Genome-Wide Transcript Analysis of Bradyrhizobium japonicum Bacteroids in Soybean Root Nodules

The transcriptome of endosymbiotic Bradyrhizobium japonicum bacteroids was assessed, using RNA extracted from determinate soybean root nodules. Results were compared with the transcript profiles of B. japonicum cells grown in either aerobic or microaerobic culture. Microoxia is a known trigger for the induction of symbiotically relevant genes. In fact, one third of the genes induced in bacteroids at day 21 after inoculation are congruent with those up-regulated in culture by a decreased oxygen concentration. The other induced genes, however, may be regulated by cues other than oxygen limitation. Both groups of genes provide a rich source for the possible discovery of novel functions related to symbiosis. Samples taken at different timepoints in nodule development have led to the distinction of genes expressed early and late in bacteroids. The experimental approach applied here is also useful for B. japonicum mutant analyses. As an example, we compared the transcriptome of wild-type bacteroids with that of bacteroids formed by a mutant defective in the RNA polymerase transcription factor σ54. This led to a collection of hitherto unrecognized B. japonicum genes potentially transcribed in planta in a σ54-dependent manner.

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Strain distribution and in planta production of an extracellular polysaccharide depolymerase from Bradyrhizobium japonicum

Canadian Journal of Microbiology ◽

10.1139/m92-139 ◽

1992 ◽

Vol 38 (8) ◽

pp. 857-861 ◽

Cited By ~ 3

Author(s):

Michael F. Dunn ◽

Arthur L. Karr

Keyword(s):

Bradyrhizobium Japonicum ◽

Root Nodules ◽

Extracellular Polysaccharide ◽

Extracellular Polysaccharides ◽

Neutral Sugar ◽

In Planta ◽

In Vitro Production ◽

Polysaccharide Composition ◽

Vitro Production

Thirty-four strains of Bradyrhizobium japonicum were screened for the in vitro production of an extracellular polysaccharide depolymerase active against the B. japonicum acidic extracellular polysaccharide that contains mannose, glucose, galactose, and 4-O-methylgalactose as neutral sugar components. Over 90% of tested strains producing this type of extracellular polysaccharide also produced the extracellular polysaccharide depolymerase, whereas strains producing a compositionally different extracellular polysaccharide did not. In addition, representatives of species related to B. japonicum by extracellular polysaccharide composition or host range were also phenotypically depolymerase negative. Depolymerase was also present in soybean root nodules formed by B. japonicum strain 2143. In contrast to the cell-associated depolymerase activity found in free-living cells of this strain, most of the depolymerase activity present in nodules is free of the bacteroids. The widespread occurrence of the depolymerase among B. japonicum strains and the spatiotemporal distribution of its activity in planta are consistent with the enzyme playing a role in the removal of surface extracellular polysaccharide from the microorganism during the infection of nodulation process. Key words: Bradyrhizobium japonicum, soybean, extracellular polysaccharides, extracellular polysaccharide depolymerase, bacteroids.

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Starch content and activities of starch-metabolizing enzymes in effective and ineffective root nodules of soybean

Canadian Journal of Botany ◽

10.1139/b91-094 ◽

1991 ◽

Vol 69 (4) ◽

pp. 697-701 ◽

Cited By ~ 15

Author(s):

Sharon I. Forrest ◽

Desh Pal S. Verma ◽

Rajinder S. Dhindsa

Keyword(s):

Nitrogen Fixation ◽

Starch Content ◽

Root Nodules ◽

Starch Synthase ◽

Nodule Development ◽

Wild Type ◽

Starch Phosphorylase ◽

High Concentration ◽

Key Words Nitrogen Fixation ◽

Glycine Max L

Starch content and activities of some enzymes of starch metabolism were determined in wild-type, N2-fixing (fix+) nodules and in two non-N2-fixing (fix−) nodules induced by Bradyrhizobium japonicum mutant strains, T5-95 and T8-1, on soybean (Glycine max L.) roots. The T5-95 nodules are similar to wild type in ultrastructure, but the T8-1 nodules are different in that the bacteroids are not released from the infection thread. After initial accumulation to relatively high concentration, starch was depleted during nitrogen fixation in fix+ nodules. However, in fix− nodules, the accumulated starch was not metabolized. The activity of starch-bound starch synthase (EC 2.4.1.21) declined in fix+ nodules but remained high in fix− nodules. The activity of α-amylase (EC 3.2.1.1) was only slightly higher than wild type in T5-95 but was four times higher than wild type in T8-1 nodules. The activity of starch phosphorylase (EC 2.4.1.1) increased in all nodule types from 14 to 21 days postinfection. A positive correlation was observed between the capacity of nodules to fix N2 and their capacity to degrade starch. Collectively, these results support the concept that starch accumulated during early stages of nodule development is metabolized to supply energy for nitrogen fixation and to meet the metabolic demands of bacteroids. Key words: nitrogen fixation, starch content, effective and ineffective nodules, starch synthase, starch phosphorylase, α-amylase.

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Morphogenesis of root nodules in white clover. II. The effect of mutation in genes nod IJ of the microsymbiont upon the nodule structure

Acta Societatis Botanicorum Poloniae ◽

10.5586/asbp.1998.001 ◽

2014 ◽

Vol 67 (1) ◽

pp. 5-22

Author(s):

Barbara Łotocka ◽

Joanna Kopcińska ◽

Władysław Golinowski

Keyword(s):

White Clover ◽

Rhizobium Leguminosarum ◽

Root Nodules ◽

Single Cells ◽

Wild Strain ◽

Root Hairs ◽

Nodule Development ◽

Wild Type ◽

Delayed Reaction ◽

Infection Threads

Morphogenesis of ineffective root nodules initiated on the roots of white clover 'Astra' by the <em>Rhizobium leguminosarum</em> biovar. <em>trifolii</em> strains ANU261 (Tn5 insertion in nod 1 gene) and ANU262 (Tn5 insertion in nod J gene) was investigated. Following changes were observed, as compared to the wild-type nodulation: the exaggerated, not delayed reaction of root hairs; the delay in nodulation with the number of nodules the same as in plants inoculated with a wild strain; the formation and organization of the nodule primordium not changed in comparison with the wild-type nodules; infection threads abnormally branched and diffusing with bacteria deprived of light zone and enriched with storage material; infected cells of bacteroidal tissue abnormally strongly osmiophilic and only slightly vacuolated; symbiosomes with very narrowed peribacteroidal space, subject to premature degradation; abnormal accumulation of starch in the nodule tissues; nodule development blocked at the stage of laterally situated meristem and single nodule bundle; inhibition of divisions in the meristem and vacuolation of its cells; the appearance of single cells with colonies of saprophytic rhizobia embedded in the fibrillar matrix in the old, degraded regions of the bacteroidal tissue.

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Particle density and protein composition of the peribacteroid membrane from soybean root nodules is affected by mutation in the microsymbiont Bradyrhizobium japonicum

Planta ◽

10.1007/bf00394780 ◽

1988 ◽

Vol 174 (2) ◽

pp. 263-270 ◽

Cited By ~ 18

Author(s):

Dietrich Werner ◽

Erhard M�rschel ◽

Christine Garbers ◽

Stefan Bassarab ◽

Robert B. Mellor

Keyword(s):

Bradyrhizobium Japonicum ◽

Particle Density ◽

Root Nodules ◽

Protein Composition ◽

Peribacteroid Membrane ◽

Soybean Root

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An Iron Uptake Operon Required for Proper Nodule Development in the Bradyrhizobium japonicum-Soybean Symbiosis

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-18-0950 ◽

2005 ◽

Vol 18 (9) ◽

pp. 950-959 ◽

Cited By ~ 23

Author(s):

Heather P. Benson ◽

Eric Boncompagni ◽

Mary Lou Guerinot

Keyword(s):

Bradyrhizobium Japonicum ◽

Iron Acquisition ◽

Outer Membrane Proteins ◽

Protein Characterization ◽

Nodule Development ◽

In Planta ◽

Siderophore Receptors ◽

Infected Cells ◽

Inner Membrane Protein ◽

Mutant Forms

Rhizobia live in the soil or enter into a nitrogen-fixing symbiosis with a suitable host plant. Each environment presents different challenges with respect to iron acquisition. The soybean symbiont Bradyrhizobium japonicum 61A152 can utilize a variety of siderophores (Fe[III]-specific ligands). Purification of iron-regulated outer membrane proteins had previously allowed the cloning of a gene, fegA, from B. ja-ponicum 61A152, whose predicted protein shares significant amino acid similarity with known TonB-dependent siderophore receptors. Here, we show that fegA is in an operon with a gene, fegB, that is predicted to encode an inner membrane protein. Characterization of fegAB and fegB mutants shows that both fegA and fegB are required for utilization of the siderophore ferrichrome. Whereas the fegB mutant forms a normal symbiosis, the fegAB mutant has a dramatic phenotype in planta. Six weeks after inoculation with a fegAB strain, soybean nodules do not contain leghemoglobin and do not fix nitrogen. Infected cells contain few symbiosomes and are filled with vesicles. As ferrichrome is a fungal siderophore not likely to be available in nodules, the symbiotic defect suggests that the fegAB operon is serving a different function in planta, possibly one involved in signaling between the two partners.

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Increase in root nodulation and crop yield of soybean by native Bradyrhizobium japonicum strains

Botanica Orientalis Journal of Plant Science ◽

10.3126/botor.v6i0.2902 ◽

2010 ◽

Vol 6 ◽

pp. 1-3

Author(s):

Namraj Dhami ◽

Braj Nandan Prasad

Keyword(s):

Glycine Max ◽

Crop Yield ◽

Bradyrhizobium Japonicum ◽

Root Nodules ◽

Bromothymol Blue ◽

Strong Positive Correlation ◽

N Content ◽

Root Nodulation ◽

Soybean Root ◽

Gram Staining

Native strains of Bradyrhizobium japonicum were tested for their effectiveness on nodulation, crop yield and nitrogen fixation in soybean (Glycine max). B. japonicum strains were isolated from soybean root nodules collected from different agro-climatic regions of Far Western Nepal, viz. Dipayal (607 m asl), Dadeldhura (1097 m asl), Silgadhi (1209 m asl) and Bajura (1524 m asl). The strains were characterized by studying colony characteristics, growth response with Congo red and Bromothymol blue, and Gram staining. The native bradyrhizobial strains were authenticated by performing infection test on soybean seedlings. All the four strains were found compatible and effective on root nodulation, crop yield and soil nitrogen (N) content. Inoculation of these strains increased soybean root nodulation by 247-343% and crop yield by 45-204%. There was strong positive correlation (r = 0.982) between number of root nodules and crop yield, which suggest that optimization of root nodulation by inoculating compatible and effective B. japonicum strains significantly increase the soybean crop yield. Soil N content of inoculated experimental pots was increased by 13-33%. However, variability among different strains was observed in their effect on root nodulation and yield performance. B. japonicum strain collected from Silgadhi was found to be the most effective in increasing nodule number and crop yield by 343% and 204% respectively. Key-words: Glycine max; gram staining; inoculation; nitrogen fixationDOI: 10.3126/botor.v6i0.2902 Botanica Orientalis - Journal of Plant Science (2009) 6: 1-3

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A Link between Arabinose Utilization and Oxalotrophy in Bradyrhizobium japonicum

Applied and Environmental Microbiology ◽

10.1128/aem.03314-13 ◽

2014 ◽

Vol 80 (7) ◽

pp. 2094-2101 ◽

Cited By ~ 9

Author(s):

Marion Koch ◽

Nathanaël Delmotte ◽

Christian H. Ahrens ◽

Ulrich Omasits ◽

Kathrin Schneider ◽

...

Keyword(s):

Bradyrhizobium Japonicum ◽

Host Plants ◽

Root Nodule ◽

Coenzyme A ◽

Root Nodules ◽

Proteome Analysis ◽

Wild Type ◽

Content Type ◽

Mutant Cells ◽

Respective Host

ABSTRACTRhizobia have a versatile catabolism that allows them to compete successfully with other microorganisms for nutrients in the soil and in the rhizosphere of their respective host plants. In this study,Bradyrhizobium japonicumUSDA 110 was found to be able to utilize oxalate as the sole carbon source. A proteome analysis of cells grown in minimal medium containing arabinose suggested that oxalate oxidation extends the arabinose degradation branch via glycolaldehyde. A mutant of the key pathway genesoxc(for oxalyl-coenzyme A decarboxylase) andfrc(for formyl-coenzyme A transferase) was constructed and shown to be (i) impaired in growth on arabinose and (ii) unable to grow on oxalate. Oxalate was detected in roots and, at elevated levels, in root nodules of four differentB. japonicumhost plants. Mixed-inoculation experiments with wild-type andoxc-frcmutant cells revealed that oxalotrophy might be a beneficial trait ofB. japonicumat some stage during legume root nodule colonization.

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Iron-Dependent Cytochrome c1 Expression Is Mediated by the Status of Heme in Bradyrhizobium japonicum

Journal of Bacteriology ◽

10.1128/jb.187.15.5084-5089.2005 ◽

2005 ◽

Vol 187 (15) ◽

pp. 5084-5089 ◽

Cited By ~ 7

Author(s):

Tao Gao ◽

Mark R. O'Brian

Keyword(s):

Bradyrhizobium Japonicum ◽

Aminolevulinic Acid ◽

Iron Status ◽

Covalent Binding ◽

Dependent Manner ◽

Wild Type ◽

Prosthetic Group ◽

Protein Levels ◽

Genes Encoding ◽

In The Wild

ABSTRACT The heme prosthetic group of heme proteins contains iron, which can be a limiting nutrient. Here, we show that cytochrome c 1 protein from Bradyrhizobium japonicum was strongly affected by the iron status, with low expression in cells grown under iron limitation. This control was not affected in mutants encoding the iron regulator Irr or Fur. Furthermore, cytochrome c 1 mRNA was not influenced by the iron status, suggesting control at a posttranscriptional step. Cytochrome c 1 protein levels were very low in mutants defective in the genes encoding δ-aminolevulinic acid (ALA) synthase and ferrochelatase, enzymes that catalyze the first and final steps of the heme biosynthetic pathway, respectively. Iron-dependent cytochrome c 1 expression was restored in the ALA synthase mutant by supplementation of the medium with the heme precursor ALA. Supplementation with heme resulted in high levels of cytochrome c 1 protein in the wild type and in both mutants, but expression was no longer iron dependent. Cytochrome c 1 is synthesized as a protein precursor fused with cytochrome b. A plasmid-borne construct encoding only cytochrome c 1 was expressed in an iron- and heme-dependent manner similar to that of the wild-type gene, indicating that control by those effectors is not linked to posttranslational processing of the fusion protein. Mutation of the cytochrome c 1 cysteines involved in covalent binding to heme nearly abolished immunodetectable protein. Thus, defects in heme synthesis or heme binding abrogate cytochrome c 1 accumulation, apparently due to protein degradation. We suggest that iron-dependent cytochrome c 1 expression is mediated by heme availability for heme protein formation

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Transcription Profiling of Soybean Nodulation by Bradyrhizobium japonicum

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-21-5-0631 ◽

2008 ◽

Vol 21 (5) ◽

pp. 631-645 ◽

Cited By ~ 64

Author(s):

Laurent Brechenmacher ◽

Moon-Young Kim ◽

Marisol Benitez ◽

Min Li ◽

Trupti Joshi ◽

...

Keyword(s):

Nitrogen Fixation ◽

Bradyrhizobium Japonicum ◽

Root Nodules ◽

Expression Patterns ◽

Root Hairs ◽

Defense Responses ◽

Nodule Development ◽

Plant Defense Responses ◽

Polymerase Chain ◽

High Level

Legumes interact with nodulating bacteria that convert atmospheric nitrogen into ammonia for plant use. This nitrogen fixation takes place within root nodules that form after infection of root hairs by compatible rhizobia. Using cDNA microarrays, we monitored gene expression in soybean (Glycine max) inoculated with the nodulating bacterium Bradyrhizobium japonicum 4, 8, and 16 days after inoculation, timepoints that coincide with nodule development and the onset of nitrogen fixation. This experiment identified several thousand genes that were differentially expressed in response to B. japonicum inoculation. Expression of 27 genes was analyzed by quantitative reverse transcriptase-polymerase chain reaction, and their expression patterns mimicked the microarray results, confirming integrity of analyses. The microarray results suggest that B. japonicum reduces plant defense responses during nodule development. In addition, the data revealed a high level of regulatory complexity (transcriptional, post-transcriptional, translational, post-translational) that is likely essential for development of the symbiosis and adjustment to an altered nutritional status.

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