Characterization of the iron transporter DMT1 (NRAMP2/DCT1) in red blood cells of normal and anemic mk/mkmice

Blood ◽  
2001 ◽  
Vol 98 (13) ◽  
pp. 3823-3830 ◽  
Author(s):  
François Canonne-Hergaux ◽  
An-Sheng Zhang ◽  
Prem Ponka ◽  
Philippe Gros
Keyword(s):  
Iron Uptake ◽  
Iron Acquisition ◽  
Uptake System ◽  
Erythroid Cells ◽  
Iron Release ◽  
Loss Of Function ◽  
Divalent Metal Transporter 1 ◽  
Peripheral Tissues ◽  
Divalent Metal Transporter ◽  
In Erythroid Cells

Abstract Divalent metal transporter 1 (DMT1) is the major transferrin-independent iron uptake system at the apical pole of intestinal cells, but it may also transport iron across the membrane of acidified endosomes in peripheral tissues. Iron transport and expression of the 2 isoforms of DMT1 was studied in erythroid cells that consume large quantities of iron for biosynthesis of hemoglobin. In mk/mk mice that express a loss-of-function mutant variant of DMT1, reticulocytes have a decreased cellular iron uptake and iron incorporation into heme. Interestingly, iron release from transferrin inside the endosome is normal in mk/mkreticulocytes, suggesting a subsequent defect in Fe++ transport across the endosomal membrane. Studies by immunoblotting using membrane fractions from peripheral blood or spleen from normal mice where reticulocytosis was induced by erythropoietin (EPO) or phenylhydrazine (PHZ) treatment suggest that DMT1 is coexpressed with transferrin receptor (TfR) in erythroid cells. Coexpression of DMT1 and TfR in reticulocytes was also detected by double immunofluorescence and confocal microscopy. Experiments with isoform-specific anti-DMT1 antiserum strongly suggest that it is the non–iron-response element containing isoform II of DMT1 that is predominantly expressed by the erythroid cells. As opposed to wild-type reticulocytes, mk/mk reticulocytes express little if any DMT1, despite robust expression of TfR, suggesting a possible effect of the mutation on stability and targeting of DMT1 isoform II in these cells. Together, these results provide further evidence that DMT1 plays a central role in iron acquisition via the transferrin cycle in erythroid cells.

scholarly journals Regulation of divalent metal transporter 1 (DMT1) non-IRE isoform by the microRNA Let-7d in erythroid cells

Haematologica ◽  
2010 ◽  
Vol 95 (8) ◽  
pp. 1244-1252 ◽  
Author(s):  
I. Andolfo ◽  
L. De Falco ◽  
R. Asci ◽  
R. Russo ◽  
S. Colucci ◽  
...  
Keyword(s):  
Divalent Metal ◽  
Erythroid Cells ◽  
Divalent Metal Transporter 1 ◽  
Divalent Metal Transporter ◽  
Metal Transporter ◽  
In Erythroid Cells

Copper repletion enhances apical iron uptake and transepithelial iron transport by Caco-2 cells

2002 ◽  
Vol 282 (3) ◽  
pp. G527-G533 ◽  
Author(s):  
Okhee Han ◽  
Marianne Wessling-Resnick
Keyword(s):  
Iron Uptake ◽  
Iron Transport ◽  
Cell Monolayer ◽  
Transepithelial Transport ◽  
Apical Surface ◽  
Divalent Metal Transporter 1 ◽  
Divalent Metal Transporter ◽  
Metal Transporter ◽  
Copper Status ◽  
Copper Supplementation

The influence of copper status on Caco-2 cell apical iron uptake and transepithelial transport was examined. Cells grown for 7–8 days in media supplemented with 1 μM CuCl2had 10-fold higher cellular levels of copper compared with control. Copper supplementation did not affect the integrity of differentiated Caco-2 cell monolayers grown on microporous membranes. Copper-repleted cells displayed increased uptake of iron as well as increased transport of iron across the cell monolayer. Northern blot analysis revealed that expression of the apical iron transporter divalent metal transporter-1 (DMT1), the basolateral transporter ferroportin-1 (Fpn1), and the putative ferroxidase hephaestin (Heph) was upregulated by copper supplementation, whereas the recently identified ferrireductase duodenal cytochrome b (Dcytb) was not. These results suggest that DMT1, Fpn1, and Heph are involved in the iron uptake process modulated by copper status. Although a clear role for Dcytb was not identified, an apical surface ferrireductase was modulated by copper status, suggesting that its function also contributes to the enhanced iron uptake by copper-repleted cells. A model is proposed wherein copper promotes iron depletion of intestinal Caco-2 cells, creating a deficiency state that induces upregulation of iron transport factors.

scholarly journals The Divalent Metal Transporter 1 (DMT1) Is Required for Iron Uptake and Normal Development of Oligodendrocyte Progenitor Cells

2018 ◽  
Vol 38 (43) ◽  
pp. 9142-9159 ◽  
Author(s):  
Veronica T. Cheli ◽  
Diara A. Santiago González ◽  
Leandro N. Marziali ◽  
Norma N. Zamora ◽  
María E. Guitart ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Iron Uptake ◽  
Normal Development ◽  
Divalent Metal ◽  
Oligodendrocyte Progenitor Cells ◽  
Divalent Metal Transporter 1 ◽  
Oligodendrocyte Progenitor ◽  
Divalent Metal Transporter ◽  
Metal Transporter

scholarly journals Hepcidin inhibits apical iron uptake in intestinal cells

2008 ◽  
Vol 294 (1) ◽  
pp. G192-G198 ◽  
Author(s):  
Natalia P. Mena ◽  
Andrés Esparza ◽  
Victoria Tapia ◽  
Pamela Valdés ◽  
Marco T. Núñez
Keyword(s):  
Iron Uptake ◽  
Basolateral Membrane ◽  
Intestinal Cells ◽  
Divalent Metal Transporter 1 ◽  
Divalent Metal Transporter ◽  
Cell Monolayers ◽  
Metal Transporter ◽  
Protein Levels ◽  
J774 Cells ◽  
Iron Export

Hepcidin (Hepc) is considered a key mediator in iron trafficking. Although the mechanism of Hepc action in macrophages is fairly well established, much less is known about its action in intestinal cells, one of the main targets of Hepc. The current study investigated the effects of physiologically generated Hepc on iron transport in Caco-2 cell monolayers and rat duodenal segments compared with the effects on the J774 macrophage cell line. Addition of Hepc to Caco-2 cells or rat duodenal segments strongly inhibited apical 55Fe uptake without apparent effects on the transfer of 55Fe from the cells to the basolateral medium. Concurrently, the levels of divalent metal transporter 1 (DMT1) mRNA and protein in Caco-2 cells decreased while the mRNA and protein levels of the iron export transporter ferroportin did not change. Plasma membrane localization of ferroportin was studied by selective biotinylation of apical and basolateral membrane domains; Hepc induced rapid internalization of ferroportin in J774 cells but not in Caco-2 cells These results indicate that the effect of Hepc is cell dependent: in macrophages it inhibits iron export by inducing ferroportin degradation, whereas in enterocytes it inhibits apical iron uptake by inhibiting DMT1 transcription. Our results highlight the crucial role of Hepc in the control of intestinal iron absorption.

Functional consequences of the human DMT1 (SLC11A2) mutation on protein expression and iron uptake

Blood ◽  
2005 ◽  
Vol 106 (12) ◽  
pp. 3985-3987 ◽  
Author(s):  
Monika Priwitzerova ◽  
Guangjun Nie ◽  
Alex D. Sheftel ◽  
Dagmar Pospisilova ◽  
Vladimir Divoky ◽  
...  
Keyword(s):  
Protein Expression ◽  
Iron Uptake ◽  
Chinese Hamster ◽  
Microcytic Anemia ◽  
Homozygous Mutation ◽  
Divalent Metal Transporter 1 ◽  
Divalent Metal Transporter ◽  
Hypochromic Microcytic Anemia ◽  
Functional Consequences ◽  
Uptake Activity

We have previously described a case of severe hypochromic microcytic anemia caused by a homozygous mutation in the divalent metal transporter 1 (DMT1 1285G > C). This mutation encodes for an amino acid substitution (E399D) and causes preferential skipping of exon 12 during processing of the DMT1 mRNA. To examine the functional consequences of this mutation, full-length DMT1 transcript with the patient's point mutation or a DMT1 transcript with exon 12 deleted was expressed in Chinese hamster ovary (CHO) cells. Our results demonstrate that the E399D substitution has no effect on protein expression and function. In contrast, deletion of exon 12 led to a decreased expression of the protein and disruption of its subcellular localization and iron uptake activity. We hypothesize that the residual protein in hematopoietic cells represents the functional E399D DMT1 variant, but because of its quantitative reduction, the iron uptake activity of DMT1 in the patient's erythroid cells is severely suppressed.

The Path of Iron from Plasma Transferrin to Hemoglobin in Developing Red Blood Cells.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. sci-26-sci-26
Author(s):  
Prem Ponka ◽  
An-Shen Zhang ◽  
Alex Sheftel ◽  
Orian S. Shirihai
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Biosynthetic Pathway ◽  
Protoporphyrin Ix ◽  
Heme Iron ◽  
Erythroid Cells ◽  
Divalent Metal Transporter ◽  
Receptor Complexes ◽  
Endosomal Acidification ◽  
In Erythroid Cells

Abstract An exquisite relationship between iron and heme in hemoglobin-synthesizing cells makes blood red. Erythroid cells are the most avid consumers of iron (Fe) in the organism and synthesize heme at a breakneck speed. Additionally, there is virtually no free Fe or heme detectable during hemoglobin (Hb) synthesis. Developing red blood cells (RBC) can take up Fe only from the plasma glycoprotein transferrin (Tf). Delivery of iron to these cells occurs following the binding of Tf to its cognate receptors on the cell membrane. The Tf-receptor complexes are then internalized via endocytosis, and iron is released from Tf by a process involving endosomal acidification. Iron, following its reduction to Fe2+ by Steap3, is then transported across the endosomal membrane by the divalent metal transporter, DMT1. However, the post-endosomal path of Fe in the developing RBC remains elusive or is, at best, controversial. It has been commonly accepted that a low molecular weight intermediate chaperones Fe in transit from endosomes to mitochondria and other sites of utilization; however, this much sought iron-binding intermediate has never been identified. In erythroid cells, more than 90% of iron must enter mitochondria since ferrochelatase, the final enzyme in the heme biosynthetic pathway that inserts Fe2+ into protoporphyrin IX, resides in the inner part of the inner mitochondrial membrane. In fact, in erythroid cells, strong evidence does exist for specific targeting of Fe toward mitochondria. This targeting is demonstrated in Hb-synthesizing cells in which Fe acquired from Tf continues to flow into mitochondria, even when the synthesis of protoporphyrin IX is suppressed. Based on this, we have formulated a hypothesis that in erythroid cells a transient mitochondrion-endosome interaction is involved in iron translocation to its final destination. Recently, we have collected strong experimental evidence supporting this hypothesis: we have shown that Fe, delivered to mitochondria via the Tf pathway, is unavailable to cytoplasmic chelators. Moreover, we have demonstrated that Tf-containing endosomes move and contact mitochondria in erythroid cells, that vesicular movement is required for iron delivery to mitochondria, and that “free” cytoplasmic Fe is not efficiently used for heme biosynthesis. As mentioned above, the substrate for the endosomal transporter DMT1 is Fe2+, the redox form of iron that is also the substrate for ferrochelatase. These facts make the above hypothesis quite attractive, since the “chaperone”-like function of endosomes may be one of the mechanisms that keeps the concentrations of reactive Fe2+ at extremely low levels in oxygen-rich cytosol of erythroblasts, preventing ferrous ion’s participation in a dangerous Fenton reaction. In conclusion, the delivery of iron into Hb occurs extremely efficiently, since mature erythrocytes contain about 45,000-fold more heme iron (20 mM) than non-heme iron (440 nM). These facts, together with experimental data that will be discussed, indicate that the iron transport machinery in erythroid cells is an integral part of the heme biosynthetic pathway.

scholarly journals Iron Acquisition in Mycobacterium avium subsp. paratuberculosis

2015 ◽  
Vol 198 (5) ◽  
pp. 857-866 ◽  
Author(s):  
Joyce Wang ◽  
Jalal Moolji ◽  
Alex Dufort ◽  
Alfredo Staffa ◽  
Pilar Domenech ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Iron Uptake ◽  
Genomic Island ◽  
Iron Acquisition ◽  
Laboratory Data ◽  
Uptake System ◽  
Intracellular Iron ◽  
Content Type ◽  
Iron Uptake System ◽  
Environmental Bacterium

ABSTRACTMycobacterium aviumsubsp.paratuberculosisis a host-adapted pathogen that evolved from the environmental bacteriumM. aviumsubsp.hominissuisthrough gene loss and gene acquisition. Growth ofM. aviumsubsp.paratuberculosisin the laboratory is enhanced by supplementation of the media with the iron-binding siderophore mycobactin J. Here we examined the production of mycobactins by related organisms and searched for an alternative iron uptake system inM. aviumsubsp.paratuberculosis. Through thin-layer chromatography and radiolabeled iron-uptake studies, we showed thatM. aviumsubsp.paratuberculosisis impaired for both mycobactin synthesis and iron acquisition. Consistent with these observations, we identified several mutations, including deletions, inM. aviumsubsp.paratuberculosisgenes coding for mycobactin synthesis. Using a transposon-mediated mutagenesis screen conditional on growth without myobactin, we identified a potential mycobactin-independent iron uptake system on aM. aviumsubsp.paratuberculosis-specific genomic island, LSPP15. We obtained a transposon (Tn) mutant with a disruption in the LSPP15 geneMAP3776cfor targeted study. The mutant manifests increased iron uptake as well as intracellular iron content, with genes downstream of the transposon insertion (MAP3775ctoMAP3772c[MAP3775-2c]) upregulated as the result of a polar effect. As an independent confirmation, we observed the same iron uptake phenotypes by overexpressingMAP3775-2cin wild-typeM. aviumsubsp.paratuberculosis. These data indicate that the horizontally acquired LSPP15 genes contribute to iron acquisition byM. aviumsubsp.paratuberculosis, potentially allowing the subsequent loss of siderophore production by this pathogen.IMPORTANCEMany microbes are able to scavenge iron from their surroundings by producing iron-chelating siderophores. One exception isMycobacterium aviumsubsp.paratuberculosis, a fastidious, slow-growing animal pathogen whose growth needs to be supported by exogenous mycobacterial siderophore (mycobactin) in the laboratory. Data presented here demonstrate that, compared to other closely relatedM. aviumsubspecies, mycobactin production and iron uptake are different inM. aviumsubsp.paratuberculosis, and these phenotypes may be caused by numerous deletions in its mycobactin biosynthesis pathway. Using a genomic approach, supplemented by targeted genetic and biochemical studies, we identified that LSPP15, a horizontally acquired genomic island, may encode an alternative iron uptake system. These findings shed light on the potential physiological consequence of horizontal gene transfer inM. aviumsubsp.paratuberculosisevolution.

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