The Diguanylate Cyclase SadC Is a Central Player in Gac/Rsm-Mediated Biofilm Formation in Pseudomonas aeruginosa

Moscoso, Joana A.; Jaeger, Tina; Valentini, Martina; Hui, Kailyn; Jenal, Urs; Filloux, Alain

doi:10.1128/jb.01850-14

Cited by 91 publications

(101 citation statements)

References 60 publications

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“…These results are similar to those described for double mutants lacking each of these DGCs in combination with removal of BifA (21). SadC is also a primary contributor to the c-di-GMP pool that promotes matrix production when the Gac/Rsm regulatory cascade is active in the P. aeruginosa strain PAK (34).…”

Section: Colony Matrix Distribution Phenotypes Suggest That Phenazinesupporting

confidence: 76%

Electron-shuttling antibiotics structure bacterial communities by modulating cellular levels of c-di-GMP

Okegbe

Fields

Cole

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

101

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Diverse organisms secrete redox-active antibiotics, which can be used as extracellular electron shuttles by resistant microbes. Shuttlemediated metabolism can support survival when substrates are available not locally but rather at a distance. Such conditions arise in multicellular communities, where the formation of chemical gradients leads to resource limitation for cells at depth. In the pathogenic bacterium Pseudomonas aeruginosa PA14, antibiotics called phenazines act as oxidants to balance the intracellular redox state of cells in anoxic biofilm subzones. PA14 colony biofilms show a profound morphogenic response to phenazines resulting from electron acceptor-dependent inhibition of ECM production. This effect is reminiscent of the developmental responses of some eukaryotic systems to redox control, but for bacterial systems its mechanistic basis has not been well defined. Here, we identify the regulatory protein RmcA and show that it links redox conditions to PA14 colony morphogenesis by modulating levels of bis-(3′,5′)-cyclic-dimeric-guanosine (c-di-GMP), a second messenger that stimulates matrix production, in response to phenazine availability. RmcA contains four Per-Arnt-Sim (PAS) domains and domains with the potential to catalyze the synthesis and degradation of c-di-GMP. Our results suggest that phenazine production modulates RmcA activity such that the protein degrades c-di-GMP and thereby inhibits matrix production during oxidizing conditions. RmcA thus forms a mechanistic link between cellular redox sensing and community morphogenesis analogous to the functions performed by PAS-domain-containing regulatory proteins found in complex eukaryotes.W hen microbial cells cannot import or are physically separated from metabolic electron donors or acceptors, diffusible compounds can act as electron carriers and support survival on these substrates (1, 2). These conditions arise in the presence of poised-potential electrodes or insoluble minerals, such as iron oxides (3-5), and in multicellular communities (biofilms) where the formation of chemical gradients leads to oxidant limitation for cells at depth (6-9). Diverse microbes secrete redoxactive compounds with the capacity to function as electron shuttles (10-12). In the pathogenic bacterium Pseudomonas aeruginosa PA14, electron-shuttling antibiotics called phenazines support survival on poised-potential electrodes and balance the intracellular redox state of cells in anoxic biofilm subzones (1, 9).Similar to those formed by many species of microbes, colonies of PA14 develop intricate wrinkle structures on agar-solidified growth media (13). Phenazines profoundly alter PA14 colony morphogenesis, inhibiting the onset of wrinkle formation and changing the organization of wrinkles (12) (Fig. 1A). Modeling of resource availability within colonies suggests that the earlier increase in the colony surface area-to-volume ratio in phenazine-null (Δphz) mutants maximizes access to oxygen for cells that would otherwise become limited for oxidant (14). Measurements of...

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Section: Colony Matrix Distribution Phenotypes Suggest That Phenazinesupporting

confidence: 76%

Electron-shuttling antibiotics structure bacterial communities by modulating cellular levels of c-di-GMP

Okegbe

Fields

Cole

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

101

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“…Through overexpression of DGC's and PDE's Moscoso et al provided evidence that the switch between type III and type VI secretion occurs as a direct result of changes in c-di-GMP metabolism. It was subsequently demonstrated that the SadC DGC is a central link between the RetS/GacS/LadS system and c-di-GMP signaling in P. aeruginosa (51). In the present study, we have added to these findings by demonstrating that c-di-GMP metabolism may change the expression of virulence factors through interference with the VfrcAMP signaling system.…”

Section: Discussionmentioning

confidence: 73%

The Cyclic AMP-Vfr Signaling Pathway in Pseudomonas aeruginosa Is Inhibited by Cyclic Di-GMP

Almblad¹,

Harrison²,

Rybtke³

et al. 2015

J. Bacteriol.

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The opportunistic human pathogen Pseudomonas aeruginosa expresses numerous acute virulence factors in the initial phase of infection, and during long-term colonization it undergoes adaptations that optimize survival in the human host. Adaptive changes that often occur during chronic infection give rise to rugose small colony variants (RSCVs), which are hyper-biofilmforming mutants that commonly possess mutations that increase production of the biofilm-promoting secondary messenger cyclic di-GMP (c-di-GMP). We show that RSCVs display a decreased production of acute virulence factors as a direct result of elevated c-di-GMP content. Overproduction of c-di-GMP causes a decrease in the transcription of virulence factor genes that are regulated by the global virulence regulator Vfr. The low level of Vfr-dependent transcription is caused by a low level of its coactivator, cyclic AMP (cAMP), which is decreased in response to a high level of c-di-GMP. Mutations that cause reversion of the RSCV phenotype concomitantly reactivate Vfr-cAMP signaling. Attempts to uncover the mechanism underlying the observed c-di-GMP-mediated lowering of cAMP content provided evidence that it is not caused by inhibition of adenylate cyclase production or activity and that it is not caused by activation of cAMP phosphodiesterase activity. In addition to the studies of the RSCVs, we present evidence that the deeper layers of wild-type P. aeruginosa biofilms have high c-di-GMP levels and low cAMP levels. IMPORTANCEOur work suggests that cross talk between c-di-GMP and cAMP signaling pathways results in downregulation of acute virulence factors in P. aeruginosa biofilm infections. Knowledge about this cross-regulation adds to our understanding of virulence traits and immune evasion by P. aeruginosa in chronic infections and may provide new approaches to eradicate biofilm infections.

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“…The hptB gene encodes a histidine phosphotransfer protein involved in a complex regulatory cascade (47)(48)(49). The ⌬hptB mutant of P. aeruginosa PAK has been demonstrated to produce hyperbiofilms and to show elevated Pel polysaccharide production and elevated c-di-GMP levels (48,50). By mass spectrometry analysis, we found that, in the P. aeruginosa PA14 strain, the ⌬hptB mutant produces 4.3-fold more c-di-GMP than the wild type, while the ⌬bifA mutant produces 12.8-fold more c-di-GMP than the wild type.…”

Section: Resultsmentioning

confidence: 99%

PilZ Domain Protein FlgZ Mediates Cyclic Di-GMP-Dependent Swarming Motility Control in Pseudomonas aeruginosa

et al. 2016

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The second messenger cyclic diguanylate (c-di-GMP) is an important regulator of motility in many bacterial species. In Pseudomonas aeruginosa, elevated levels of c-di-GMP promote biofilm formation and repress flagellum-driven swarming motility. The rotation of P. aeruginosa's polar flagellum is controlled by two distinct stator complexes, MotAB, which cannot support swarming motility, and MotCD, which promotes swarming motility. Here we show that when c-di-GMP levels are elevated, swarming motility is repressed by the PilZ domain-containing protein FlgZ and by Pel polysaccharide production. We demonstrate that FlgZ interacts specifically with the motility-promoting stator protein MotC in a c-di-GMP-dependent manner and that a functional green fluorescent protein (GFP)-FlgZ fusion protein shows significantly reduced polar localization in a strain lacking the MotCD stator. Our results establish FlgZ as a c-di-GMP receptor affecting swarming motility by P. aeruginosa and support a model wherein c-di-GMP-bound FlgZ impedes motility via its interaction with the MotCD stator. IMPORTANCEThe regulation of surface-associated motility plays an important role in bacterial surface colonization and biofilm formation. c-di-GMP signaling is a widespread means of controlling bacterial motility, and yet the mechanism whereby this signal controls surface-associated motility in P. aeruginosa remains poorly understood. Here we identify a PilZ domain-containing c-di-GMP effector protein that contributes to c-di-GMP-mediated repression of swarming motility by P. aeruginosa. We provide evidence that this effector, FlgZ, impacts swarming motility via its interactions with flagellar stator protein MotC. Thus, we propose a new mechanism for c-di-GMP-mediated regulation of motility for a bacterium with two flagellar stator sets, increasing our understanding of surface-associated behaviors, a key prerequisite to identifying ways to control the formation of biofilm communities. Cyclic diguanylate (c-di-GMP) is a ubiquitous bacterial second messenger responsible for regulating a range of cellular processes, including motility and biofilm formation (1). In general, low intracellular c-di-GMP levels are associated with motile lifestyles, while elevated levels of c-di-GMP promote surface attachment and sessile lifestyles (2, 3). c-di-GMP is synthesized from two molecules of GTP by diguanylate cyclases (DGCs) and degraded by c-di-GMP-specific phosphodiesterases (PDEs) (1, 4). Many DGCs and PDEs involved in motility regulation have been characterized, but the mechanisms by which c-di-GMP regulates motility are poorly understood in Pseudomonas aeruginosa and in other bacterial species.To regulate numerous biological functions, c-di-GMP binds to specific effector proteins or RNA (reviewed in reference 5). Recent studies have focused on identifying these c-di-GMP effectors and their mechanisms for regulating c-di-GMP-dependent processes. One class of effectors is the PilZ domain-containing protein family, which is characterized by conserved ...

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The Diguanylate Cyclase SadC Is a Central Player in Gac/Rsm-Mediated Biofilm Formation in Pseudomonas aeruginosa

Cited by 91 publications

References 60 publications

Electron-shuttling antibiotics structure bacterial communities by modulating cellular levels of c-di-GMP

Electron-shuttling antibiotics structure bacterial communities by modulating cellular levels of c-di-GMP

The Cyclic AMP-Vfr Signaling Pathway in Pseudomonas aeruginosa Is Inhibited by Cyclic Di-GMP

PilZ Domain Protein FlgZ Mediates Cyclic Di-GMP-Dependent Swarming Motility Control in Pseudomonas aeruginosa

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