SuhB Regulates the Motile-Sessile Switch in Pseudomonas aeruginosa through the Gac/Rsm Pathway and c-di-GMP Signaling.
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ABSTRACT:
SUBMITTER: Li K
PROVIDER: S-EPMC5462983 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:In Pseudomonas aeruginosa, the second messenger cyclic-di-GMP and Gac/Rsm signaling pathways are associated with the transition from acute to chronic infection. Therefore, identification of the molecular mechanisms that govern lifestyle choice in bacteria is very important. Here, we identified a novel cyclic-di-GMP modulator, PrtR, which was shown to repress pyocin production by inhibition of PrtN and activate the type III secretion system (T3SS) through PtrB. Compared to a wild-type strain or a prtN mutant, the prtR prtN double mutant exhibited a wrinkly colony and hyperbiofilm phenotype, as well as an increase in intracellular c-di-GMP levels. Interestingly, a diguanylate cyclase (DGC) gene, siaD, was repressed by PrtR. Further experiments revealed that PrtR directly interacts with SiaD and facilitates the accumulation of c-di-GMP in cells. We also demonstrated that PrtR regulates the activity of the Gac/Rsm system, thus affecting expression of the T3SS and type VI secretion system (T6SS) and the formation of biofilm. Taken together, the present findings indicate that PrtR, as a c-di-GMP modulator, plays key roles in the adaptation to opportunistic infection of P. aeruginosa Additionally, this study revealed a novel mechanism for PrtR-mediated regulation of the lifestyle transition via the Gac/Rsm and c-di-GMP signaling networks.
| S-EPMC7822137 | biostudies-literature
Project description:The interaction of Pseudomonas aeruginosa with surfaces has been described as a two-stage process requiring distinct signaling events and the reciprocal modulation of small RNAs (sRNAs). However, little is known regarding the relationship between sRNA-modulating pathways active under planktonic or surface-associated growth conditions. Here, we demonstrate that SagS (PA2824), the cognate sensor of HptB, links sRNA-modulating activities via the Gac/HptB/Rsm system postattachment to the signal transduction network BfiSR, previously demonstrated to be required for the development of P. aeruginosa. Consistent with the role of SagS in the GacA-dependent HtpB signaling pathway, inactivation of sagS resulted in hyperattachment, an HptB-dependent increase in rsmYZ, increased Psl polysaccharide production, and increased virulence. Moreover, sagS inactivation rescued attachment but abrogated biofilm formation by the ?gacA and ?hptB mutant strains. The ?sagS strain was impaired in biofilm formation at a stage similar to that of the previously described two-component system BfiSR. Expression of bfiR but not bfiS restored ?sagS biofilm formation independently of rsmYZ. We demonstrate that SagS interacts directly with BfiS and only indirectly with BfiR, with the direct and specific interaction between these two membrane-bound sensors resulting in the modulation of the phosphorylation state of BfiS in a growth-mode-dependent manner. SagS plays an important role in P. aeruginosa virulence in a manner opposite to that of BfiS. Our findings indicate that SagS acts as a switch by linking the GacA-dependent sensory system under planktonic conditions to the suppression of sRNAs postattachment and to BfiSR, required for the development of P. aeruginosa biofilms, in a sequential and stage-specific manner.
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Project description:Pseudomonas aeruginosa is a Gram-negative opportunistic human pathogen and a threat for immunocompromised and cystic fibrosis patients. It is responsible for acute and chronic infections and can switch between these lifestyles upon taking an informed decision involving complex regulatory networks. The RetS/LadS/Gac/Rsm network and the cyclic-di-GMP (c-di-GMP) signaling pathways are both central to this phenomenon redirecting the P. aeruginosa population toward a biofilm mode of growth, which is associated with chronic infections. While these two pathways were traditionally studied independently from each other, we recently showed that cellular levels of c-di-GMP are increased in the hyperbiofilm retS mutant. Here, we have formally established the link between the two networks by showing that the SadC diguanylate cyclase is central to the Gac/Rsm-associated phenotypes, notably, biofilm formation. Importantly, SadC is involved in the signaling that converges onto the RsmA translational repressor either via RetS/LadS or via HptB/HsbR. Although the level of expression of the sadC gene does not seem to be impacted by the regulatory cascade, the production of the SadC protein is tightly repressed by RsmA. This adds to the growing complexity of the signaling network associated with c-di-GMP in P. aeruginosa. While this organism possesses more than 40 c-di-GMP-related enzymes, it remains unclear how signaling specificity is maintained within the c-di-GMP network. The finding that SadC but no other diguanylate cyclase is related to the formation of biofilm governed by the Gac/Rsm pathway further contributes to understanding of this insulation mechanism.
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Project description:The cyclic di-GMP (c-di-GMP) second messenger represents a signaling system that regulates many bacterial behaviors and is of key importance for driving the lifestyle switch between motile loner cells and biofilm formers. This review provides an up-to-date compendium of c-di-GMP pathways connected to biofilm formation, biofilm-associated motilities, and other functionalities in the ubiquitous and opportunistic human pathogen Pseudomonas aeruginosa This bacterium is frequently adopted as a model organism to study bacterial biofilm formation. Importantly, its versatility and adaptation capabilities are linked with a broad range of complex regulatory networks, including a large set of genes involved in c-di-GMP biosynthesis, degradation, and transmission.
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| S-EPMC7682387 | biostudies-literature
Project description:Flagellar motility is critical for surface attachment and biofilm formation in many bacteria. A key regulator of flagellar motility in Pseudomonas aeruginosa and other microbes is cyclic diguanylate (c-di-GMP). High levels of this second messenger repress motility and stimulate biofilm formation. c-di-GMP levels regulate motility in P. aeruginosa in part by influencing the localization of its two flagellar stator sets, MotAB and MotCD. Here, we show that while c-di-GMP can influence stator localization, stators can in turn impact c-di-GMP levels. We demonstrate that the swarming motility-driving stator MotC physically interacts with the transmembrane region of the diguanylate cyclase SadC. Furthermore, we demonstrate that this interaction is capable of stimulating SadC activity. We propose a model by which the MotCD stator set interacts with SadC to stimulate c-di-GMP production under conditions not permissive to motility. This regulation implies a positive-feedback loop in which c-di-GMP signaling events cause MotCD stators to disengage from the motor; then disengaged stators stimulate c-di-GMP production to reinforce a biofilm mode of growth. Our studies help to define the bidirectional interactions between c-di-GMP and the flagellar machinery.IMPORTANCE The ability of bacterial cells to control motility during early steps in biofilm formation is critical for the transition to a nonmotile, biofilm lifestyle. Recent studies have clearly demonstrated the ability of c-di-GMP to control motility via a number of mechanisms, including through controlling transcription of motility-related genes and modulating motor function. Here, we provide evidence that motor components can in turn impact c-di-GMP levels. We propose that communication between motor components and the c-di-GMP synthesis machinery allows the cell to have a robust and sensitive switching mechanism to control motility during early events in biofilm formation.
| S-EPMC6707927 | biostudies-literature
Project description:C-di-GMP signaling can directly influence bacterial behavior by affecting the functionality of c-di-GMP-binding proteins. In addition, c-di-GMP can exert an indirect and more global effect on gene transcription or translation, e.g. via riboswitches or by binding to transcription factors. In this study, we investigated the effects of changes in intracellular c-di-GMP levels on gene expression and protein production in the opportunistic pathogen Pseudomonas aeruginosa. We induced c-di-GMP production via an ectopically introduced diguanylate cyclase and recorded the transcriptional, translational as well as proteomic profile of the cells. We demonstrate that rising levels of c-di-GMP in P. aeruginosa under growth conditions otherwise characterized by low c-di-GMP levels immediately cause a switch to a non-motile, auto-aggregative phenotype. This switch became apparent before any c-di-GMP-dependent role on transcription, translation, or protein abundance could be observed. Our results indicate that sudden rises in global c-di-GMP pools affect the P. aeruginosa phenotype via an alteration of protein functionality, rather than an impact on global gene transcription or translation.
2024-09-03 | GSE183394 | GEO
Project description:The signaling molecule cyclic di-GMP (cdG) controls the switch between bacterial motility and biofilm production, and fluctuations in cellular levels of cdG have been implicated in Vibrio cholerae pathogenesis. Intracellular concentrations of cdG are controlled by the interplay of diguanylate cyclase (DGC) enzymes, which synthesize cdG to promote biofilms, and phosphodiesterase (PDE) enzymes, which hydrolyse cdG to drive motility. To track the complete regulatory logic of how V. cholerae responds to changing cdG levels, we followed a time course of overexpression of either the V. harveyi diguanylate cyclase QrgB or a variant of QrgB lacking catalytic activity (QrgB*). We find that QrgB increases cdG levels relative to QrgB* for 30 minutes after overexpression, but the effect of QrgB on cdG levels plateaus at 30 minutes, indicating tight adaptive control of cdG levels. In contrast, loss of VpsR, a master regulator activating biofilm formation upon binding to cdG, leads to higher baseline levels of cdG and continuously increasing cdG through 60 minutes after QrgB induction, revealing the existence of a negative feedback loop on cdG levels operating through VpsR. Through a combination of RNA polymerase ChIP-seq, RNA-seq, and genetic approaches, we show that the PDE CdgC is activated by VpsR at high cdG concentrations, mediating this negative feedback on cdG levels. We further identify a transcript encoded within, and antisense to, the cdgC open reading frame which we name sRNA negative regulator of CdgC (SnrC). RNA polymerase ChIP-seq and RNA-seq demonstrate SnrC to be expressed specifically under conditions of high cdG in the absence of VpsR. Ectopic SnrC expression increases cdG levels in a manner depending on CdgC, demonstrating that its effect on cdG levels is likely through interference with CdgC production. Further, although cells lacking cdgC exhibit enhanced biofilm formation, these mutants are outcompeted by wild type V. cholerae in colonization assays that reward a combination of attachment, dispersal, and motility. These results underscore the importance of negative feedback regulation of cdG to maintain appropriate homeostatic levels for efficient transitioning between biofilm formation and motility, both of which are necessary over the course of the V. cholerae infection cycle.
2024-10-09 | GSE274216 | GEO
Project description:The signaling molecule cyclic di-GMP (cdG) controls the switch between bacterial motility and biofilm production, and fluctuations in cellular levels of cdG have been implicated in Vibrio cholerae pathogenesis. Intracellular concentrations of cdG are controlled by the interplay of diguanylate cyclase (DGC) enzymes, which synthesize cdG to promote biofilms, and phosphodiesterase (PDE) enzymes, which hydrolyse cdG to drive motility. To track the complete regulatory logic of how V. cholerae responds to changing cdG levels, we followed a time course of overexpression of either the V. harveyi diguanylate cyclase QrgB or a variant of QrgB lacking catalytic activity (QrgB*). We find that QrgB increases cdG levels relative to QrgB* for 30 minutes after overexpression, but the effect of QrgB on cdG levels plateaus at 30 minutes, indicating tight adaptive control of cdG levels. In contrast, loss of VpsR, a master regulator activating biofilm formation upon binding to cdG, leads to higher baseline levels of cdG and continuously increasing cdG through 60 minutes after QrgB induction, revealing the existence of a negative feedback loop on cdG levels operating through VpsR. Through a combination of RNA polymerase ChIP-seq, RNA-seq, and genetic approaches, we show that the PDE CdgC is activated by VpsR at high cdG concentrations, mediating this negative feedback on cdG levels. We further identify a transcript encoded within, and antisense to, the cdgC open reading frame which we name sRNA negative regulator of CdgC (SnrC). RNA polymerase ChIP-seq and RNA-seq demonstrate SnrC to be expressed specifically under conditions of high cdG in the absence of VpsR. Ectopic SnrC expression increases cdG levels in a manner depending on CdgC, demonstrating that its effect on cdG levels is likely through interference with CdgC production. Further, although cells lacking cdgC exhibit enhanced biofilm formation, these mutants are outcompeted by wild type V. cholerae in colonization assays that reward a combination of attachment, dispersal, and motility. These results underscore the importance of negative feedback regulation of cdG to maintain appropriate homeostatic levels for efficient transitioning between biofilm formation and motility, both of which are necessary over the course of the V. cholerae infection cycle.
2024-10-09 | GSE274215 | GEO
Project description:The transcriptional regulator BrlR is a member of the MerR family of multidrug transport activators that contributes to the high-level drug tolerance of Pseudomonas aeruginosa biofilms. While MerR regulators are known to activate both the expression of multidrug efflux pump genes and their own transcription upon inducer binding, little is known about BrlR activation. We demonstrate using promoter reporter strains, in vivo and in vitro?DNA-binding assays combined with 5'RACE, that BrlR binds to its own promoter, likely via a MerR-like palindromic sequence. Unlike known MerR multidrug transport activators, BrlR and brlR expression are not activated by multidrug transporter substrates. Instead, BrlR-DNA binding was enhanced by the secondary messenger c-di-GMP. In addition to enhanced BrlR-DNA binding, c-di-GMP levels contributed to PbrlR promoter activity in initial attached cells with elevated c-di-GMP levels correlating with increased expression of brlR. While not harbouring amino acid motifs resembling previously defined c-di-GMP-binding domains, BrlR was found to bind c-di-GMP?in vitro at a ratio of one c-di-GMP per two BrlR. Cross-linking assays confirmed dimer formation to be enhanced in the presence of elevated c-di-GMP levels. Our findings demonstrate BrlR to be an unusual MerR-family member in that BrlR function and expression require the secondary messenger c-di-GMP.
| S-EPMC4000578 | biostudies-literature