scholarly journals The mammalian transferrin-independent iron transport system may involve a surface ferrireductase activity

1994 ◽  
Vol 302 (3) ◽  
pp. 875-879 ◽  
Author(s):  
I Jordan ◽  
J Kaplan
Keyword(s):  
Transport System ◽  
Hela Cells ◽  
Iron Uptake ◽  
Iron Transport ◽  
Ferric Iron ◽  
Ferric Citrate ◽  
Ammonium Citrate ◽  
Transport Systems ◽  
Ferric Ammonium Citrate ◽  
Citrate Accumulation

Mammalian cells accumulate iron from ferric citrate or ferric nitrilotriacetate through the activity of a transferrin-independent iron transport system [Sturrock, Alexander, Lamb, Craven and Kaplan (1990) J. Biol. Chem. 265, 3139-3145]. The uptake system might recognize and transport ferric-anion complexes, or cells may reduce ferric iron at the surface and then transport ferrous iron. To distinguish between these possibilities we exposed cells to either [59Fe]ferric citrate or ferric [14C]citrate and determined whether accumulation of iron was accompanied by the obligatory accumulation of citrate. In HeLa cells and human skin fibroblasts the rate of accumulation of iron was three to five times greater than that of citrate. Incubation of fibroblasts with ferric citrate or ferric ammonium citrate resulted in an enhanced accumulation of iron and citrate; the molar ratio of accumulation approaching unity. A similar rate of citrate accumulation, however, was observed when ferric citrate-incubated cells were exposed to [14C]citrate alone. Further studies demonstrated the independence of iron and citrate accumulation: addition of unlabelled citrate to cells decreased the uptake of labelled citrate without affecting the accumulation of 59Fe; iron uptake was decreased by the addition of ferrous chelators whereas the uptake of citrate was unaffected; reduction of ferric iron by ascorbate increased the uptake of iron but had no effect on the uptake of citrate. When HeLa cells were depleted of calcium, iron uptake decreased, but there was little effect on citrate uptake. These results indicate that transport of iron does not require the obligatory transport of citrate and vice versa. The mammalian transferrin-independent iron transport system appears functionally similar to iron transport systems in both the bacterial and plant kingdoms which require the activities of both a surface reductase and a ferrous metal transporter.

scholarly journals Disruption of a Novel Iron Transport System Reverses Oxidative Stress Phenotypes of a dpr Mutant Strain of Streptococcus mutans

2018 ◽  
Vol 200 (14) ◽  
Author(s):  
Tridib Ganguly ◽  
Jessica K. Kajfasz ◽  
James H. Miller ◽  
Eric Rabinowitz ◽  
Lívia C. C. Galvão ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Streptococcus Mutans ◽  
Hydroxyl Radicals ◽  
Transport System ◽  
Iron Uptake ◽  
Iron Transport ◽  
Transport Systems ◽  
Phenotypic Characterization ◽  
Content Type

ABSTRACT The Dps-like peroxide resistance protein (Dpr) is essential for H 2 O 2 stress tolerance and aerobic growth of the oral pathogen Streptococcus mutans . Dpr accumulates during oxidative stress, protecting the cell by sequestering iron ions and thereby preventing the generation of toxic hydroxyl radicals that result from the interaction of iron with H 2 O 2 . Previously, we reported that the SpxA1 and SpxA2 regulators positively regulate expression of dpr in S. mutans . Using an antibody raised against S. mutans Dpr, we confirmed at the protein level the central and cooperative nature of SpxA1 and SpxA2 regulation in Dpr production. During phenotypic characterization of the S. mutans Δ dpr strain, we observed the appearance of distinct colony variants, which sometimes lost the oxidative stress sensitivity typical of Δ dpr strains. Whole-genome sequencing of these phenotypically distinct Δ dpr isolates revealed that a putative iron transporter operon, smu995-smu998 , was a genomic hot spot with multiple single nucleotide polymorphisms identified within the different isolates. Deletion of smu995 or the entire smu995-smu998 operon in the Δ dpr background strain completely reversed the oxidative stress-sensitive phenotypes associated with dpr inactivation. Conversely, inactivation of genes encoding the ferrous iron transport system FeoABC did not alleviate phenotypes of the Δ dpr strain. Preliminary characterization of strains lacking smu995-smu998 , feoABC , and the iron/manganese transporter gene sloABC revealed the interactive nature of these three systems in iron transport but also indicated that there may be additional iron uptake systems in S. mutans . IMPORTANCE The dental caries-associated pathogen Streptococcus mutans routinely encounters oxidative stress within the human plaque biofilm. Previous studies revealed that the iron-binding protein Dpr confers protection toward oxidative stress by limiting free iron availability, which is associated with the generation of toxic hydroxyl radicals. Here, we report the identification of spontaneously occurring mutations within Δ dpr strains. Several of those mutations were mapped to the operon smu995-smu998 , revealing a previously uncharacterized system that appears to be important in iron acquisition. Disruption of the smu995-smu998 operon resulted in reversion of the stress-sensitive phenotype typical of a Δ dpr strain. Our data suggest that the Smu995-Smu998 system works along with other known metal transport systems of S. mutans , i.e., FeoABC and SloABC, to coordinate iron uptake.

scholarly journals Genes Essential to Iron Transport in the Cyanobacterium Synechocystis sp. Strain PCC 6803

2001 ◽  
Vol 183 (9) ◽  
pp. 2779-2784 ◽  
Author(s):  
Hirokazu Katoh ◽  
Natsu Hagino ◽  
Arthur R. Grossman ◽  
Teruo Ogawa
Keyword(s):  
Iron Uptake ◽  
Iron Transport ◽  
Ferric Iron ◽  
Iron Acquisition ◽  
Complete Medium ◽  
Iron Starvation ◽  
Transporter Family ◽  
Genes Encoding ◽  
In Cells ◽  
Pcc 6803

ABSTRACT Genes encoding polypeptides of an ATP binding cassette (ABC)-type ferric iron transporter that plays a major role in iron acquisition inSynechocystis sp. strain PCC 6803 were identified. These genes are slr1295, slr0513, slr0327, and recently reportedsll1878 (Katoh et al., J. Bacteriol. 182:6523–6524, 2000) and were designated futA1, futA2, futB, andfutC, respectively, for their involvement in ferric iron uptake. Inactivation of these genes individually or futA1and futA2 together greatly reduced the activity of ferric iron uptake in cells grown in complete medium or iron-deprived medium. All the fut genes are expressed in cells grown in complete medium, and expression was enhanced by iron starvation. ThefutA1 and futA2 genes appear to encode periplasmic proteins that play a redundant role in iron binding. The deduced products of futB and futC genes contain nucleotide-binding motifs and belong to the ABC transporter family of inner-membrane-bound and membrane-associated proteins, respectively. These results and sequence similarities among the four genes suggest that the Fut system is related to the Sfu/Fbp family of iron transporters. Inactivation of slr1392, a homologue offeoB in Escherichia coli, greatly reduced the activity of ferrous iron transport. This system is induced by intracellular low iron concentrations that are achieved in cells exposed to iron-free medium or in the fut-less mutants grown in complete medium.

scholarly journals Vibrio cholerae VciB Mediates Iron Reduction

2017 ◽  
Vol 199 (12) ◽  
Author(s):  
Eric D. Peng ◽  
Shelley M. Payne
Keyword(s):  
Electron Transport ◽  
Vibrio Cholerae ◽  
Ferrous Iron ◽  
Electron Transport Chain ◽  
Iron Reduction ◽  
Iron Transport ◽  
Ferric Iron ◽  
Transport Systems ◽  
Content Type ◽  
Transport Chain

ABSTRACT Vibrio cholerae is the causative agent of the severe diarrheal disease cholera. V. cholerae thrives within the human host, where it replicates to high numbers, but it also persists within the aquatic environments of ocean and brackish water. To survive within these nutritionally diverse environments, V. cholerae must encode the necessary tools to acquire the essential nutrient iron in all forms it may encounter. A prior study of systems involved in iron transport in V. cholerae revealed the existence of vciB, which, while unable to directly transport iron, stimulates the transport of iron through ferrous (Fe2+) iron transport systems. We demonstrate here a role for VciB in V. cholerae in which VciB stimulates the reduction of Fe3+ to Fe2+, which can be subsequently transported into the cell with the ferrous iron transporter Feo. Iron reduction is independent of functional iron transport but is associated with the electron transport chain. Comparative analysis of VciB orthologs suggests a similar role for other proteins in the VciB family. Our data indicate that VciB is a dimer located in the inner membrane with three transmembrane segments and a large periplasmic loop. Directed mutagenesis of the protein reveals two highly conserved histidine residues required for function. Taken together, our results support a model whereby VciB reduces ferric iron using energy from the electron transport chain. IMPORTANCE Vibrio cholerae is a prolific human pathogen and environmental organism. The acquisition of essential nutrients such as iron is critical for replication, and V. cholerae encodes a number of mechanisms to use iron from diverse environments. Here, we describe the V. cholerae protein VciB that increases the reduction of oxidized ferric iron (Fe3+) to the ferrous form (Fe2+), thus promoting iron acquisition through ferrous iron transporters. Analysis of VciB orthologs in Burkholderia and Aeromonas spp. suggest that they have a similar activity, allowing a functional assignment for this previously uncharacterized protein family. This study builds upon our understanding of proteins known to mediate iron reduction in bacteria.

Chemicals That Stimulate Iron Uptake – A Novel Approach to Treatment of Iron Deficiency,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3180-3180
Author(s):  
Zhen Li
Keyword(s):  
Iron Deficiency ◽  
Small Molecules ◽  
Iron Uptake ◽  
Iron Absorption ◽  
K562 Cells ◽  
Divalent Metal ◽  
Ammonium Citrate ◽  
Ferric Ammonium Citrate ◽  
Caco2 Cells ◽  
Ferric Ammonium

Abstract 3180 Iron (Fe) is an essential nutrient required for all cells, especially for erythrocyte hemoglobin synthesis which requires absorption of 1–2 mg of iron from the gastrointestinal tract. Iron deficiency as a result of inadequate dietary uptake has multiple consequences including anemia and abnormal neurologic development in children and is a global public health concern. Enterocytes in the duodenum, the site of iron absorption, can extract about 10% of dietary Fe. Nonetheless for numerous reasons simple iron supplementation has not solved the worldwide epidemic of iron deficiency. We hypothesized that small molecules which could potentiate iron uptake into cells would allow enterocytes to absorb an increased amount of dietary iron and could be beneficial in limiting iron deficiency. To identify molecules that would accelerate Fe uptake we used a high through-put screening system in conjunction with a reporter system of K562 cells loaded with the divalent metal chelator calcein whose fluorescence is quenched with chelation of Fe2+. Small molecules that stimulated Fe uptake were defined as causing increased calcein fluorescence quenching compared to Fe alone. K562 cells were exposed to 0.1 μM calcein for 10 minutes, thoroughly washed, and 1 × 105 cells plated into each well of multiple 96-well plates. After equilibration of the plates at 37° C, aliquots of the individual components of an in-house chemical library of ∼12,000 compounds dissolved in DMSO were screened in duplicate or triplicate and fluorescence measurements made at 0 and 30 min after addition of 10 μM FeNH4SO4 in a Synergy IV plate reader. 30 chemicals were identified that stimulated iron-induced quenching of calcein fluorescence. The stimulation was verified by dose response curves and by assaying the effect on non-transferrin bound 55Fe uptake. None of the stimulators were cytotoxic for up to at least 3 days. The lead compound, LS081, had an IC50 = 1.22 ± 0.48 μM for 55Fe uptake in K562 cells compared to controls. LS081 was also used to examine the iron uptake in Caco2 cells grown in bicameral chambers, a model system to study intestinal iron absorption. LS081 significantly increased 55Fe uptake into Caco2 cells with a very rapid influx of 55Fe in the first 5 min after Fe was offered to the apical surface followed by a ∼ 4-fold increased uptake over the next 90 min. 55Fe transport across the basolateral surface into the basal chamber also increased ∼ 4 fold. The increased 55Fe transport in caco2 cells is more prominent at lower pH of 5.5 compare to pH 7.5 suggesting LS081 acted on a common divalent metal uptake pathway. Mice treated with LS081 + ferric ammonium citrate via oral gavage for two weeks significantly increased (p < 0.001 by unpaired t-test compared to ferric ammonium citrate alone) the level of ferritin, the iron storage protein, in the liver, demonstrating the absorption of LS081 from intestinal cells. In summary, using high through-put screening technique we identified small molecules that stimulate iron uptake and could be used as a drug for iron deficiency. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Secreted Pyomelanin of Legionella pneumophila Promotes Bacterial Iron Uptake and Growth under Iron-Limiting Conditions

2013 ◽  
Vol 81 (11) ◽  
pp. 4182-4191 ◽  
Author(s):  
Huaixin Zheng ◽  
Christa H. Chatfield ◽  
Mark R. Liles ◽  
Nicholas P. Cianciotto
Keyword(s):  
Legionella Pneumophila ◽  
Ferrous Iron ◽  
Iron Uptake ◽  
Iron Transport ◽  
Ferric Iron ◽  
Iron Acquisition ◽  
Ferric Hydroxide ◽  
Supernatant Fraction ◽  
Content Type ◽  
Ferrous Iron Transport

ABSTRACTIron acquisition is critical to the growth and virulence ofLegionella pneumophila. Previously, we found thatL. pneumophilauses both a ferrisiderophore pathway and ferrous iron transport to obtain iron. We now report that two molecules secreted byL. pneumophila, homogentisic acid (HGA) and its polymerized variant (HGA-melanin, a pyomelanin), are able to directly mediate the reduction of various ferric iron salts. Furthermore, HGA, synthetic HGA-melanin, and HGA-melanin derived from bacterial supernatants enhanced the ability ofL. pneumophilaand other species ofLegionellato take up radiolabeled iron. Enhanced iron uptake was not observed with a ferrous iron transport mutant. Thus, HGA and HGA-melanin mediate ferric iron reduction, with the resulting ferrous iron being available to the bacterium for uptake. Upon further testing ofL. pneumophilaculture supernatants, we found that significant amounts of ferric and ferrous iron were associated with secreted HGA-melanin. Importantly, a pyomelanin-containing fraction obtained from a wild-type culture supernatant was able to stimulate the growth of iron-starved legionellae. That the corresponding supernatant fraction obtained from a nonpigmented mutant culture did not stimulate growth demonstrated that HGA-melanin is able to both promote iron uptake and enhance growth under iron-limiting conditions. Indicative of a complementary role in iron acquisition, HGA-melanin levels were inversely related to the levels of siderophore activity. Compatible with a role in the ecology and pathogenesis ofL. pneumophila, HGA and HGA-melanin were effective at reducing and releasing iron from both insoluble ferric hydroxide and the mammalian iron chelates ferritin and transferrin.

scholarly journals Ferric citrate decreases ruminal hydrogen sulphide concentrations in feedlot cattle fed diets high in sulphate

2013 ◽  
Vol 111 (2) ◽  
pp. 261-269 ◽  
Author(s):  
Mary E. Drewnoski ◽  
Perry Doane ◽  
Stephanie L. Hansen
Keyword(s):  
Hydrogen Sulphide ◽  
Rumen Fluid ◽  
Control Diet ◽  
Gas Production ◽  
Feedlot Cattle ◽  
Ferric Citrate ◽  
Ammonium Citrate ◽  
Ferric Ammonium Citrate ◽  
Ferric Ammonium

Dissimilatory reduction of sulphate by sulphate-reducing bacteria in the rumen produces sulphide, which can lead to a build-up of the toxic gas hydrogen sulphide (H2S) in the rumen when increased concentrations of sulphate are consumed by ruminants. We hypothesised that adding ferric Fe would competitively inhibit ruminal sulphate reduction. The effects of five concentrations and two sources (ferric citrate or ferric ammonium citrate) of ferric Fe were examinedin vitro(n6 per treatment). Rumen fluid was collected from a steer that was adapted to a high-concentrate, high-sulphate diet (0·51 % S). The addition of either source of ferric Fe decreased (P< 0·01) H2S concentrations without affecting gas production (P= 0·38), fluid pH (P= 0·80) orin vitroDM digestibility (P= 0·38) after a 24 h incubation. Anin vivoexperiment was conducted using eight ruminally fistulated steers (543 (sem12) kg) in a replicated Latin square with four periods and four treatments. The treatments included a high-concentrate, high-sulphate control diet (0·46 % S) or the control diet plus ferric ammonium citrate at concentrations of 200, 300 or 400 mg Fe/kg diet DM. The inclusion of ferric Fe did not affect DM intake (P= 0·21). There was a linear (P< 0·01) decrease in the concentration of ruminal H2S as the addition of ferric Fe concentrations increased. Ferric citrate appears to be an effective way to decrease ruminal H2S concentrations, which could allow producers to safely increase the inclusion of ethanol co-products.

scholarly journals Characterization of Ferric and Ferrous Iron Transport Systems in Vibrio cholerae

2006 ◽  
Vol 188 (18) ◽  
pp. 6515-6523 ◽  
Author(s):  
Elizabeth E. Wyckoff ◽  
Alexandra R. Mey ◽  
Andreas Leimbach ◽  
Carolyn F. Fisher ◽  
Shelley M. Payne
Keyword(s):  
Vibrio Cholerae ◽  
Transport System ◽  
Iron Transport ◽  
Iron Acquisition ◽  
Shigella Flexneri ◽  
Transport Systems ◽  
Infant Mouse ◽  
Mouse Intestine ◽  
Dependent Transport

ABSTRACT Vibrio cholerae has multiple iron acquisition systems, including TonB-dependent transport of heme and of the catechol siderophore vibriobactin. Strains defective in both of these systems grow well in laboratory media and in the infant mouse intestine, indicating the presence of additional iron acquisition systems. Previously uncharacterized potential iron transport systems, including a homologue of the ferrous transporter Feo and a periplasmic binding protein-dependent ATP binding cassette (ABC) transport system, termed Fbp, were identified in the V. cholerae genome sequence. Clones encoding either the Feo or the Fbp system exhibited characteristics of iron transporters: both repressed the expression of lacZ cloned under the control of a Fur-regulated promoter in Escherichia coli and also conferred growth on a Shigella flexneri mutant that has a severe defect in iron transport. Two other ABC transporters were also evaluated but were negative by these assays. Transport of radioactive iron by the Feo system into the S. flexneri iron transport mutant was stimulated by the reducing agent ascorbate, consistent with Feo functioning as a ferrous transporter. Conversely, ascorbate inhibited transport by the Fbp system, suggesting that it transports ferric iron. The growth of V. cholerae strains carrying mutations in one or more of the potential iron transport genes indicated that both Feo and Fbp contribute to iron acquisition. However, a mutant defective in the vibriobactin, Fbp, and Feo systems was not attenuated in a suckling mouse model, suggesting that at least one other iron transport system can be used in vivo.

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