Project description:Certain alpha- and beta-proteobacteria, the rhizobia, are able to infect legume roots, elicit root nodules, and live therein as endosymbiotic, nitrogen-fixing bacteroids. Host recognition and specificity are the results of consecutive programming events in bacteria and host plants in which important signaling molecules, e.g. plant flavonoids and rhizobial lipooligosaccharides, play key roles. Here, we introduce a new aspect of this symbiosis: the adaptive response to hosts. In contrast to host specificity, which determines early steps in bacteria-plant interaction, the adaptation to hosts refers to late events in mature bacteroids where specific genes are transcribed and translated that help the endosymbionts to meet the disparate environmental requirements imposed by the hosts in which they live. This concept was elaborated with Bradyrhizobium japonicum and three different legumes (soybean, cowpea, siratro). We systematically analyzed and compared the transcriptomes as well as the proteomes in bacteroids from root nodules of the three hosts. Transcripts and proteins were thus identified which are induced in only one of the three hosts. We then focused on those determinants that were congruent in the two data sets of host-specific transcripts and proteins, and arrived at 20 for soybean, 7 for siratro, and 4 for cowpea. One conspicuous gene cluster for a predicted ABC-type transporter, differentially expressed in siratro, was deleted. The corresponding mutant had a symbiotic defect on siratro rather than on soybean or cowpea. This result demonstrates the value of the applied approach and corroborates the host-specific adaptation concept. B. japonicum transcriptome was determined for the three different host plants
Project description:Certain alpha- and beta-proteobacteria, the rhizobia, are able to infect legume roots, elicit root nodules, and live therein as endosymbiotic, nitrogen-fixing bacteroids. Host recognition and specificity are the results of consecutive programming events in bacteria and host plants in which important signaling molecules, e.g. plant flavonoids and rhizobial lipooligosaccharides, play key roles. Here, we introduce a new aspect of this symbiosis: the adaptive response to hosts. In contrast to host specificity, which determines early steps in bacteria-plant interaction, the adaptation to hosts refers to late events in mature bacteroids where specific genes are transcribed and translated that help the endosymbionts to meet the disparate environmental requirements imposed by the hosts in which they live. This concept was elaborated with Bradyrhizobium japonicum and three different legumes (soybean, cowpea, siratro). We systematically analyzed and compared the transcriptomes as well as the proteomes in bacteroids from root nodules of the three hosts. Transcripts and proteins were thus identified which are induced in only one of the three hosts. We then focused on those determinants that were congruent in the two data sets of host-specific transcripts and proteins, and arrived at 20 for soybean, 7 for siratro, and 4 for cowpea. One conspicuous gene cluster for a predicted ABC-type transporter, differentially expressed in siratro, was deleted. The corresponding mutant had a symbiotic defect on siratro rather than on soybean or cowpea. This result demonstrates the value of the applied approach and corroborates the host-specific adaptation concept.
Project description:Brassinosteroid (BR) is an essential hormone in plant growth and development. BR signaling pathway has been extensively studied, in which Brassinazole resistant 1 (BZR1) functions as a key regulator. Here, we carried out a functional study of the homolog of BZR1 in Medicago truncatula, whose expression was induced in nodules upon rhizobial inoculation. We identified a loss-of-function mutant mtbzr1-1 and generated 35S:MtBZR1 transgenic lines for further analysis at the genetic level. Both the mutant and the overexpressor lines of MtBZR1 showed no obvious phenotypic changes under normal growth condition. After rhizobial inoculation, however, the shoot and root dry mass was reduced in mtbzr1-1 compared with the wild-type, caused by partially impaired nodule development. The transcriptomic analysis identified 1,319 differentially expressed genes in mtbzr1-1 compared with wild-type, many of which are involved in nodule development and secondary metabolite biosynthesis. Our results demonstrate an essential role of MtBZR1 in nodule development in M. truncatula, shedding light on the potential role of BR in legume-rhizobium symbiosis.
Project description:A transcriptome of Cluster II Frankia in nitrogen-fixing root-nodule symbiosis with the host plant, Datisca glomerata, was obtained by Illumina sequencing and mapping to the corresponding published genome (NCBI Bioproject PRJNA46257). Major metabolic pathways detected in Cluster II Frankia in symbiosis with Datisca glomerata were comparable to those described as up-regulated in the Frankia alni-Alnus glutinosa symbiosis (N Alloisio et al, MPMI 23(5):593-607, 2010): nitrogenase biosynthesis, tricarboxylic acid cycle, respiratory-chain related functions, oxidation protection, and terpenoid biosynthesis. These functions are consistent with the primary activities of Frankia in root nodules, e.g. to carry out the energetically-demanding fixation of atmospheric dinitrogen to ammonium, and to maintain internal reducing conditions. Expression of genes coding for amino-acid biosynthetic pathways, including arginine as reported previously (AM Berry et al. Funct Plant Biol 38, 645–652, 2011) was detected. A striking difference from other Frankia strains, revealed in the transcriptome of the Cluster II Frankia in symbiosis, was the expression of homologs of rhizobial nodulation genes, nodA, nodB and nodC.
Project description:Legumes perform symbiotic nitrogen fixation through rhizobial bacteroids housed in specialised root nodules. The biochemical process is energy‐intensive and consumes a huge carbon source to generate sufficient reducing power. To maintain the symbiosis, malate is supplied by legume nodules to bacteroids as their major carbon and energy source in return for ammonium ions and nitrogenous compounds. To sustain the carbon supply to bacteroids, nodule cells undergo drastic reorganisation of carbon metabolism. Here, a comprehensive quantitative comparison of the mitochondrial proteomes between root nodules and uninoculated roots was performed using data‐independent acquisition proteomics, revealing the modulations in nodule mitochondrial proteins and pathways in response to carbon reallocation. Corroborated our findings with that from the literature, we believe nodules preferably allocate cytosolic phosphoenolpyruvates towards malate synthesis in lieu of pyruvate synthesis, and nodule mitochondria prefer malate over pyruvate as the primary source of NADH for ATP production. Moreover, the differential regulation of respiratory chain‐associated proteins suggests that nodule mitochondria could enhance the efficiencies of complexes I and IV for ATP synthesis. This study highlighted a quantitative proteomic view of the mitochondrial adaptation in soybean nodules.
Project description:Nitrogen assimilation in plants is a tightly regulated process that integrates developmental and environmental signals. The legume-rhizobial symbiosis results in the formation of a specialized organ called root nodule, where the rhizobia fixes atmospheric nitrogen into ammonia. Ammonia is assimilated by the plant enzyme glutamine synthetase, which is specifically inhibited by PPT. The expression of key genes related to the regulation of root nodule metabolism will likely be affected by glutamine synthetase inhibition. We used microarrays to detail the global programme of gene expression in response to Glutamine synthetase inhibition in root nodules and identified genes differentially expressed over a time course.
Project description:We have undertaken a detailed study to identify mechanisms regulating expression of NCRs. We used a custom Affymetrix oligonucleotide microarray to examine the expression changes of 566 NCRs in different stages of nodule development. Additionally, rhizobial mutants were used to understand the importance of the rhizobial components in induction of NCRs. Early NCRs were detected during the initial infection of rhizobia in nodules and continue to be expressed into the late stages of nodule development. Late NCRs were induced concomittant with bacteroid development in the nodules. The induction of these groups of genes was correlated with the number and morphology of rhizobia in the nodule.
Project description:We have undertaken a detailed study to identify mechanisms regulating expression of NCRs. We used a custom Affymetrix oligonucleotide microarray to examine the expression changes of 566 NCRs in different stages of nodule development. Additionally, rhizobial mutants were used to understand the importance of the rhizobial components in induction of NCRs. Early NCRs were detected during the initial infection of rhizobia in nodules and continue to be expressed into the late stages of nodule development. Late NCRs were induced concomittant with bacteroid development in the nodules. The induction of these groups of genes was correlated with the number and morphology of rhizobia in the nodule. We used a custom Affymetrix chip containing 684 probe sequences of Medicago DEFLs to explore the expression patterns of NCRs in nodules inoculated with Sinorhizobium meliloti 1021(Sm1021) at marked developmental stages and nodules inoculated with various mutants derived from Sm1021 totalling 14 different treatments. Each treatment was supported by three biological replicates giving a grand total of 42 samples.
Project description:Paraburkholderia phymatum belongs to the β-subclass of proteobacteria. It has recently been shown to be able to nodulate and fix nitrogen in symbiosis with several mimosoid and papillionoid legumes. In contrast to symbiosis of legumes with α-proteobacteria, very little is known about the molecular determinants underlying the successful establishment of this mutualistic relationship with β-proteobacteria. In this study, we analyzed RNA-seq data of free-living P. phymatum growing under nitrogen replete and limited conditions, the latter partially mimicking the situation in nitrogen deprived soils. Among the genes up-regulated under nitrogen limitation, we found genes involved in exopolysaccharide production and motility, two traits relevant for plant root infection. Next, RNA-seq data of P. phymatum grown under free-living conditions and from symbiotic root nodules of Phaseolus vulgaris (common bean) were generated and compared. Among the genes highly up-regulated during symbiosis, we identified an operon encoding a potential cytochrome o ubiquinol oxidase (Bphy_3646-49). Bean root nodules induced by a cyoB mutant strain showed reduced nitrogenase and nitrogen fixation abilities suggesting an important role of the cytochrome for respiration inside the nodule. Analysis of mutant strains for RNA polymerase transcription factor rpoN (σ54) and its activator NifA indicated that – similar to the situation in α-rhizobia – P. phymatum RpoN and NifA are key regulators during symbiosis with P. vulgaris.