzymes involved in alfalfa (Medicago sativa) root nodule nitrogen and carbon metabolism were localized by in situ hybridization. A serial section approach allowed the construction of a map that reflects the relative distribution of these transcripts. In 33-day-old root nodules, the expression of nifH, NADH-dependent glutamate synthase (NADH-GOGAT; EC 1.4.1.14) and a cytosolic isoform of glutamine synthetase (GS13; GS; EC 6.3.1.2) were localized predominantly in a 5- to 15-cell-wide region in the distal part of the nitrogen-fixing zone. This zone was also the region of high expression for leghemoglobin, a second cytosolic glutamine synthetase isoform (GS100), aspartate aminotransferase-2 (AAT-2; EC 2.6.1.1), asparagine synthetase (AS; 6.3.5.4), phosphoenolpyruvate carboxylase (PEPC; EC 4.1.1.31), and sucrose synthase (SuSy; EC 2.4.1.13). This suggests that, in 33-day-old alfalfa root nodules, nitrogen fixation is restricted to this 5- to 15-cell-wide area. The continued significant expression of the GS100 subclass of GS and AS in the proximal part of the nitrogen-fixing zone implicates these gene products in nitrogen remobilization. A low constitutive expression of NADH-dependent glutamate dehydrogenase (NADH-GDH; EC 1.4.1.2) was observed throughout the nodule. The transcript distribution map will be used as a navigational tool to assist in developing strategies for the genetic engineering of alfalfa root nodules for enhanced nitrogen assimilation.

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Alfalfa Root Nodule Invasion Efficiency Is Dependent on Sinorhizobium melilotiPolysaccharides

Journal of Bacteriology ◽

10.1128/jb.182.15.4310-4318.2000 ◽

2000 ◽

Vol 182 (15) ◽

pp. 4310-4318 ◽

Cited By ~ 144

Author(s):

Brett J. Pellock ◽

Hai-Ping Cheng ◽

Graham C. Walker

Keyword(s):

Molecular Weight ◽

Sinorhizobium Meliloti ◽

Root Nodule ◽

Fluorescent Protein ◽

Infection Thread ◽

Low Molecular Weight ◽

Infection Threads ◽

Green Fluorescent ◽

K Antigen ◽

Alfalfa Root

ABSTRACT The soil bacterium Sinorhizobium meliloti is capable of entering into a nitrogen-fixing symbiosis with Medicago sativa (alfalfa). Particular low-molecular-weight forms of certain polysaccharides produced by S. meliloti are crucial for establishing this symbiosis. Alfalfa nodule invasion by S. meliloti can be mediated by any one of three symbiotically important polysaccharides: succinoglycan, EPS II, or K antigen (also referred to as KPS). Using green fluorescent protein-labeled S. meliloti cells, we have shown that there are significant differences in the details and efficiencies of nodule invasion mediated by these polysaccharides. Succinoglycan is highly efficient in mediating both infection thread initiation and extension. However, EPS II is significantly less efficient than succinoglycan at mediating both invasion steps, and K antigen is significantly less efficient than succinoglycan at mediating infection thread extension. In the case of EPS II-mediated symbioses, the reduction in invasion efficiency results in stunted host plant growth relative to plants inoculated with succinoglycan or K-antigen-producing strains. Additionally, EPS II- and K-antigen-mediated infection threads are 8 to 10 times more likely to have aberrant morphologies than those mediated by succinoglycan. These data have important implications for understanding how S. meliloti polysaccharides are functioning in the plant-bacterium interaction, and models are discussed.

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Involvement of Glutaredoxin and Thioredoxin Systems in the Nitrogen-Fixing Symbiosis between Legumes and Rhizobia

Antioxidants ◽

10.3390/antiox7120182 ◽

2018 ◽

Vol 7 (12) ◽

pp. 182 ◽

Cited By ~ 5

Author(s):

Geneviève Alloing ◽

Karine Mandon ◽

Eric Boncompagni ◽

Françoise Montrichard ◽

Pierre Frendo

Keyword(s):

Sinorhizobium Meliloti ◽

Root Nodule ◽

Redox Balance ◽

Atmospheric Nitrogen ◽

Nitrogen Fixing ◽

Symbiotic Interaction ◽

Bacterial Differentiation ◽

Genes Encoding ◽

Transcriptomics Data

Leguminous plants can form a symbiotic relationship with Rhizobium bacteria, during which plants provide bacteria with carbohydrates and an environment appropriate to their metabolism, in return for fixed atmospheric nitrogen. The symbiotic interaction leads to the formation of a new organ, the root nodule, where a coordinated differentiation of plant cells and bacteria occurs. The establishment and functioning of nitrogen-fixing symbiosis involves a redox control important for both the plant-bacteria crosstalk and the regulation of nodule metabolism. In this review, we discuss the involvement of thioredoxin and glutaredoxin systems in the two symbiotic partners during symbiosis. The crucial role of glutathione in redox balance and S-metabolism is presented. We also highlight the specific role of some thioredoxin and glutaredoxin systems in bacterial differentiation. Transcriptomics data concerning genes encoding components and targets of thioredoxin and glutaredoxin systems in connection with the developmental step of the nodule are also considered in the model system Medicago truncatula–Sinorhizobium meliloti.

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Sinorhizobium meliloti Mutants Deficient in Phosphatidylserine Decarboxylase Accumulate Phosphatidylserine and Are Strongly Affected during Symbiosis with Alfalfa

Journal of Bacteriology ◽

10.1128/jb.00610-08 ◽

2008 ◽

Vol 190 (20) ◽

pp. 6846-6856 ◽

Cited By ~ 14

Author(s):

Miguel Angel Vences-Guzmán ◽

Otto Geiger ◽

Christian Sohlenkamp

Keyword(s):

Sinorhizobium Meliloti ◽

Minimal Medium ◽

Root Nodule ◽

Membrane Lipids ◽

Nodule Formation ◽

Deficient Mutant ◽

Nitrogen Fixing ◽

Wild Type ◽

Root Nodule Symbiosis ◽

Nodule Symbiosis

ABSTRACT Sinorhizobium meliloti contains phosphatidylglycerol, cardiolipin, phosphatidylcholine, and phosphatidylethanolamine (PE) as major membrane lipids. PE is formed in two steps. In the first step, phosphatidylserine synthase (Pss) condenses serine with CDP-diglyceride to form phosphatidylserine (PS), and in the second step, PS is decarboxylated by phosphatidylserine decarboxylase (Psd) to form PE. In this study we identified the sinorhizobial psd gene coding for Psd. A sinorhizobial mutant deficient in psd is unable to form PE but accumulates the anionic phospholipid PS. Properties of PE-deficient mutants lacking either Pss or Psd were compared with those of the S. meliloti wild type. Whereas both PE-deficient mutants grew in a wild-type-like manner on many complex media, they were unable to grow on minimal medium containing high phosphate concentrations. Surprisingly, the psd-deficient mutant could grow on minimal medium containing low concentrations of inorganic phosphate, while the pss-deficient mutant could not. Addition of choline to the minimal medium rescued growth of the pss-deficient mutant, CS111, to some extent but inhibited growth of the psd-deficient mutant, MAV01. When the two distinct PE-deficient mutants were analyzed for their ability to form a nitrogen-fixing root nodule symbiosis with their alfalfa host plant, they behaved strikingly differently. The Pss-deficient mutant, CS111, initiated nodule formation at about the same time point as the wild type but did form about 30% fewer nodules than the wild type. In contrast, the PS-accumulating mutant, MAV01, initiated nodule formation much later than the wild type and formed 90% fewer nodules than the wild type. The few nodules formed by MAV01 seemed to be almost devoid of bacteria and were unable to fix nitrogen. Leaves of alfalfa plants inoculated with the mutant MAV01 were yellowish, indicating that the plants were starved for nitrogen. Therefore, changes in lipid composition, including the accumulation of bacterial PS, prevent the establishment of a nitrogen-fixing root nodule symbiosis.

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Synchronous expression of leghaemoglobin genes inMedicago truncatula during nitrogen-fixing root nodule development and response to exogenously supplied nitrate

Plant Molecular Biology ◽

10.1007/bf00040629 ◽

1991 ◽

Vol 17 (3) ◽

pp. 335-349 ◽

Cited By ~ 36

Author(s):

Philippe Gallusci ◽

Annie Dedieu ◽

Etienne P. Journet ◽

Thierry Huguet ◽

David G. Barker

Keyword(s):

Root Nodule ◽

Nodule Development ◽

Nitrogen Fixing

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The Symbiosis Regulator CbrA Modulates a Complex Regulatory Network Affecting the Flagellar Apparatus and Cell Envelope Proteins

Journal of Bacteriology ◽

10.1128/jb.01834-06 ◽

2007 ◽

Vol 189 (9) ◽

pp. 3591-3602 ◽

Cited By ~ 38

Author(s):

Katherine E. Gibson ◽

Melanie J. Barnett ◽

Carol J. Toman ◽

Sharon R. Long ◽

Graham C. Walker

Keyword(s):

Sinorhizobium Meliloti ◽

Cell Envelope ◽

Global Analysis ◽

Flagellar Apparatus ◽

Nodule Development ◽

Bacterial Invasion ◽

Nitrogen Fixing ◽

Plant Host ◽

Legume Plant ◽

A Cell

ABSTRACT Sinorhizobium meliloti participates in a nitrogen-fixing symbiosis with legume plant host species of the genera Medicago, Melilotus, and Trigonella. We recently identified an S. meliloti two-component sensory histidine kinase, CbrA, which is absolutely required to establish a successful symbiosis with Medicago sativa (K. E. Gibson, G. R. Campbell, J. Lloret, and G. C. Walker, J. Bacteriol. 188:4508-4521, 2006). In addition to having a symbiotic defect, the cbrA::Tn5 mutant also has free-living phenotypes that suggest a cell envelope perturbation. Because the bases for these phenotypes are not well understood, we undertook an identification of CbrA-regulated genes. We performed a microarray analysis and compared the transcriptome of the cbrA::Tn5 mutant to that of the wild type. Our global analysis of gene expression identified 162 genes that are differentially expressed in the cbrA::Tn5 mutant, including those encoding proteins involved in motility and chemotaxis, metabolism, and cell envelope function. With regard to those genes with a known role in symbiosis, we observed increased expression of nine genes with overlapping functions in bacterial invasion of its host, which suggests that the mutant could be competent for invasion. Since these CbrA-repressed genes are vital to the invasion process, it appears that down-regulation of CbrA activity is important at this stage of nodule development. In contrast, our previous work showed that CbrA is required for bacteria to establish themselves within the host as nitrogen-fixing symbionts. Therefore, we propose a model in which CbrA functions as a developmental switch during symbiosis.

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Sinorhizobium melilotiGlutathione Reductase Is Required for both Redox Homeostasis and Symbiosis

Applied and Environmental Microbiology ◽

10.1128/aem.01937-17 ◽

2017 ◽

Vol 84 (3) ◽

Cited By ~ 3

Author(s):

Guirong Tang ◽

Ningning Li ◽

Yumin Liu ◽

Liangliang Yu ◽

Junhui Yan ◽

...

Keyword(s):

Nitrogen Fixation ◽

Sinorhizobium Meliloti ◽

De Novo ◽

Redox Homeostasis ◽

Nodule Development ◽

Glutathione Synthetase ◽

Insertion Mutation ◽

Nitrogen Fixing ◽

Content Type ◽

In Cells

ABSTRACTGlutathione (l-γ-glutamyl-l-cysteinylglycine) (GSH), one of the key antioxidants inSinorhizobium meliloti, is required for the development of alfalfa (Medicago sativa) nitrogen-fixing nodules. Glutathione exists as either reduced glutathione (GSH) or oxidized glutathione (GSSG), and its content is regulated by two pathways inS. meliloti. The first pathway is thede novosynthesis of glutathione from its constituent amino acids, namely, Glu, Cys, and Gly, catalyzed by γ-glutamylcysteine synthetase (GshA) and glutathione synthetase (GshB). The second pathway is the recycling of GSSG via glutathione reductase (GR). However, whether theS. melilotiGR functions similarly to GshA and GshB1 during symbiotic interactions with alfalfa remains unknown. In this study, a plasmid insertion mutation of theS. melilotigorgene, which encodes GR, was constructed, and the mutant exhibited delayed alfalfa nodulation, with 75% reduction in nitrogen-fixing capacity. Thegormutant demonstrated increased accumulation of GSSG and a decreased GSH/GSSG ratio in cells. The mutant also showed defective growth in rich broth and minimal broth and was more sensitive to the oxidants H2O2and sodium nitroprusside. Interestingly, the expression ofgshA,gshB1,katA, andkatBwas induced in the mutant. These findings reveal that the recycling of glutathione is important forS. melilotito maintain redox homeostasis and to interact symbiotically with alfalfa.IMPORTANCEThe antioxidant glutathione is regulated by its synthetase and reductase in cells. In the symbiotic bacteriumS. meliloti, thede novosynthesis of glutathione is essential for alfalfa nodulation and nitrogen fixation. In this study, we observed that the recycling of glutathione from GSSG not only was required for redox homeostasis and oxidative stress protection inS. meliloticells but also contributed to alfalfa nodule development and competition capacity. Our findings demonstrate that the recycling of glutathione plays a key role in nitrogen fixation symbiosis.

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Nodule-Specific Cysteine-Rich Peptides Negatively Regulate Nitrogen-Fixing Symbiosis in a Strain-Specific Manner in Medicago truncatula

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-08-17-0207-r ◽

2018 ◽

Vol 31 (2) ◽

pp. 240-248 ◽

Cited By ~ 17

Author(s):

Qi Wang ◽

Jinge Liu ◽

Hua Li ◽

Shengming Yang ◽

Peter Körmöczi ◽

...

Keyword(s):

Medicago Truncatula ◽

Sinorhizobium Meliloti ◽

Molecular Mechanisms ◽

Nodule Development ◽

Nitrogen Fixing ◽

Plant Genotype ◽

Specific Manner ◽

Meliloti Strain ◽

High Level ◽

Symbiotic Partner

Medicago truncatula shows a high level of specificity when interacting with its symbiotic partner Sinorhizobium meliloti. This specificity is mainly manifested at the nitrogen-fixing stage of nodule development, such that a particular bacterial strain forms nitrogen-fixing nodules (Nod+/Fix+) on one plant genotype but ineffective nodules (Nod+/Fix−) on another. Recent studies have just begun to reveal the underlying molecular mechanisms that control this specificity. The S. meliloti strain A145 induces the formation of Fix+ nodules on the accession DZA315.16 but Fix− nodules on Jemalong A17. A previous study reported that the formation of Fix− nodules on Jemalong A17 by S. meliloti A145 was conditioned by a single recessive allele named Mtsym6. Here we demonstrate that the specificity associated with S. meliloti A145 is controlled by multiple genes in M. truncatula, including NFS1 and NFS2 that encode nodule-specific cysteine-rich (NCR) peptides. The two NCR peptides acted dominantly to block rather than promote nitrogen fixation by S. meliloti A145. These two NCR peptides are the same ones that negatively regulate nitrogen-fixing symbiosis associated with S. meliloti Rm41.

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