Azithromycin Inhibits Expression of the GacA-Dependent Small RNAs RsmY and RsmZ in Pseudomonas aeruginosa

Pérez‐Martínez, Isabel; Haas, Dieter

doi:10.1128/aac.01801-10

Cited by 68 publications

(67 citation statements)

References 49 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Data from this study and others have shown that rsmYZ transcription requires RsmA (Fig. 4B) (57). We also found that plasmid-expressed RsmA results in a significant increase in expression of the P rsmY-lacZ and P rsmZ-lacZ reporters.…”

Section: Discussionsupporting

confidence: 61%

See 1 more Smart Citation

The AlgZR Two-Component System Recalibrates the RsmAYZ Posttranscriptional Regulatory System To Inhibit Expression of the Pseudomonas aeruginosa Type III Secretion System

Intile

Diaz

Urbanowski

et al. 2013

Journal of Bacteriology

124

View full text Add to dashboard Cite

bPseudomonas aeruginosa causes chronic airway infections in cystic fibrosis (CF) patients. A classic feature of CF airway isolates is the mucoid phenotype. Mucoidy arises through mutation of the mucA anti-sigma factor and subsequent activation of the AlgU regulon. Inactivation of mucA also results in reduced expression of the Vfr transcription factor. Vfr regulates several important virulence factors, including a type III secretion system (T3SS). In the present study, we report that ExsA expression, the master regulator of T3SS gene expression, is further reduced in mucA mutants through a Vfr-independent mechanism involving the RsmAYZ regulatory system. RsmA is an RNA binding protein required for T3SS gene expression. Genetic experiments suggest that the AlgZR two-component system, part of the AlgU regulon, inhibits ExsA expression by increasing the expression of RsmY and RsmZ, two small noncoding RNAs that sequester RsmA from target mRNAs. Epistasis analyses revealed that increasing the concentration of free RsmA, through either rsmYZ deletion or increased RsmA expression, partially restored T3SS gene expression in the mucA mutant. Furthermore, increasing RsmA availability in combination with Vfr complementation fully restored T3SS expression. Recalibration of the RsmAYZ system by AlgZR, however, did not alter the expression of other selected RsmAdependent targets. We account for this observation by showing that ExsA expression is more sensitive to changes in free RsmA than other members of the RsmA regulon. Together, these data indicate that recalibration of the RsmAYZ system partially accounts for reduced T3SS gene expression in mucA mutants. Pseudomonas aeruginosa is a Gram-negative bacterium with the ability to cause both acute and chronic infections (1, 2). Immunocompromised individuals often develop acute P. aeruginosa infections that can become systemic, with lethal sepsis as a common outcome (3). Isolates from acute P. aeruginosa infections are generally characterized by the presence of a functional type III secretion system (T3SS) (4, 5). The T3SS injects multiple effectors directly into the host cell cytoplasm, where they function to deregulate signal transduction mechanisms and promote phagocytic avoidance and host cell killing (6-8). In contrast to acute infections, the airways of most adults with the genetic disorder cystic fibrosis (CF) are chronically colonized with P. aeruginosa. CF patients are initially colonized with environmental strains of P. aeruginosa that subsequently undergo diversification via mutation and selection by the CF airway environment to generate a population of cells adapted for persistence, host immune defense, antibiotic resistance, and biofilm formation (9-11).A common phenotype observed in CF isolates is the loss of T3SS gene expression (12, 13). One mechanism that accounts for reduced T3SS expression involves the MucA/AlgU signal transduction system (see Fig. 8 for a model). MucA is a membranebound anti-sigma factor that sequesters the alternative sigma factor AlgU to c...

show abstract

Section: Discussionsupporting

confidence: 61%

“…4B) (57). We observed the same using P rsmY-lacZ and P rsmZ-lacZ transcriptional reporters and also saw that plasmid-expressed RsmA results in a significant increase in P rsmY-lacZ and P rsmZ-lacZ reporter activities (Fig.…”

Section: Figsupporting

confidence: 60%

The AlgZR Two-Component System Recalibrates the RsmAYZ Posttranscriptional Regulatory System To Inhibit Expression of the Pseudomonas aeruginosa Type III Secretion System

Intile

Diaz

Urbanowski

et al. 2013

Journal of Bacteriology

124

View full text Add to dashboard Cite

show abstract

“…Previous works aiming to elucidate QS inhibitory mechanisms have reported that the QS inhibitor azithromycin inhibits the expression of rsmY and rsmZ transcription through the PA0588-PA0584 gene cluster (31). However, our transcriptomic data did not show that iberin affected the expression of the PA0588-PA0584 gene cluster, which suggests that iberin uses a different mechanism from that of azithromycin to inhibit the expression of rsmY and rsmZ.…”

Section: Resultscontrasting

confidence: 53%

Comparative Systems Biology Analysis To Study the Mode of Action of the Isothiocyanate Compound Iberin on Pseudomonas aeruginosa

Tan

Liu

Chua

et al. 2014

Antimicrob Agents Chemother

View full text Add to dashboard Cite

Food is now recognized as a natural resource of novel antimicrobial agents, including those that target the virulence mechanisms of bacterial pathogens. Iberin, an isothiocyanate compound from horseradish, was recently identified as a quorum-sensing inhibitor (QSI) of the bacterial pathogen Pseudomonas aeruginosa. In this study, we used a comparative systems biology approach to unravel the molecular mechanisms of the effects of iberin on QS and virulence factor expression of P. aeruginosa. Our study shows that the two systems biology methods used (i.e., RNA sequencing and proteomics) complement each other and provide a thorough overview of the impact of iberin on P. aeruginosa. RNA sequencing-based transcriptomics showed that iberin inhibits the expression of the GacA-dependent small regulatory RNAs RsmY and RsmZ; this was verified by using gfp-based transcriptional reporter fusions with the rsmY or rsmZ promoter regions. Isobaric tags for relative and absolute quantitation (iTRAQ) proteomics showed that iberin reduces the abundance of the LadS protein, an activator of GacS. Taken together, the findings suggest that the mode of QS inhibition in iberin is through downregulation of the Gac/Rsm QS network, which in turn leads to the repression of QS-regulated virulence factors, such as pyoverdine, chitinase, and protease IV. Lastly, as expected from the observed repression of small regulatory RNA synthesis, we also show that iberin effectively reduces biofilm formation. This suggests that small regulatory RNAs might serve as potential targets in the future development of therapies against pathogens that use QS for controlling virulence factor expression and assume the biofilm mode of growth in the process of causing disease. Q uorum sensing (QS) is a cell-to-cell communication system widely distributed among bacteria in which small diffusible signal molecules are employed to regulate gene expression in a dose-dependent manner (1). After reaching a threshold concentration, the QS signal molecules will bind to and activate their receptors, which results in a coordinated population expression of QS-regulated genes. These genes include those that upregulate the synthesis of QS signal molecules (autoinduction) but, more importantly, they also include genes that encode virulence factors required for bacterial infections (2). Thus, QS inhibitors (QSIs) have been proposed as antipathogenic agents and have been shown to attenuate the capability of pathogens to cause infections (3, 4). QSIs possess different modes of action, including interfering with the synthesis of quorum-sensing signaling molecules (5) or competitively binding to the QS signal receptors (6). The regulation of the bacterial QS systems is complex, and this further expands the targets for the design of novel QSIs (7,8).Isothiocyanates (ITCs) are biologically active compounds found in cruciferous vegetables and have gained research interest as cancer chemopreventive agents (9). Sulforaphane (SFN) (10), allyl isothiocyanate (AITC) (11), and phenethyl isothioc...

show abstract

“…Two bands hybridized to the rsmY probe, as has been seen previously ( Fig. 5D) (30,38). Little difference was seen between a ΔalgR mutant strain and the wild-type PAO1 (data not shown).…”

Section: Resultsmentioning

confidence: 50%

“…The regulation of rsmZ is more complex than rsmY and includes regulators other than GacA (38,40,41). Primer extension analysis confirmed the location of the transcriptional start site in P. aeruginosa that was predicted from P. fluorescens (Fig.…”

Section: Resultsmentioning

confidence: 53%

The Pseudomonas aeruginosa Two-Component Regulator AlgR Directly Activates rsmA Expression in a Phosphorylation-Independent Manner

2017

View full text Add to dashboard Cite

Pseudomonas aeruginosa is an important pathogen of the immunocompromised, causing both acute and chronic infections. In cystic fibrosis (CF) patients, P. aeruginosa causes chronic disease. The impressive sensory network of P. aeruginosa allows the bacterium to sense and respond to a variety of stimuli found in diverse environments. Transcriptional regulators, including alternative sigma factors and response regulators, integrate signals changing gene expression, allowing P. aeruginosa to cause infection. The two-component transcriptional regulator AlgR is important in P. aeruginosa pathogenesis in both acute and chronic infections. In chronic infections, AlgR and the alternative sigma factor AlgU activate the genes responsible for alginate production. Previous work demonstrated that AlgU controls rsmA expression. RsmA is a posttranscriptional regulator that is antagonized by two small RNAs, RsmY and RsmZ. In this work, we demonstrate that AlgR directly activates rsmA expression from the same promoter as AlgU. In addition, phosphorylation was not necessary for AlgR activation of rsmA using algR and algZ mutant strains. AlgU and AlgR appear to affect the antagonizing small RNAs rsmY and rsmZ indirectly. RsmA was active in a mucA22 mutant strain using leader fusions of two RsmA targets, tssA1 and hcnA. AlgU and AlgR were necessary for posttranscriptional regulation of tssA1 and hcnA. Altogether, our work demonstrates that the alginate regulators AlgU and AlgR are important in the control of the RsmA posttranscriptional regulatory system. These findings suggest that RsmA plays an unknown role in mucoid strains due to AlgU and AlgR activities.IMPORTANCE P. aeruginosa infections are difficult to treat and frequently cause significant mortality in CF patients. Understanding the mechanisms of persistence is important. Our work has demonstrated that the alginate regulatory system also significantly impacts the posttranscriptional regulator system RsmA/Y/Z. We demonstrate that AlgR directly activates rsmA expression, and this impacts the RsmA regulon. This leads to the possibility that the RsmA/Y/Z system plays a role in helping P. aeruginosa persist during chronic infection. In addition, this furthers our understanding of the reach of the alginate regulators AlgU and AlgR.KEYWORDS P. aeruginosa, RsmA, AlgR, mucoid, Pseudomonas aeruginosa, cystic fibrosis, mucA, two-component regulatory systems T he opportunistic pathogen Pseudomonas aeruginosa possesses multiple virulence factors for causing disease. This allows P. aeruginosa to cause both acute and chronic infections. Acute infecting strains are characterized by the presence of type IV pili (T4P), flagella, and a type III secretion system (T3SS) (1-3). In contrast, chronic infecting strains diversify (4, 5) and frequently do not express T3SS, T4P, or flagella (6, 7). Chronic infecting strains often form biofilms composed of exopolysaccharides, such as alginate, and signal a decline in lung function in CF patients (8-11). Alginate

show abstract

Azithromycin Inhibits Expression of the GacA-Dependent Small RNAs RsmY and RsmZ in Pseudomonas aeruginosa

Cited by 68 publications

References 49 publications

The AlgZR Two-Component System Recalibrates the RsmAYZ Posttranscriptional Regulatory System To Inhibit Expression of the Pseudomonas aeruginosa Type III Secretion System

The AlgZR Two-Component System Recalibrates the RsmAYZ Posttranscriptional Regulatory System To Inhibit Expression of the Pseudomonas aeruginosa Type III Secretion System

Comparative Systems Biology Analysis To Study the Mode of Action of the Isothiocyanate Compound Iberin on Pseudomonas aeruginosa

The Pseudomonas aeruginosa Two-Component Regulator AlgR Directly Activates rsmA Expression in a Phosphorylation-Independent Manner

Contact Info

Product

Resources

About