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Identification of a functional fur gene in Bradyrhizobium japonicum.

ABSTRACT: The recent identification of the iron response regulator (Irr) in Bradyrhizobium japonicum raised the question of whether the global regulator Fur is present in that organism. A fur gene homolog was isolated by the functional complementation of an Escherichia coli fur mutant. The B. japonicum Fur bound to a Fur box DNA element in vitro, and a fur mutant grown in iron-replete medium was derepressed for iron uptake activity. Thus, B. japonicum expresses at least two regulators of iron metabolism.

SUBMITTER: Hamza I

PROVIDER: S-EPMC94108 | biostudies-literature | 1999 Sep

REPOSITORIES: biostudies-literature

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Identification of a functional fur gene in Bradyrhizobium japonicum.

Hamza I I Hassett R R O'Brian M R MR

Journal of bacteriology 19990901 18

The recent identification of the iron response regulator (Irr) in Bradyrhizobium japonicum raised the question of whether the global regulator Fur is present in that organism. A fur gene homolog was isolated by the functional complementation of an Escherichia coli fur mutant. The B. japonicum Fur bound to a Fur box DNA element in vitro, and a fur mutant grown in iron-replete medium was derepressed for iron uptake activity. Thus, B. japonicum expresses at least two regulators of iron metabolism. ...[more]

PMID: 10482529

Similar Datasets

A dominant-negative fur mutation in Bradyrhizobium japonicum.

Project description:In many bacteria, the ferric uptake regulator (Fur) protein plays a central role in the regulation of iron uptake genes. Because iron figures prominently in the agriculturally important symbiosis between soybean and its nitrogen-fixing endosymbiont Bradyrhizobium japonicum, we wanted to assess the role of Fur in the interaction. We identified a fur mutant by selecting for manganese resistance. Manganese interacts with the Fur protein and represses iron uptake genes. In the presence of high levels of manganese, bacteria with a wild-type copy of the fur gene repress iron uptake systems and starve for iron, whereas fur mutants fail to repress iron uptake systems and survive. The B. japonicum fur mutant, as expected, fails to repress iron-regulated outer membrane proteins in the presence of iron. Unexpectedly, a wild-type copy of the fur gene cannot complement the fur mutant. Expression of the fur mutant allele in wild-type cells leads to a fur phenotype. Unlike a B. japonicum fur-null mutant, the strain carrying the dominant-negative fur mutation is unable to form functional, nitrogen-fixing nodules on soybean, mung bean, or cowpea, suggesting a role for a Fur-regulated protein or proteins in the symbiosis.

| S-EPMC344408 | biostudies-literature

The ftsA gene as a molecular marker for phylogenetic studies in Bradyrhizobium and identification of Bradyrhizobium japonicum.

Project description:The use of ftsA gene sequences for taxonomic studies of the genus Bradyrhizobium bacteria was assessed. The ftsA gene codes for an actin-like protein involved in prokaryotic cell division. Up to now, this gene has not been used as a phylogenetic marker for analysis of bacteria establishing root nodule symbiosis with Fabaceae plants. In this study, the ftsA gene sequences obtained for bradyrhizobia forming N2 fixing symbiosis with four Genisteae tribe plants growing in Poland and most of the type strains of the genus Bradyrhizobium species were analyzed and evaluated as molecular markers for phylogenetic studies of these bacteria for the first time. The ftsA gene sequences of all bradyrhizobial strains with completely or partially sequenced genomes, available in the GenBank database, were also included into the analysis. The phylogeny of the ftsA gene was compared to the phylogenies of other chromosomal genes commonly used in the studies of Bradyrhizobium bacteria. The results showed that the phylogenies of ftsA and the core genes recA and glnII were congruent, making the ftsA gene useful as a phylogenetic marker. Analysis of the ftsA gene sequences revealed a single-nucleotide polymorphism unique to Bradyrhizobium japonicum strains, and the potential use of this SNP for identification of this species was discussed.

| S-EPMC6373400 | biostudies-literature

Metal-specific control of gene expression mediated by Bradyrhizobium japonicum Mur and Escherichia coli Fur is determined by the cellular context.

Project description:Bradyrhizobium japonicum Mur and Escherichia coli Fur are manganese- and iron-responsive transcriptional regulators, respectively, that belong to the same protein family. Here, we show that neither Mur nor Fur discriminate between Fe(2+) and Mn(2+) in vitro nor is there a metal preference for conferral of DNA-binding activity on the purified proteins. When expressed in E. coli, B. japonicum Mur responded to iron, but not manganese, as determined by in vivo promoter occupancy and transcriptional repression activity. Moreover, E. coli Fur activity was manganese-dependent in B. japonicum. Total and chelatable iron levels were higher in E. coli than in B. japonicum under identical growth conditions, and Mur responded to iron in a B. japonicum iron export mutant that accumulated high levels of the metal. However, elevated manganese content in E. coli did not confer activity on Fur or Mur, suggesting a regulatory pool of manganese in B. japonicum that is absent in E. coli. We conclude that the metal selectivity of Mur and Fur depends on the cellular context in which they function, not on intrinsic properties of the proteins. Also, the novel iron sensing mechanism found in the rhizobia may be an evolutionary adaptation to the cellular manganese status.

| S-EPMC4925281 | biostudies-literature

Characterization of the Bradyrhizobium japonicum ftsH gene and its product.

Project description:The Bradyrhizobium japonicum ftsH gene was cloned by using a set of widely applicable degenerated oligonucleotides. Western blot experiments indicated that the FtsH protein was produced under standard growth conditions and that it was not heat inducible. Attempts to delete the ftsH gene in B. japonicum failed, suggesting a pivotal cellular function of this gene. The expression of B. japonicum ftsH in an ftsH-negative Escherichia coli strain significantly enhanced the fitness of this mutant and reduced the steady-state level of sigma(32).

| S-EPMC103706 | biostudies-literature

The Bradyrhizobium japonicum frcB gene encodes a diheme ferric reductase.

Project description:Iron utilization by bacteria in aerobic environments involves uptake as a ferric chelate from the environment, followed by reduction to the ferrous form. Ferric iron reduction is poorly understood in most bacterial species. Here, we identified Bradyrhizobium japonicum frcB (bll3557) as a gene adjacent to, and coregulated with, the pyoR gene (blr3555) encoding the outer membrane receptor for transport of a ferric pyoverdine. FrcB is a membrane-bound, diheme protein, characteristic of eukaryotic ferric reductases. Heme was essential for FrcB stability, as were conserved histidine residues in the protein that likely coordinate the heme moieties. Expression of the frcB gene in Escherichia coli conferred ferric reductase activity on those cells. Furthermore, reduced heme in purified FrcB was oxidized by ferric iron in vitro. B. japonicum cells showed inducible ferric reductase activity in iron-limited cells that was diminished in an frcB mutant. Steady-state levels of frcB mRNA were strongly induced under iron-limiting conditions, but transcript levels were low and unresponsive to iron in an irr mutant lacking the global iron response transcriptional regulator Irr. Thus, Irr positively controls the frcB gene. FrcB belongs to a family of previously uncharacterized proteins found in many proteobacteria and some cyanobacteria. This suggests that membrane-bound, heme-containing ferric reductase proteins are not confined to eukaryotes but may be common in bacteria.

| S-EPMC3147698 | biostudies-literature

Bradyrhizobium japonicum has two differentially regulated, functional homologs of the sigma 54 gene (rpoN).

Project description:Recognition of -24/-12-type promoters by RNA polymerase requires a special sigma factor, sigma 54 (RpoN NtrA GlnF). In the nitrogen-fixing soybean symbiont Bradyrhizobium japonicum, two functional, highly conserved rpoN genes (rpoN1 and rpoN2) were identified and sequenced. The two predicted B. japonicum RpoN protein sequences were 87% identical, and both showed different levels of homology to the RpoN proteins of other bacteria. Downstream of rpoN2 (but not of rpoN1), two additional open reading frames were identified that corresponded to open reading frames located at similar positions in Klebsiella pneumoniae and Pseudomonas putida. Both B. japonicum rpoN genes complemented the succinate- and nitrate-negative phenotypes of a Rhizobium meliloti rpoN mutant. B. japonicum strains carrying single or double rpoN mutations were still able to utilize C4-dicarboxylates as a carbon source and histidine, proline, or arginine as a nitrogen source, whereas the ability to assimilate nitrate required expression of at least one of the two rpN genes. In symbiosis both rpoN genes could replace each other functionally. The rpoN1/2 double mutant induced about twice as many nodules on soybeans as did the wild type, and these nodules lacked nitrogen fixation activity completely. Transcription of a nifH'-'lacZ fusion was not activated in the rpoN1/2 mutant background, whereas expression of a fixR'-'lacZ fusion in this mutant was affected only marginally. By using rpoN'-'lacZ fusions, rpoN1 expression was shown to be activated at least sevenfold in microaerobiosis as compared with that in aerobiosis, and this type of regulation involved fixLJ. Expression of rpoN2 was observed under all conditions tested and was increased fivefold in an rpoN2 mutant. The data suggested that the rpoN1 gene was regulated in response to oxygen, whereas the rpoN2 gene was negatively autoregulated.

| S-EPMC207233 | biostudies-other

Identification and characterization of hupT, a gene involved in negative regulation of hydrogen oxidation in Bradyrhizobium japonicum.

Project description:The Bradyrhizobium japonicum hupT gene was sequenced, and its gene product was found to be homologous to NtrB-like histidine kinases. A hupT mutant expresses higher levels of hydrogenase activity than the wild-type strain under hydrogenase-inducing conditions (i.e., microaerobiosis plus hydrogen, or symbiosis), whereas in noninduced hupT cells, hupSL expression is derepressed but does not lead to hydrogenase activity. We conclude that HupT is involved in the repression of HupSL synthesis at the transcriptional level but that enzymatic activation requires inducing conditions.

| S-EPMC94000 | biostudies-literature

A single rRNA gene region in Bradyrhizobium japonicum.

Project description:Bradyrhizobium japonicum contains only a single rRNA (rrn) gene region, despite its comparatively large genome size of 8,700 kb. The nucleotide sequence revealed an organization of rRNA and tRNA genes that is frequently found in bacteria: 5'-rrs (16S rRNA)-ileT (tRNA(Ile))-alaT (tRNA(Ala))-rrl (23S rRNA)-rrf (5S rRNA)-3'. The 5' end of the primary transcript, one of the 16S rRNA processing sites, and the 5' end of the mature 16S rRNA were determined by primer extension. DNA hybridization experiments showed that the slowly growing Bradyrhizobium strains generally have only a single copy of the 16S rRNA gene, whereas the faster-growing Rhizobium species contain three rrs copies.

| S-EPMC177297 | biostudies-other

DNA Microarray-Based Identification of Genes Regulated by NtrC in Bradyrhizobium japonicum.

Project description:The Bradyrhizobium japonicum NtrBC two-component system is a critical regulator of cellular nitrogen metabolism, including the acquisition and catabolism of nitrogenous compounds. To better define the roles of this system, genome-wide transcriptional profiling was performed to identify the NtrC regulon during the response to nitrogen limitation. Upon cells perceiving low intracellular nitrogen, they stimulate the phosphorylation of NtrC, which induces genes responsible for alteration of the core glutamine synthetase/glutamate synthase nitrogen assimilation pathway, including the genes for the glutamine synthetases and PII proteins. In addition, genes responsible for the import and utilization of multiple nitrogen sources, specifically nitrate and nitrite, were upregulated by NtrC activation. Mutational analysis of a candidate nitrite reductase revealed a role for NtrC in regulating the assimilation of nitrite, since mutations in both ntrC and the gene encoding the candidate nitrite reductase abolished the ability to grow on nitrite as a sole nitrogen source.

| S-EPMC4510186 | biostudies-literature

The Bradyrhizobium japonicum proline biosynthesis gene proC is essential for symbiosis.

Project description:Plant host-derived proline is proposed to serve as an energy source for rhizobia in the rhizosphere and in symbiotic root nodules. The Bradyrhizobium japonicum proC gene was isolated, and a proC mutant strain that behaved as a strict proline auxotroph in culture was constructed. The proC strain elicited undeveloped nodules on soybeans that lacked nitrogen fixation activity and plant hemoglobin. We conclude that the proC gene is essential for symbiosis and suggest that the mutant does not obtain an exogenous supply of proline in association with soybeans sufficient to satisfy its auxotrophy.

| S-EPMC92483 | biostudies-literature

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