The TolC Protein of Legionella pneumophila Plays a Major Role in Multi-Drug Resistance and the Early Steps of Host Invasion

Ferhat, Mourad; Atlan, Danièle; Vianney, Anne; Lazzaroni, Jean-Claude; Doublet, Patricia; Gilbert, Christophe

doi:10.1371/journal.pone.0007732

Cited by 49 publications

(29 citation statements)

References 41 publications

(47 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Jacobi and Heuner identified two genes, lssB and lssD, of L. pneumophila as part of a putative T1SS, based on similarity with toxin transporter systems of Vibrio cholerae, Salmonella typhi, and E. coli (14). Moreover, we showed in a previous study that a tolC gene is present in the L. pneumophila genome and that it is involved in early steps of host invasion, as a ⌬tolC mutant exhibits impaired virulence for amoebas and macrophages (32). A similar phenotype has also been observed in a ⌬rtxA mutant strain (26,27).…”

Section: Resultsmentioning

confidence: 61%

Functional Type 1 Secretion System Involved in Legionella pneumophila Virulence

et al. 2014

Self Cite

View full text Add to dashboard Cite

e Legionella pneumophila is a Gram-negative pathogen found mainly in water, either in a free-living form or within infected protozoans, where it replicates. This bacterium can also infect humans by inhalation of contaminated aerosols, causing a severe form of pneumonia called legionellosis or Legionnaires' disease. The involvement of type II and IV secretion systems in the virulence of L. pneumophila is now well documented. Despite bioinformatic studies showing that a type I secretion system (T1SS) could be present in this pathogen, the functionality of this system based on the LssB, LssD, and TolC proteins has never been established. Here, we report the demonstration of the functionality of the T1SS, as well as its role in the infectious cycle of L. pneumophila. Using deletion mutants and fusion proteins, we demonstrated that the repeats-in-toxin protein RtxA is secreted through an LssB-LssD-TolC-dependent mechanism. Moreover, fluorescence monitoring and confocal microscopy showed that this T1SS is required for entry into the host cell, although it seems dispensable to the intracellular cycle. Together, these results underline the active participation of L. pneumophila, via its T1SS, in its internalization into host cells.

show abstract

Section: Resultsmentioning

confidence: 61%

Functional Type 1 Secretion System Involved in Legionella pneumophila Virulence

et al. 2014

Self Cite

View full text Add to dashboard Cite

show abstract

“…To obtain L. pneumophila Lens mutants defective for the lpl0780 gene, a homologous-recombination strategy was chosen as previously described (25) The 400-bp upstream and downstream regions of the gene of interest were amplified by PCR, digested by NotI, and then cloned into pAV695. A kanamycin resistance cassette was amplified from pKD13 (with AV80 and AV81 primers [see Table S3 in the supplemental material]), digested by SalI, and cloned inside the encoding sequence of the gene in pAV695.…”

Section: Methodsmentioning

confidence: 99%

Three Antagonistic Cyclic di-GMP-Catabolizing Enzymes Promote Differential Dot/Icm Effector Delivery and Intracellular Survival at the Early Steps of Legionella pneumophila Infection

et al. 2014

Self Cite

View full text Add to dashboard Cite

g Legionella pneumophila is an intracellular pathogen which replicates within protozoan cells and can accidently infect alveolar macrophages, causing an acute pneumonia in humans. The second messenger cyclic di-GMP (c-di-GMP) has been shown to play key roles in the regulation of various bacterial processes, including virulence. While investigating the function of the 22 potential c-di-GMP-metabolizing enzymes of the L. pneumophila Lens strain, we found three that directly contribute to its ability to infect both protozoan and mammalian cells. These three enzymes display diguanylate cyclase (Lpl0780), phosphodiesterase (Lpl1118), and bifunctional diguanylate cyclase/phosphodiesterase (Lpl0922) activities, which are all required for the survival and intracellular replication of L. pneumophila. Mutants with deletions of the corresponding genes are efficiently taken up by phagocytic cells but are partially defective for the escape of the Legionella-containing vacuole (LCV) from the host degradative endocytic pathway and result in lower survival. In addition, Lpl1118 is required for efficient endoplasmic reticulum recruitment to the LCV. Trafficking and biogenesis of the LCV are dependent upon the orchestrated actions of several type 4 secretion system Dot/ Icm effectors proteins, which exhibit differentially altered translocation in the three mutants. While translocation of some effectors remained unchanged, others appeared over-and undertranslocated. A general translocation offset of the large repertoire of Dot/Icm effectors may be responsible for the observed defects in the trafficking and biogenesis of the LCV. Our results suggest that L. pneumophila uses cyclic di-GMP signaling to fine-tune effector delivery and ensure effective evasion of the host degradative pathways and establishment of a replicative vacuole. C yclic-di-GMP (c-di-GMP) is a nearly ubiquitous bacterial second messenger involved in signaling pathways that control lifestyle transitions, among which are the transition from the motile to the sessile state and, for pathogenic bacteria, the transition from pathogenic to environmental lifestyles or from acute/planktonic infections to chronic/biofilm infections (1-4). c-di-GMP is produced from GTP by diguanylate cyclases (DGCs), is degraded by phosphodiesterases (PDEs), and acts through various effector proteins or RNA riboswitches (5-8). The catalytic site of DGCs is associated with the GGDEF domain, while PDE activity involves either the EAL or the HD-GYP domains. Genes encoding these proteins have been identified in all major bacterial phyla and are particularly expanded in gammaproteobacteria, with a median of 22 genes per genome (9). Consistently, the genomes of gammaproteobacterium Legionella pneumophila strains encode from 22 to 24 GGDEF/EAL proteins (10, 11). Few are specific to each strain, but most of them are highly conserved in L. pneumophila strains (11). c-di-GMP seems to be strongly involved in Legionella cell signaling, since the 22 GGDEF/EAL proteins represent 13.5% of proteins contain...

show abstract

“…However, the fact that in vitro exposure of Legionella to erythromycin or ciprofloxacin selects mostly lowlevel resistance may be an indication of efflux participation (360), while target mutation-derived moxifloxacin resistance causes high-level MIC increases (8-to 512-fold) (361). The genomes of L. pneumophila strains (3.4 to 3.5 Mb) are smaller than that of E. coli K-12 (4.64 Mb) but still possess a large number of genes encoding efflux pumps, membrane fusion proteins, and OM channel proteins (362,363). Inactivation of the TolC efflux channel (36% identity to E. coli TolC) increases the susceptibility to erythromycin (16-fold reduction in the MIC) and to benzalkonium chloride, deoxycholate, ethidium bromide, norfloxacin, novobiocin, and rhodamine 6G (2-to 8-fold decreases in MIC values) (362,363).…”

Section: Legionella Sppmentioning

confidence: 99%

“…The genomes of L. pneumophila strains (3.4 to 3.5 Mb) are smaller than that of E. coli K-12 (4.64 Mb) but still possess a large number of genes encoding efflux pumps, membrane fusion proteins, and OM channel proteins (362,363). Inactivation of the TolC efflux channel (36% identity to E. coli TolC) increases the susceptibility to erythromycin (16-fold reduction in the MIC) and to benzalkonium chloride, deoxycholate, ethidium bromide, norfloxacin, novobiocin, and rhodamine 6G (2-to 8-fold decreases in MIC values) (362,363). Moreover, TolC is also involved in the secretion of a lipid-containing surfactant that promotes Legionella motility and displays activity against Legionella (363).…”

Section: Legionella Sppmentioning

confidence: 99%

“…Inactivation of the TolC efflux channel (36% identity to E. coli TolC) increases the susceptibility to erythromycin (16-fold reduction in the MIC) and to benzalkonium chloride, deoxycholate, ethidium bromide, norfloxacin, novobiocin, and rhodamine 6G (2-to 8-fold decreases in MIC values) (362,363). Moreover, TolC is also involved in the secretion of a lipid-containing surfactant that promotes Legionella motility and displays activity against Legionella (363). Given that intracellular killing of L. pneumophila by antibiotics is required for legionellosis therapy, drug efflux could be an important factor affecting the efficacy of in vivo antibiotic regimens.…”

Section: Legionella Sppmentioning

confidence: 99%

See 1 more Smart Citation

The Challenge of Efflux-Mediated Antibiotic Resistance in Gram-Negative Bacteria

2015

View full text Add to dashboard Cite

SUMMARY The global emergence of multidrug-resistant Gram-negative bacteria is a growing threat to antibiotic therapy. The chromosomally encoded drug efflux mechanisms that are ubiquitous in these bacteria greatly contribute to antibiotic resistance and present a major challenge for antibiotic development. Multidrug pumps, particularly those represented by the clinically relevant AcrAB-TolC and Mex pumps of the resistance-nodulation-division (RND) superfamily, not only mediate intrinsic and acquired multidrug resistance (MDR) but also are involved in other functions, including the bacterial stress response and pathogenicity. Additionally, efflux pumps interact synergistically with other resistance mechanisms (e.g., with the outer membrane permeability barrier) to increase resistance levels. Since the discovery of RND pumps in the early 1990s, remarkable scientific and technological advances have allowed for an in-depth understanding of the structural and biochemical basis, substrate profiles, molecular regulation, and inhibition of MDR pumps. However, the development of clinically useful efflux pump inhibitors and/or new antibiotics that can bypass pump effects continues to be a challenge. Plasmid-borne efflux pump genes (including those for RND pumps) have increasingly been identified. This article highlights the recent progress obtained for organisms of clinical significance, together with methodological considerations for the characterization of MDR pumps.

show abstract

The TolC Protein of Legionella pneumophila Plays a Major Role in Multi-Drug Resistance and the Early Steps of Host Invasion

Cited by 49 publications

References 41 publications

Functional Type 1 Secretion System Involved in Legionella pneumophila Virulence

Functional Type 1 Secretion System Involved in Legionella pneumophila Virulence

Three Antagonistic Cyclic di-GMP-Catabolizing Enzymes Promote Differential Dot/Icm Effector Delivery and Intracellular Survival at the Early Steps of Legionella pneumophila Infection

The Challenge of Efflux-Mediated Antibiotic Resistance in Gram-Negative Bacteria

Contact Info

Product

Resources

About