Abstract:A 3.9-kb fragment of the genome of Pseudomonas putida H, containing the complete zwf-pgl-eda-operon, encoding glucose 6-phosphate dehydrogenase, 6-phosphogluconolactonase and 2-keto-3-deoxy-6-phosphogluconate-aldolase, respectively, and part of the divergently transcribed regulatory gene, hexR, was cloned and analyzed. The nucleotide sequences of these genes showed high similarities to the corresponding DNA sequences of P. putida KT2440 and also to sequences of Pseudomonas aeruginosa PAO1. Derivatives of strai… Show more
“…Complementing previous studies on the global hexose metabolism repressor, HexR, in P. putida [21-24], our results suggested that involvement of HexR in regulation of lsc expression might be a selective adaptation of the plant pathogen, P. syringae , to its well-studied infection cycle [42,43]. Once Lsc is secreted, cellular resources needed for its synthesis such as amino acyl residues are not available for the cell anymore.…”
Section: Discussionsupporting
confidence: 80%
“…Analysis of the nucleotide sequences upstream of lscB and lscC , which are almost identical at nucleotide level [2,40], revealed the presence of a sequence (nucleotides +113 to +127 with respect to the corresponding translational start site) with high similarity to the conserved motif TTGTN 7–8 ACAA previously shown to be the DNA binding site of HexR in P. putida [22-24] (Figure 1). This finding suggested a putative binding of HexR of P. syringae to the upstream sequence of both lsc genes.Figure 1
Nucleotide sequence of the upstream region of
lscB
.…”
BackgroundPseudomonas syringae pv. glycinea PG4180 causes bacterial blight on soybean plants and enters the leaf tissue through stomata or open wounds, where it encounters a sucrose-rich milieu. Sucrose is utilized by invading bacteria via the secreted enzyme, levansucrase (Lsc), liberating glucose and forming the polyfructan levan. P. syringae PG4180 possesses two functional lsc alleles transcribed at virulence-promoting low temperatures.ResultsWe hypothesized that transcription of lsc is controlled by the hexose metabolism repressor, HexR, since potential HexR binding sites were identified upstream of both lsc genes. A hexR mutant of PG4180 was significantly growth-impaired when incubated with sucrose or glucose as sole carbon source, but exhibited wild type growth when arabinose was provided. Analyses of lsc expression resulted in higher transcript and protein levels in the hexR mutant as compared to the wild type. The hexR mutant’s ability to multiply in planta was reduced. HexR did not seem to impact hrp gene expression as evidenced by the hexR mutant’s unaltered hypersensitive response in tobacco and its unmodified protein secretion pattern as compared to the wild type under hrp-inducing conditions.ConclusionsOur data suggested a co-regulation of genes involved in extra-cellular sugar acquisition with those involved in intra-cellular energy-providing metabolic pathways in P. syringae.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-015-0349-0) contains supplementary material, which is available to authorized users.
“…Complementing previous studies on the global hexose metabolism repressor, HexR, in P. putida [21-24], our results suggested that involvement of HexR in regulation of lsc expression might be a selective adaptation of the plant pathogen, P. syringae , to its well-studied infection cycle [42,43]. Once Lsc is secreted, cellular resources needed for its synthesis such as amino acyl residues are not available for the cell anymore.…”
Section: Discussionsupporting
confidence: 80%
“…Analysis of the nucleotide sequences upstream of lscB and lscC , which are almost identical at nucleotide level [2,40], revealed the presence of a sequence (nucleotides +113 to +127 with respect to the corresponding translational start site) with high similarity to the conserved motif TTGTN 7–8 ACAA previously shown to be the DNA binding site of HexR in P. putida [22-24] (Figure 1). This finding suggested a putative binding of HexR of P. syringae to the upstream sequence of both lsc genes.Figure 1
Nucleotide sequence of the upstream region of
lscB
.…”
BackgroundPseudomonas syringae pv. glycinea PG4180 causes bacterial blight on soybean plants and enters the leaf tissue through stomata or open wounds, where it encounters a sucrose-rich milieu. Sucrose is utilized by invading bacteria via the secreted enzyme, levansucrase (Lsc), liberating glucose and forming the polyfructan levan. P. syringae PG4180 possesses two functional lsc alleles transcribed at virulence-promoting low temperatures.ResultsWe hypothesized that transcription of lsc is controlled by the hexose metabolism repressor, HexR, since potential HexR binding sites were identified upstream of both lsc genes. A hexR mutant of PG4180 was significantly growth-impaired when incubated with sucrose or glucose as sole carbon source, but exhibited wild type growth when arabinose was provided. Analyses of lsc expression resulted in higher transcript and protein levels in the hexR mutant as compared to the wild type. The hexR mutant’s ability to multiply in planta was reduced. HexR did not seem to impact hrp gene expression as evidenced by the hexR mutant’s unaltered hypersensitive response in tobacco and its unmodified protein secretion pattern as compared to the wild type under hrp-inducing conditions.ConclusionsOur data suggested a co-regulation of genes involved in extra-cellular sugar acquisition with those involved in intra-cellular energy-providing metabolic pathways in P. syringae.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-015-0349-0) contains supplementary material, which is available to authorized users.
“…Unlike the genomes of G. sulfurreducens and most other Geobacteraceae , which encode the enzymes of only the non-oxidative branch of the pentose phosphate pathway, the G. metallireducens genome includes a cluster of oxidative pentose phosphate pathway enzyme genes: 6-phosphogluconolactonase (Gmet_2618, 30% identical to the Pseudomonas putida enzyme [82]), glucose-6-phosphate dehydrogenase (Gmet_2619, 50% identical to the Nostoc punctiforme enzyme [83]), and 6-phosphogluconate dehydrogenase (Gmet_2620, 36% identical to YqeC of B. subtilis [84]), along with two ribose-5-phosphate isomerase isoenzymes (Gmet_2621 and Gmet_1604 = GSU1606, 39% and 44% identical to RpiB of E. coli [85]). Thus, G. metallireducens apparently generates biosynthetic reducing equivalents in the form of NADPH from carbohydrates.…”
BackgroundThe genome sequence of Geobacter metallireducens is the second to be completed from the metal-respiring genus Geobacter, and is compared in this report to that of Geobacter sulfurreducens in order to understand their metabolic, physiological and regulatory similarities and differences.ResultsThe experimentally observed greater metabolic versatility of G. metallireducens versus G. sulfurreducens is borne out by the presence of more numerous genes for metabolism of organic acids including acetate, propionate, and pyruvate. Although G. metallireducens lacks a dicarboxylic acid transporter, it has acquired a second putative succinate dehydrogenase/fumarate reductase complex, suggesting that respiration of fumarate was important until recently in its evolutionary history. Vestiges of the molybdate (ModE) regulon of G. sulfurreducens can be detected in G. metallireducens, which has lost the global regulatory protein ModE but retained some putative ModE-binding sites and multiplied certain genes of molybdenum cofactor biosynthesis. Several enzymes of amino acid metabolism are of different origin in the two species, but significant patterns of gene organization are conserved. Whereas most Geobacteraceae are predicted to obtain biosynthetic reducing equivalents from electron transfer pathways via a ferredoxin oxidoreductase, G. metallireducens can derive them from the oxidative pentose phosphate pathway. In addition to the evidence of greater metabolic versatility, the G. metallireducens genome is also remarkable for the abundance of multicopy nucleotide sequences found in intergenic regions and even within genes.ConclusionThe genomic evidence suggests that metabolism, physiology and regulation of gene expression in G. metallireducens may be dramatically different from other Geobacteraceae.
“…In Pseudomonas species, glycolysis is linked with the EntnerDoudoroff (ED) pathway, which is extensively studied in Pseudomonas putida and contains two operons (zwf-pgl-eda and eddglk-gltR2-gltS) (55). The transcription of these operons is controlled by the repressor HexR, which directly binds to an inverted repeat (TTGTN 7-8 ACAA) in these two promoters, which is released by the specific binding of the ED pathway intermediate 2-keto-3-deoxy-6-phosphogluconate (KDPG) to HexR (24).…”
Section: One-dose-based Microarray Analyses (Highlighted In Red Inmentioning
e Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from human pathogens Staphylococcus aureus and Pseudomonas aeruginosa can be readily inhibited by reactive oxygen species (ROS)-mediated direct oxidation of their catalytic active cysteines. Because of the rapid degradation of H 2 O 2 by bacterial catalase, only steady-state but not one-dose treatment with H 2 O 2 rapidly induces glycolysis and the pentose phosphate pathway (PPP). We conducted transcriptome sequencing (RNA-seq) analyses to globally profile the bacterial transcriptomes in response to a steady level of H 2 O 2 , which revealed profound transcriptional changes, including the induced expression of glycolytic genes in both bacteria. Our results revealed that the inactivation of GAPDH by H 2 O 2 induces metabolic levels of glycolysis and the PPP; the elevated levels of fructose 1,6-biphosphate (FBP) and 2-keto-3-deoxy-6-phosphogluconate (KDPG) lead to dissociation of their corresponding glycolytic repressors (GapR and HexR, respectively) from their cognate promoters, thus resulting in derepression of the glycolytic genes to overcome H 2 O 2 -stalled glycolysis in S. aureus and P. aeruginosa, respectively. Both GapR and HexR may directly sense oxidative stresses, such as menadione.
P athogenic bacteria, such as Pseudomonas aeruginosa andStaphylococcus aureus, need to conquer high concentrations of reactive oxygen species (ROS) that are produced by host phagocytic cells for sustained virulence (1). To this end, these bacteria use ROS-reactive small molecules, such as glutathione (GSH) and melanin in P. aeruginosa as well as coenzyme A and staphyloxanthin in S. aureus. These pathogens also produce a group of ROSdetoxifying enzymes, such as catalase, superoxide dismutase, hydroperoxide reductase, thioredoxin, and glutaredoxin, whose expression is induced by oxidative stress (2-4).It has been well documented that P. aeruginosa and S. aureus also mount global transcriptional changes by utilizing a group of thiol-based ROS-active transcription regulators, such as OxyR, SoxR, MgrA, OhrR, SarA, SarZ, MexR, OspR, CymR, AirSR, and AgrA (5-15). Upon oxidative stress, the specific cysteine groups in these regulatory proteins form sulfenic acids or disulfides, thus inducing conformational changes that attenuate their DNA binding affinities. Our previous work showed that a thiol-based, oxidation-sensing mechanism is utilized by these human pathogens to sense the host immune response and regulate a global change of their properties. ROS leads to activation of defense systems to reduce the oxidative threat as well as a major shift in the life forms of the pathogens (6, 11).ROS can efficiently oxidize the thiol group of active and allosteric cysteines in bacterial proteins, causing changes in their functions. Previously, we employed an isotopic orthogonal proteolysis-activity-based protein profiling (isoTOP-ABPP) technology to identify around 200 oxidation-sensitive cysteines and further determined that several of these proteins perform important redox-active functions...
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