Project description:During the legume-rhizobium symbiosis, free-living soil bacteria known as rhizobia trigger the formation of root nodules. The rhizobia infect these organs and adopt an intracellular lifestyle within the symbiotic nodule cells where they become nitrogen-fixing bacteroids. Several legume lineages enforce their symbionts into an extreme cellular differentiation, comprising cell enlargement and genome endoreduplication. The antimicrobial peptide transporter BclA is a major determinant of this differentiation process in Bradyrhizobium sp. ORS285, a symbiont of Aeschynomene spp.. In the absence of BclA, Bradyrhizobium sp. ORS285 proceeds until the intracellular infection of nodule cells but the bacteria cannot differentiate into enlarged polyploid bacteroids and fix nitrogen. The nodule bacteria of the bclA mutant constitute thus an intermediate stage between the free-living soil bacteria and the intracellular nitrogen-fixing bacteroids. Metabolomics on whole nodules of Aeschynomene afraspera and Aeschynomene indica infected with the ORS285 wild type or the bclA mutant revealed 47 metabolites that differentially accumulated concomitantly with bacteroid differentiation. Bacterial transcriptome analysis of these nodules discriminated nodule-induced genes that are specific to differentiated and nitrogen-fixing bacteroids and others that are activated in the host microenvironment irrespective of bacterial differentiation and nitrogen fixation. These analyses demonstrated that the intracellular settling of the rhizobia in the symbiotic nodule cells is accompanied with a first transcriptome switch involving several hundreds of upregulated and downregulated genes and a second switch accompanying the bacteroid differentiation, involving less genes but that are expressed to extremely elevated levels. The transcriptomes further highlighted the dynamics of oxygen and redox regulation of gene expression during nodule formation and we discovered that bclA represses the expression of non-ribosomal peptide synthetase gene clusters suggesting a non-symbiotic function of BclA. Together, our data uncover the metabolic and gene expression changes that accompany the transition from intracellular bacteria into differentiated nitrogen-fixing bacteroids.
Project description:To circumvent the paucity of nitrogen sources in the soil Legume plants evolved a symbiotic interaction with nitrogen-fixing soil bacteria called rhizobia. During symbiosis, legumes form root organs called nodules, where bacteria are housed intracellularly and become active nitrogen fixers known as bacteroids. Depending on their host plant, bacteroids can adopt different morphotypes, being either unmodified (U), elongated (E) or spherical (S). E- and S-typr bacteroids undergo a terminal differentiation leading to irreversible morphological changes and DNA endoreduplication. Previous studies suggest that differentiated bacteroids display an increased symbiotic efficiency (E>U & S>U). In this study, we used a combination of Aeschynomene species inducing E- and S-type bacteroids in symbiosis with Bradyrhizobium sp. ORS285 to show that S- performed better than E-type bacteroids. Thus, we performed a transcriptomic analysis on E- and S-type bacteroids to identify the bacterial functions involved in each bacteroid type.
Project description:Metabolomics and transcriptomics of Bradyrhizobium diazoefficiens-induced root nodules Bradyrhizobium diazoefficiens is a nitrogen-fixing endosymbiont, which can grow inside root-nodule cells of the agriculturally important soybean and other host plants. Our previous studies described B. diazoefficiens host-specific global expression changes occurring during legume infection at the transcript and protein level. In order to further characterize nodule metabolism, we here determine by flow injection -time of flight mass spectrometry analysis the metabolome of i) nodules and roots from four different B. diazoefficiens host plants, ii) soybean nodules harvested at different time points during nodule development, and iii) soybean nodules infected by two strains mutated in key genes for nitrogen fixation, respectively. Ribose (soybean), tartaric acid (mungbean), hydroxybutanoyloxybutanoate (siratro) and catechol (cowpea) were among the metabolites found to be specifically elevated in one of the respective host plants. While the level of C4-dicarboxylic acids decreased during soybean nodule development, we observed an accumulation of trehalose-phosphate at 21 days post infection (dpi). Moreover, nodules from non-nitrogen-fixing bacteroids (nifA and nifH mutants) showed specific metabolic alterations; these were also supported by transcriptomics data that was generated for the two mutant strains and were helpful to separate for some examples the respective bacterial and plant contributions to the metabolic profile. The alterations included signs of nitrogen limitation in both mutants, and an increased level of a phytoalexin in nodules induced by the nifA mutant, suggesting that the tissue of these nodules exhibits defense and stress reactions.
Project description:To investigate the gene expression levels of Medicago truncatula roots after beneficial fungi Gongronella sp. w5 inoculated.Gongronella sp. w5 promoted M. truncatula growth and caused the accumulation of sucrose in M. truncatula root tissue at 16 day-post-inoculation (dpi) without invading into the root cells. The transport of photosynthetic product sucrose to the rhizosphere by M. truncatula root cells was accelerated by upregulating the SWEET gene.
Project description:part of GSE8478: Genome-wide transcript analysis of Bradyrhizobium japonicum bacteroids in soybean root nodules This SuperSeries is composed of the SubSeries listed below.
Project description:Global bottom-up proteomics analysis of proteins purified from soybean root nodules infected with either WT or nifH- mutant Bradyrhizobium japonicum. Nine glycoproteins containing Lewis-a N-glycans, with 3 distinct Lewis-a epitopes (Hex:5 HexNAc:4 dHex:3 Pent:1, Hex:4 HexNAc:4 dHex:2 Pent:1, and Hex:4 HexNAc:3 dHex:2 Pent:1) were observed. Proteins purified from WT and nifH- infected soybean root nodules (five biological replicates each) were reduced using dithiothreitol, alkylated with iodoacetamide and trypsin digested followed by C18 SPE clean-up and LC-MS/MS analysis. Raw data files were processed using FragPipe v17.1, then output files 'combined_modified_peptide.tsv' and 'combined_protein.tsv' were used to identify glycopeptides and for global protein quantitation. These files, along with Excel files containing global quantitation analysis files for soybean nodule (Glycine max) and bacterial (Bradyrhizobium), are available in directory 'Quantification/MSFragger_results'. Glycopeptide data were also processed with PMI Byonic, and Excel file results are available in directory 'Quantification/PMI_Byonic_results'.
Project description: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 (dai), time points that coincided 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 qRT-PCR 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. Keywords = symbiosis Keywords = nodulation Keywords = rhizobium Keywords = defense Keywords = ANOVA Keywords = plant loop design, 7 samples, 7 comparison, 2 technical repeats including dye swaps, 4 biological repeats
Project description:Legume plants can form root organs called nodules where they house intracellular symbiotic rhizobium bacteria. Within nodule cells, rhizobia differentiate into bacteroids, which fix nitrogen for the benefit of the plant. Depending on the combination of host plants and rhizobial strains, the output of rhizobium-legume interactions is varying from non-fixing associations to symbioses that are highly beneficial for the plant. Bradyrhizobium diazoefficiens USDA110 was isolated as a soybean symbiont but it can also establish a functional symbiotic interaction with Aeschynomene afraspera. In contrast to soybean, A. afraspera triggers terminal bacteroid differentiation, a process involving bacterial cell elongation, polyploidy and membrane permeability leading to loss of bacterial viability while plants increase their symbiotic benefit. A combination of plant metabolomics, bacterial proteomics and transcriptomics along with cytological analyses was used to study the physiology of USDA110 bacteroids in these two host plants. We show that USDA110 establish a poorly efficient symbiosis with A. afraspera, despite the full activation of the bacterial symbiotic program. We found molecular signatures of high level of stress in A. afraspera bacteroids whereas those of terminal bacteroid differentiation were only partially activated. Finally, we show that in A. afraspera, USDA110 bacteroids undergo an atypical terminal differentiation hallmarked by the disconnection of the canonical features of this process. This study pinpoints how a rhizobium strain can adapt its physiology to a new host and cope with terminal differentiation when it did not co-evolve with such a host.