YlaN is an iron(II) binding protein that functions to relieve Fur-mediated repression of gene expression inStaphylococcus aureus (original) (raw)
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Molecular characterization of the ferric-uptake regulator, Fur, from Staphylococcus aureus
Iron is an essential nutrient for the survival and pathogenesis of bacteria, but relatively little is known regarding its transport and regulation in staphylococci. Based on the known sequences of ferric-uptake regulatory (fur) genes from several Gram-positive and Gram-negative bacteria, a fragment containing the fur homologue was cloned from a genomic library of Staphylococcus aureus RN450. Nucleotide sequence analysis of this fragment revealed the presence of a 447 bp ORF that encodes a putative 149 aa polypeptide with an apparent molecular mass of 17 kDa. A putative ferrichrome-uptake (fhu) operon, containing the conserved Fur-binding sequences (Fur box) in the promoter region, was also cloned from the same S. aureus library. To characterize the impact of Fur on the fhu operon, fur was cloned, overexpressed as a His-tagged protein and purified by Ni 2M -affinity column chromatography. The recombinant protein was digested with enterokinase to remove the His tag. Electrophoretic mobility-shift assays indicated that Fur binds to the promoter region of the fhu operon in the presence of divalent cations. Fur also interacted with the promoter region of the recently reported sir operon that has been proposed to constitute a siderophore-transport system in S. aureus. The DNase I-protection assay revealed that Fur specifically binds to the Fur box located in the promoter region of the fhu operon. The primer-extension reaction indicated that the transcription-start site of the fhu operon was located inside the Fur box. S. aureus fur partially complemented a fur N mutation in Bacillus subtilis. The data suggest that Fur regulates iron-transport processes in S. aureus.
Microbiology, 2000
Iron is an essential nutrient for the survival and pathogenesis of bacteria, but relatively little is known regarding its transport and regulation in staphylococci. Based on the known sequences of ferric-uptake regulatory (fur) genes from several Gram-positive and Gram-negative bacteria, a fragment containing the fur homologue was cloned from a genomic library of Staphylococcus aureus RN450. Nucleotide sequence analysis of this fragment revealed the presence of a 447 bp ORF that encodes a putative 149 aa polypeptide with an apparent molecular mass of 17 kDa. A putative ferrichrome-uptake (fhu) operon, containing the conserved Fur-binding sequences (Fur box) in the promoter region, was also cloned from the same S. aureus library. To characterize the impact of Fur on the fhu operon, fur was cloned, overexpressed as a His-tagged protein and purified by Ni 2M-affinity column chromatography. The recombinant protein was digested with enterokinase to remove the His tag. Electrophoretic mobility-shift assays indicated that Fur binds to the promoter region of the fhu operon in the presence of divalent cations. Fur also interacted with the promoter region of the recently reported sir operon that has been proposed to constitute a siderophore-transport system in S. aureus. The DNase I-protection assay revealed that Fur specifically binds to the Fur box located in the promoter region of the fhu operon. The primer-extension reaction indicated that the transcription-start site of the fhu operon was located inside the Fur box. S. aureus fur partially complemented a fur N mutation in Bacillus subtilis. The data suggest that Fur regulates iron-transport processes in S. aureus.
Microbes and Infection, 2006
Staphylococcus aureus can proliferate in iron-limited environments such as the mammalian host. The transcriptional profiles of 460 genes (iron-regulated, putative Fur-regulated, membrane transport, pathogenesis) obtained for S. aureus grown in iron-restricted environments in vitro and in vivo were compared in order to identify new iron-regulated genes and to evaluate their potential as possible therapeutic targets in vivo. Iron deprivation was created in vitro by 2,2-dipyridyl, and in vivo, S. aureus was grown in tissue cages implanted in mice. Bacterial RNA was obtained from each growth condition and cDNA probes were co-hybridized on DNA arrays. Thirty-six upregulated and 11 downregulated genes were commonly modulated in animals and in the low-iron medium. Real-time PCR confirmed the iron-dependent modulation of four novel genes (SACOL0161, 2170, 2369, 2431) with a Fur box motif. Some genes expressed in the dipyridyl medium were not expressed in vivo (e.g., copA, frpA, SACOL1045). Downregulated genes included an iron-storage protein gene and genes of the succinate dehydrogenase complex, reminiscent of a small RNA-dependent regulation thus far only demonstrated in Gram-negative bacteria. The expression of iron-regulated genes in distinct low-iron environments provided insight into their relative importance in vitro and in vivo and their usefulness for vaccine and drug development.
Staphylococcus aureus Redirects Central Metabolism to Increase Iron Availability
PLOS Pathogens, 2006
Staphylococcus aureus pathogenesis is significantly influenced by the iron status of the host. However, the regulatory impact of host iron sources on S. aureus gene expression remains unknown. In this study, we combine multivariable difference gel electrophoresis and mass spectrometry with multivariate statistical analyses to systematically cluster cellular protein response across distinct iron-exposure conditions. Quadruplicate samples were simultaneously analyzed for alterations in protein abundance and/or post-translational modification state in response to environmental (iron chelation, hemin treatment) or genetic (Dfur) alterations in bacterial iron exposure. We identified 120 proteins representing several coordinated biochemical pathways that are affected by changes in iron-exposure status. Highlighted in these experiments is the identification of the heme-regulated transport system (HrtAB), a novel transport system which plays a critical role in staphylococcal heme metabolism. Further, we show that regulated overproduction of acidic end-products brought on by iron starvation decreases local pH resulting in the release of iron from the host iron-sequestering protein transferrin. These findings reveal novel strategies used by S. aureus to acquire scarce nutrients in the hostile host environment and begin to define the iron and heme-dependent regulons of S. aureus. Citation: Friedman DB, Stauff DL, Pishchany G, Whitwell CW, Torres VJ, et al. (2006) Staphylococcus aureus redirects central metabolism to increase iron availability. PLoS Pathog 2(8): e87.
International Journal of Molecular Sciences
Nutritional immunity is a form of innate immunity widespread in both vertebrates and invertebrates. The term refers to a rich repertoire of mechanisms set up by the host to inhibit bacterial proliferation by sequestering trace minerals (mainly iron, but also zinc and manganese). This strategy, selected by evolution, represents an effective front-line defense against pathogens and has thus inspired the exploitation of iron restriction in the development of innovative antimicrobials or enhancers of antimicrobial therapy. This review focuses on the mechanisms of nutritional immunity, the strategies adopted by opportunistic human pathogen Staphylococcus aureus to circumvent it, and the impact of deletion mutants on the fitness, infectivity, and persistence inside the host. This information finally converges in an overview of the current development of inhibitors targeting the different stages of iron uptake, an as-yet unexploited target in the field of antistaphylococcal drug discovery.
Influence of Iron and Aeration on Staphylococcus aureus Growth, Metabolism, and Transcription
Journal of Bacteriology, 2014
Staphylococcus aureus is a prominent nosocomial pathogen and a major cause of biomaterial-associated infections. The success of S. aureus as a pathogen is due in part to its ability to adapt to stressful environments. As an example, the transition from residing in the nares to residing in the blood or deeper tissues is accompanied by changes in the availability of nutrients and elements such as oxygen and iron. As such, nutrients, oxygen, and iron are important determinants of virulence factor synthesis in S. aureus . In addition to influencing virulence factor synthesis, oxygen and iron are critical cofactors in enzymatic and electron transfer reactions; thus, a change in iron or oxygen availability alters the bacterial metabolome. Changes in metabolism create intracellular signals that alter the activity of metabolite-responsive regulators such as CodY, RpiRc, and CcpA. To assess the extent of metabolomic changes associated with oxygen and iron limitation, S. aureus cells were cul...