Transcriptome analysis of a Pseudomonas aeruginosa sn-glycerol-3-phosphate dehydrogenase mutant reveals a disruption in bioenergetics

Shuman, Jon D.; Giles, Tyler X; Carroll, Leslie P; Tabata, Kenji; Powers, Austin; Suh, Sang-Jin; Silo-Suh, Laura

doi:10.1099/mic.0.000646

Cited by 11 publications

(11 citation statements)

References 57 publications

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“…Acetate and glycerol have different entry points into P. aeruginosa central carbon metabolism and also are thought to have distinct effects on redox metabolism (11)(12)(13)(14). We therefore anticipated a carbon source-specific impact on the expression of enzymes (and corresponding fluxes) involved in the relevant pathways.…”

Section: Resultsmentioning

confidence: 99%

“…For the second and main cultures, PAO1 was grown in MOPS minimal media with 60 mM acetate or 40 mM glycerol as the sole carbon source (120 mM carbon). For 13 C flux experiments, naturally labeled acetate and glycerol were replaced with three separate tracers per carbon source to maximize data set resolution and to accurately determine substrate uptake. Naturally labeled glycerol was replaced with [1,[3][4][5][6][7][8][9][10][11][12][13] Starter cultures were prepared by inoculating LB medium with a loop of freshly plated PAO1.…”

Section: Methodsmentioning

confidence: 99%

“…Much of what we think we know has been gleaned by extrapolation from other bacterial species, and yet those species often occupy niches very different from those occupied by P. aeruginosa and display different substrate preferences. To redress this, in the current work, we used a combination of 'omics approaches (transcriptomics, proteomics, and 13 C fluxomics), coupled with reverse genetics, to systematically investigate the pathway(s) of carbon assimilation during growth on acetate and glycerol. Surprisingly, the presence of these different carbon sources not only leads to a "rewiring" of central metabolism, it also gives rise to profound changes in the levels of expression of pathogenicity-associated functions, including denitrification, redox balance mechanisms, and the electron transport chain (ETC).…”

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confidence: 99%

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Contextual Flexibility in Pseudomonas aeruginosa Central Carbon Metabolism during Growth in Single Carbon Sources

Dolan

Kohlstedt

Trigg

et al. 2020

mBio

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Pseudomonas aeruginosa is an opportunistic human pathogen, particularly noted for causing infections in the lungs of people with cystic fibrosis (CF). Previous studies have shown that the gene expression profile of P. aeruginosa appears to converge toward a common metabolic program as the organism adapts to the CF airway environment. However, we still have only a limited understanding of how these transcriptional changes impact metabolic flux at the systems level. To address this, we analyzed the transcriptome, proteome, and fluxome of P. aeruginosa grown on glycerol or acetate. These carbon sources were chosen because they are the primary breakdown products of an airway surfactant, phosphatidylcholine, which is known to be a major carbon source for P. aeruginosa in CF airways. We show that the fluxes of carbon throughout central metabolism are radically different among carbon sources. For example, the newly recognized “EDEMP cycle” (which incorporates elements of the Entner-Doudoroff [ED] pathway, the Embden-Meyerhof-Parnas [EMP] pathway, and the pentose phosphate [PP] pathway) plays an important role in supplying NADPH during growth on glycerol. In contrast, the EDEMP cycle is attenuated during growth on acetate, and instead, NADPH is primarily supplied by the reaction catalyzed by isocitrate dehydrogenase(s). Perhaps more importantly, our proteomic and transcriptomic analyses revealed a global remodeling of gene expression during growth on the different carbon sources, with unanticipated impacts on aerobic denitrification, electron transport chain architecture, and the redox economy of the cell. Collectively, these data highlight the remarkable metabolic plasticity of P. aeruginosa; that plasticity allows the organism to seamlessly segue between different carbon sources, maximizing the energetic yield from each. IMPORTANCE Pseudomonas aeruginosa is an opportunistic human pathogen that is well known for causing infections in the airways of people with cystic fibrosis. Although it is clear that P. aeruginosa is metabolically well adapted to life in the CF lung, little is currently known about how the organism metabolizes the nutrients available in the airways. In this work, we used a combination of gene expression and isotope tracer (“fluxomic”) analyses to find out exactly where the input carbon goes during growth on two CF-relevant carbon sources, acetate and glycerol (derived from the breakdown of lung surfactant). We found that carbon is routed (“fluxed”) through very different pathways during growth on these substrates and that this is accompanied by an unexpected remodeling of the cell’s electron transfer pathways. Having access to this “blueprint” is important because the metabolism of P. aeruginosa is increasingly being recognized as a target for the development of much-needed antimicrobial agents.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Contextual Flexibility in Pseudomonas aeruginosa Central Carbon Metabolism during Growth in Single Carbon Sources

Dolan

Kohlstedt

Trigg

et al. 2020

mBio

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“…It is interesting that glpD whose product Glycerol-3-phosphate (G3P) dehydrogenase catalyzes G3P transformation in the rpvA mutant is significantly increased, accompanied by a decrease in G3P and dihydroxyacetone phosphate (DHAP) compared with the wild type (Figure 14). Previous studies have confirmed that null mutations in glpD increased the persister formation in P. aeruginosa and E. coli (70, 71). GlpD and G3P synthase (GpsA) encoded by gpsA function at the juncture between respiration and glycolysis, and catalyze the interconversion of G3P and DHAP.…”

Section: Discussionmentioning

confidence: 62%

A Novel LysR-Type Global Regulator RpvA Controls Persister Formation and Virulence inStaphylococcus aureus

Han

Liu

et al. 2019

Preprint

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9Staphylococcus aureus is the leading cause of wound and nosocomial infections. 0Persister formation and virulence factors play crucial roles during S. aureus infection. 1However, the mechanisms of persister formation and its relationship to virulence in S.3 2 aureus are poorly understood. In this study, we screened a transposon mutant library 3 3 and identified a LysR-type global transcriptional regulator NWMN_0037, which we 3 4 called RpvA, for regulator of persistence and virulence, whose mutation leads to 3 5 higher susceptibility to antibiotics ampicillin and norfloxacin and various stresses 3 6including oxidative stress, heat, and starvation in late exponential and early stationary 3 7 phase. Interestingly, the rpvA mutant was highly attenuated for virulence compared 3 8with the parent S. aureus Newman strain as shown by a much higher lethal dose, 3 9 reduced ability to survive in macrophages and to form abscess in the mouse model. 4 0 Transcriptional profiling and metabolomic analysis revealed that RpvA could repress 4 1 multiple genes including gapR, gapA, tpi, pgm, eno, glpD, and acs expression and 4 2 enhance production of numerous intermediate metabolites including 4 3 dihydroxyacetone phosphate, 2-phosphoglycerate, acetyl-CoA, glycerol 3-phosphate, 4 4 L-glutamate in the cells. The differentially expressed genes and altered production of 4 5 metabolites are distributed in global metabolism including carbohydrate metabolism, 4 6 amino acid metabolism, energy metabolism and metabolism of cofactors and vitamins. 4 7These metabolic adjustments could cause the cell to go into dormancy, thus promoting 4 8 S. aureus to convert to persisters. In addition, RpvA could upregulate the expression 4 9 of virulence genes including hla, hlgA, hlgB, hlgC, lukF, lukS, lukD, sea and coa, and 5 0

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“…glpD encodes for a protein involved in glycerol metabolism. Mutations and changes its activity can affect the metabolism of amino acids, induce a stress response with rpoS[168], and lower alignate production making isolates much more virulent[169]. The R274H mutation was in only a single CF strain of PA and 38/639 non-CF strains.MetG, or methionyl-tRNA ligase, is required to initiate and elongate protein synthesis.…”

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confidence: 99%

Genome-wide association study of Pseudomonas aeruginosa adaptations to the cystic fibrosis lung

Noah¹

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Pseudomonas aeruginosa is an opportunistic pathogen that chronically infects the lungs of patients with cystic fibrosis. We know that these bacteria evolve within the lung environment, and have an idea of some of the changes that occur. I performed a genome-wide association study in P. aeruginosa using a variety of analysis methods and several datasets of gene and SNP presence/absence. I determined that the machine learning algorithms random forest and support vector machines performed well on gene presence/absence and core SNP datasets, respectively, when compared with current methods (PLINK and treeWAS). Genes and SNPs already associated with adaptation to the CF lung environment, such as mucA, gyrA, and mex genes, were found with these methods. Some hypothetical and probable proteins were also recovered, and are good candidates for future research.

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Transcriptome analysis of a Pseudomonas aeruginosa sn-glycerol-3-phosphate dehydrogenase mutant reveals a disruption in bioenergetics

Cited by 11 publications

References 57 publications

Contextual Flexibility in Pseudomonas aeruginosa Central Carbon Metabolism during Growth in Single Carbon Sources

Contextual Flexibility in Pseudomonas aeruginosa Central Carbon Metabolism during Growth in Single Carbon Sources

A Novel LysR-Type Global Regulator RpvA Controls Persister Formation and Virulence inStaphylococcus aureus

Genome-wide association study of Pseudomonas aeruginosa adaptations to the cystic fibrosis lung

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