Phenazines Regulate Nap-Dependent Denitrification in Pseudomonas aeruginosa Biofilms

Lin, Yu Cheng; Sekedat, Matthew D.; Cornell, William Cole; Silva, Gustavo M.; Okegbe, Chinweike; Price-Whelan, Alexa; Vogel, Christine; Dietrich, Lars E. P.

doi:10.1128/jb.00031-18

Cited by 34 publications

(31 citation statements)

References 65 publications

(77 reference statements)

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“…We observe reduced intensity of quinol oxidase staining in the interior of those populated wrinkles (Fig. S4), suggesting that—consistent with previous reports (7, 27)—respiration in the deeper layers of the biofilm respiration occurs anaerobically. Based on the qPCR results (Fig.…”

Section: Resultssupporting

confidence: 91%

Respiratory Heterogeneity Shapes Biofilm Formation and Host Colonization in Uropathogenic Escherichia coli

Beebout

Eberly

Werby

et al. 2019

mBio

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Biofilms are multicellular bacterial communities encased in a self-secreted extracellular matrix comprised of polysaccharides, proteinaceous fibers, and DNA. Organization of these components lends spatial organization to the biofilm community such that biofilm residents can benefit from the production of common goods while being protected from exogenous insults. Spatial organization is driven by the presence of chemical gradients, such as oxygen. Here we show that two quinol oxidases found in Escherichia coli and other bacteria organize along the biofilm oxygen gradient and that this spatially coordinated expression controls architectural integrity. Cytochrome bd, a high-affinity quinol oxidase required for aerobic respiration under hypoxic conditions, is the most abundantly expressed respiratory complex in the biofilm community. Depletion of the cytochrome bd-expressing subpopulation compromises biofilm complexity by reducing the abundance of secreted extracellular matrix as well as increasing cellular sensitivity to exogenous stresses. Interrogation of the distribution of quinol oxidases in the planktonic state revealed that ∼15% of the population expresses cytochrome bd at atmospheric oxygen concentration, and this population dominates during acute urinary tract infection. These data point toward a bet-hedging mechanism in which heterogeneous expression of respiratory complexes ensures respiratory plasticity of E. coli across diverse host niches. IMPORTANCE Biofilms are multicellular bacterial communities encased in a self-secreted extracellular matrix comprised of polysaccharides, proteinaceous fibers, and DNA. Organization of these components lends spatial organization in the biofilm community. Here we demonstrate that oxygen gradients in uropathogenic Escherichia coli (UPEC) biofilms lead to spatially distinct expression programs for quinol oxidases—components of the terminal electron transport chain. Our studies reveal that the cytochrome bd-expressing subpopulation is critical for biofilm development and matrix production. In addition, we show that quinol oxidases are heterogeneously expressed in planktonic populations and that this respiratory heterogeneity provides a fitness advantage during infection. These studies define the contributions of quinol oxidases to biofilm physiology and suggest the presence of respiratory bet-hedging behavior in UPEC.

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Section: Resultssupporting

confidence: 91%

Respiratory Heterogeneity Shapes Biofilm Formation and Host Colonization in Uropathogenic Escherichia coli

Beebout

Eberly

Werby

et al. 2019

mBio

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“…Using their large repertoire of sensory mechanisms and signal transduction pathways, bacteria can be expected to react to these gradients [12,53]. Consistently, a clear stratification of metabolic activities was observed in biofilms of various Gram-negative and Gram-positive bacteria [10,11,54–57]. When grown in macrocolony biofilms, which represent a highly structured type of biofilm (figure 1), E. coli produces the biofilm extracellular matrix only in the upper layer which is more remote from the nutrient-providing agar phase, whereas a dense network of entangled flagella is found in the bottom zone right next to the agar phase [14–16,58,59].…”

Section: Discussionmentioning

confidence: 95%

Spatial organization of different sigma factor activities and c-di-GMP signalling within the three-dimensional landscape of a bacterial biofilm

Klauck¹,

Serra²,

Possling³

et al. 2018

Open Biol.

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Bacterial biofilms are large aggregates of cells embedded in an extracellular matrix of self-produced polymers. In macrocolony biofilms of Escherichia coli, this matrix is generated in the upper biofilm layer only and shows a surprisingly complex supracellular architecture. Stratified matrix production follows the vertical nutrient gradient and requires the stationary phase σS (RpoS) subunit of RNA polymerase and the second messenger c-di-GMP. By visualizing global gene expression patterns with a newly designed fingerprint set of Gfp reporter fusions, our study reveals the spatial order of differential sigma factor activities, stringent control of ribosomal gene expression and c-di-GMP signalling in vertically cryosectioned macrocolony biofilms. Long-range physiological stratification shows a duplication of the growth-to-stationary phase pattern that integrates nutrient and oxygen gradients. In addition, distinct short-range heterogeneity occurs within specific biofilm strata and correlates with visually different zones of the refined matrix architecture. These results introduce a new conceptual framework for the control of biofilm formation and demonstrate that the intriguing extracellular matrix architecture, which determines the emergent physiological and biomechanical properties of biofilms, results from the spatial interplay of global gene regulation and microenvironmental conditions. Overall, mature bacterial macrocolony biofilms thus resemble the highly organized tissues of multicellular organisms.

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“…Aerobic denitrification has been recently suggested to function as a "bet-hedging" strategy to anticipate nitrate availability and to respond to abrupt anoxia (73,74). The oxygen-limited microenvironment in the CF airways is thought to result in P. aeruginosa growth mediated by microaerobic respiration (75).…”

Section: Discussionmentioning

confidence: 99%

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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Phenazines Regulate Nap-Dependent Denitrification in Pseudomonas aeruginosa Biofilms

Cited by 34 publications

References 65 publications

Respiratory Heterogeneity Shapes Biofilm Formation and Host Colonization in Uropathogenic Escherichia coli

Respiratory Heterogeneity Shapes Biofilm Formation and Host Colonization in Uropathogenic Escherichia coli

Spatial organization of different sigma factor activities and c-di-GMP signalling within the three-dimensional landscape of a bacterial biofilm

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

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