Analysis of the aerobactin and ferric hydroxamate uptake systems of Yersinia pestis

Forman, Stanislav; Nagiec, Michal J.; Abney, Jennifer; Perry, Robert D.; Fetherston, Jacqueline D.

doi:10.1099/mic.0.2006/004275-0

Cited by 25 publications

(44 citation statements)

References 52 publications

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“…To investigate the involvement of FhuD in the growth defect of the tatC mutant at 37°C and to look into the Tat dependency of FhuD export, we also investigated the growth of these strains in iron-limited medium containing ferrichrome. FhuD functions in the uptake of ferric-hydroxamate-type ferrichrome, which is an iron-chelating siderophore (28). As a control, we also included a ⌬ybtP mutant, as ybtP is a gene required for another independent iron uptake system.…”

Section: Resultsmentioning

confidence: 99%

Transcriptomic and Phenotypic Analysis Reveals New Functions for the Tat Pathway in Yersinia pseudotuberculosis

et al. 2016

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The twin-arginine translocation (Tat) system mediates the secretion of folded proteins that are identified via an N-terminal signal peptide in bacteria, plants, and archaea. Tat systems are associated with virulence in many bacterial pathogens, and our previous studies revealed that Tat-deficient Yersinia pseudotuberculosis was severely attenuated for virulence. Aiming to identify Tat-dependent pathways and phenotypes of relevance for in vivo infection, we analyzed the global transcriptome of parental and ⌬tatC mutant strains of Y. pseudotuberculosis during exponential and stationary growth at 26°C and 37°C. The most significant changes in the transcriptome of the ⌬tatC mutant were seen at 26°C during stationary-phase growth, and these included the altered expression of genes related to virulence, stress responses, and metabolism. Subsequent phenotypic analysis based on these transcriptome changes revealed several novel Tat-dependent phenotypes, including decreased YadA expression, impaired growth under iron-limited and high-copper conditions, as well as acidic pH and SDS. Several functionally related Tat substrates were also verified to contribute to these phenotypes. Interestingly, the phenotypic defects observed in the Tat-deficient strain were generally more pronounced than those in mutants lacking the Tat substrate predicted to contribute to that specific function. Altogether, this provides new insight into the impact of Tat deficiency on in vivo fitness and survival/replication of Y. pseudotuberculosis during infection. IMPORTANCEIn addition to its established role in mediating the secretion of housekeeping enzymes, the Tat system has been recognized as being involved in infection. In some clinically relevant bacteria, such as Pseudomonas spp., several key virulence determinants can readily be identified among the Tat substrates. In enteropathogens, such as Yersinia spp., there are no obvious virulence determinants among the Tat substrates. Tat mutants show no growth defect in vitro but are highly attenuated in in vivo. This makes Tat an attractive target for the development of novel antimicrobials. Therefore, it is important to establish the causes of the attenuation. Here, we show that the attenuation is likely due to synergistic effects of different Tat-dependent phenotypes that each contributes to lowered in vivo fitness. Bacteria have evolved several specialized secretion systems for protein export as part of their successful strategies to colonize niches where they encounter various environmental conditions. Gram-negative bacteria possess different mechanisms to transport/export proteins, including virulence-related factors from the cytoplasm across the inner membrane and/or the outer membrane, either in a single step or acting together with other secretion systems (1). Two pathways, the secretory (Sec) and the twin-arginine translocation (Tat) pathways, are involved in the translocation of proteins from the cytoplasm to the periplasm. In Gramnegative bacteria, they can cooperate with other s...

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Section: Resultsmentioning

confidence: 99%

Transcriptomic and Phenotypic Analysis Reveals New Functions for the Tat Pathway in Yersinia pseudotuberculosis

et al. 2016

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“…We have found that C. difficile encodes a putative Fhu (ferric hydroxamate uptake) ABC transporter system which is expressed under iron-limiting conditions and is repressed by Fur. Fhu transporters have been shown to function in iron acquisition in many Gram-positive as well as Gram-negative bacteria (31)(32)(33)(34). C. difficile also encodes a system homologous to the catecholate siderophore petrobactin ABC transporter Fpi/Ycl from B. subtilis (35).…”

Section: Discussionmentioning

confidence: 99%

Ferric Uptake Regulator Fur Control of Putative Iron Acquisition Systems in Clostridium difficile

Ellermeier

2015

J Bacteriol

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Clostridium difficile is an anaerobic, Gram-positive, spore-forming opportunistic pathogen and is the most common cause of hospital-acquired infectious diarrhea. Although iron acquisition in the host is a key to survival of bacterial pathogens, high levels of intracellular iron can increase oxidative damage. Therefore, expression of iron acquisition mechanisms is tightly controlled by transcriptional regulators. We identified a C. difficile homologue of the master bacterial iron regulator Fur. Using targetron mutagenesis, we generated a fur insertion mutant of C. difficile. To identify the genes regulated by Fur in C. difficile, we used microarray analysis to compare transcriptional differences between the fur mutant and the wild type when grown in highiron medium. The fur mutant had increased expression of greater than 70 transcriptional units. Using quantitative reverse transcriptase PCR (qRT-PCR), we analyzed several of the Fur-regulated genes identified by the microarray and verified that they are both iron and Fur regulated in C. difficile. Among those Fur-and iron-repressed genes were C. difficile genes encoding 7 putative cation transport systems of different classes. We found that Fur was able to bind the DNA upstream of three Fur-repressed genes in electrophoretic mobility shift assays. We also demonstrate that expression of Fur-regulated putative iron acquisition systems was increased during C. difficile infection using the hamster model. Our data suggest that C. difficile expresses multiple iron transport mechanisms in response iron depletion in vitro and in vivo. IMPORTANCEClostridium difficile is the most common cause of hospital-acquired infectious diarrhea and has been recently classified as an "urgent" antibiotic resistance threat by the CDC. To survive and cause disease, most bacterial pathogens must acquire the essential enzymatic cofactor iron. While import of adequate iron is essential for most bacterial growth, excess intracellular iron can lead to extensive oxidative damage. Thus, bacteria must regulate iron import to maintain iron homeostasis. We demonstrate here that C. difficile regulates expression of several putative iron acquisition systems using the transcriptional regulator Fur. These import mechanisms are induced under iron-limiting conditions in vitro and during C. difficile infection of the host. This suggests that during a C. difficile infection, iron availability is limited in vivo.A lmost all living organisms require iron as a cofactor for essential metabolic chemistry (1). Although iron is one of the most abundant of Earth's elements, ferric iron has very limited solubility in aqueous, nonacidic, or oxygenated environments (1). Ferric iron is most often found as iron oxides or hydroxides, which cannot be used by most organisms. Competition over bioavailable iron is fierce both among bacteria in complex microbial communities and between bacterial pathogens and their eukaryotic hosts (2).Organisms have evolved various mechanisms of iron transport to obtain this essentia...

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“…Construction of potentially virulent strains and virulence testing were performed in a CDC-approved biosafety level 3 (BSL3) laboratory following select agent regulations, using procedures approved by the University of Kentucky Institutional Biosafety Committee. Y. pestis strains were transformed with the virulence plasmid pCD1Ap by electroporation (43,45) and plated on TBA plates containing Ap (50 g/ml). The plasmid profile of transformants was analyzed, as well as their phenotype on CR agar (88) and magnesium-oxalate plates (48).…”

Section: Methodsmentioning

confidence: 99%

The Yersiniabactin Transport System Is Critical for the Pathogenesis of Bubonic and Pneumonic Plague

et al. 2010

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Iron acquisition from the host is an important step in the pathogenic process. While Yersinia pestis has multiple iron transporters, the yersiniabactin (Ybt) siderophore-dependent system plays a major role in iron acquisition in vitro and in vivo. In this study, we determined that the Ybt system is required for the use of iron bound by transferrin and lactoferrin and examined the importance of the Ybt system for virulence in mouse models of bubonic and pneumonic plague. Y. pestis mutants unable to either transport Ybt or synthesize the siderophore were both essentially avirulent via subcutaneous injection (bubonic plague model). Surprisingly, via intranasal instillation (pneumonic plague model), we saw a difference in the virulence of Ybt biosynthetic and transport mutants. Ybt biosynthetic mutants displayed an ϳ24-fold-higher 50% lethal dose (LD 50 ) than transport mutants. In contrast, under iron-restricted conditions in vitro, a Ybt transport mutant had a more severe growth defect than the Ybt biosynthetic mutant. Finally, a ⌬pgm mutant had a greater loss of virulence than the Ybt biosynthetic mutant, indicating that the 102-kb pgm locus encodes a virulence factor, in addition to Ybt, that plays a role in the pathogenesis of pneumonic plague.Nearly all organisms require trace amounts of iron. Pathogens must overcome host iron-and heme-binding proteins to cause an infection and disease. The importance of iron acquisition mechanisms has been demonstrated in a number of bacterial pathogens (14,15,27,32,85). Yersinia pestis, the causative agent of plague, has a number of proven and putative iron and heme transport systems. Of these systems, the yersiniabactin (Ybt) siderophore-dependent iron transport system plays a major role in the virulence of bubonic plague in mice (7,8,38,76,79).All identified genes required for the regulation, synthesis, and transport of Ybt, except for ybtD, are carried within a high-pathogenicity island (HPI) that has been spread among enteric pathogens but is essentially identical in the pathogenic yersiniae (13,59,79). In Y. pestis, the ϳ36-kb HPI is located within the 102-kb pgm locus; the entire pgm locus undergoes spontaneous deletion in vitro at a frequency of about 10 Ϫ5 (25, 39, 62). The Ybt system produces a siderophore composed of one salicylate, one thiazoline, and two thiazolidine rings via a nonribosomal peptide/polyketide synthesis mechanism involving high-molecular-weight protein 1 (HMWP1), HMWP2, YbtD, YbtT, YbtE, YbtU, and YbtS (76, 79, 94). The formation constant of this siderophore with ferric iron is 4 ϫ 10 36 , and the crystal structure of the ferric complex has been solved (68, 78).Iron from the Ybt-Fe complex is transported into the cell via the TonB-dependent outer membrane (OM) receptor Psn (which is also required for sensitivity to the bacteriocin pesticin) and an ABC transporter consisting of two inner membrane (IM), fused-function permease/ATP-binding proteins, YbtP and YbtQ. A mutation in any of these three genes prevents Ybt-dependent uptake of iron but does ...

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Analysis of the aerobactin and ferric hydroxamate uptake systems of Yersinia pestis

Cited by 25 publications

References 52 publications

Transcriptomic and Phenotypic Analysis Reveals New Functions for the Tat Pathway in Yersinia pseudotuberculosis

Transcriptomic and Phenotypic Analysis Reveals New Functions for the Tat Pathway in Yersinia pseudotuberculosis

Ferric Uptake Regulator Fur Control of Putative Iron Acquisition Systems in Clostridium difficile

The Yersiniabactin Transport System Is Critical for the Pathogenesis of Bubonic and Pneumonic Plague

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