Solute carrier 11a1 (Slc11a1; formerly Nramp1) regulates metabolism and release of iron acquired by phagocytic, but not transferrin-receptor-mediated, iron uptake

2002 ◽  
Vol 363 (1) ◽  
pp. 89-94 ◽  
Author(s):  
Victoriano MULERO ◽  
Susan SEARLE ◽  
Jenefer M. BLACKWELL ◽  
Jeremy H. BROCK
Keyword(s):  
Iron Uptake ◽  
Interferon Γ ◽  
Natural Resistance ◽  
Null Alleles ◽  
Iron Release ◽  
Wild Type ◽  
Bivalent Cation ◽  
Insoluble Form ◽  
Solute Carrier ◽  
Macrophage Protein

Solute carrier 11a1 (Slc11a1; formerly Nramp1; where Nramp stands for natural-resistance-associated macrophage protein) is a proton/bivalent cation antiporter that localizes to late endosomes/lysosomes and controls resistance to pathogens. In the present study the role of Slc11a1 in iron turnover is examined in macrophages transfected with Slc11a1Gly169 (wild-type) or Slc11a1Asp169 (mutant = functional null) alleles. Following direct acquisition of transferrin (Tf)-bound iron via the Tf receptor, iron uptake and release was equivalent in wild-type and mutant macrophages and was not influenced by interferon-γ/lipopolysaccharide activation. Following phagocytosis of [59Fe]Tf—anti-Tf immune complexes, iron uptake was equivalent and up-regulated similarly with activation, but intracellular distribution was markedly different. In wild-type macrophages most iron was in the soluble (60%) rather than insoluble (12%) fraction, with 28% ferritin (Ft)-bound. With activation, the soluble component increased to 82% at the expense of Ft-bound iron (< 5%). In mutant macrophages, 40–50% of iron was in insoluble form, 50–60% was soluble and < 5% was Ft-bound. Western-blot analysis confirmed failure of mutant macrophages to degrade complexes 24h after phagocytic uptake. Confocal microscopy showed that complexes were within lysosome-associated membrane protein 1-positive vesicles in wild-type and mutant macrophages at 30min and 24h, implying failure in the degradative process in mature phagosomes in mutant macrophages. NO-mediated iron release was 2.4-fold higher in activated wild-type macrophages compared with mutant macrophages. Overall, our data suggest that iron acquired by phagocytosis and degradation is retained within the phagosomal compartment in wild-type macrophages, and that NO triggers iron release by direct secretion of phagosomal contents rather than via the cytoplasm.

Genetic analysis of SLC11A1 polymorphisms in multiple sclerosis patients

2004 ◽  
Vol 10 (6) ◽  
pp. 618-620 ◽  
Author(s):  
Manuel Comabella ◽  
Laura Altet ◽  
Francesc Peris ◽  
Pablo Villoslada ◽  
Armand Sánchez ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Natural Resistance ◽  
Bivalent Cation ◽  
Slc11a1 Gene ◽  
Late Endosomes ◽  
Solute Carrier ◽  
Macrophage Protein ◽  
Multiple Sclerosis Patients ◽  
Control Study

Solute carrier 11a1 (SLC11A1; formerly NRAMP1, where NRAMP stands for natural resistance-associated macrophage protein) is a proton/bivalent cation antiporter that localizes to late endosomes/lysosomes. SLC11A1 regulates macrophage functions that are of potential importance in the induction and/or maintenance of autoimmune diseases such as rheumatoid arthritis, type 1 diabetes and Crohn’s disease. We investigated SLC11A1 gene as a candidate gene for genetic susceptibility to multiple sclerosis (MS) in our population. Four SLC11A1 gene polymorphisms (5?GT repeat, D543N, 1729 -55del4 and 1729 -271del4) were analysed in a case-control study of 195 patients with MS and 125 control subjects. We found no evidence of association between SLC11A1 polymorphisms and MS susceptibility in the Spanish population.

Transcriptional control of Nramp1: a paradigm for the repressive action of c-Myc

2004 ◽  
Vol 32 (6) ◽  
pp. 1084-1086 ◽  
Author(s):  
A.S. Lapham ◽  
E.S. Phillips ◽  
C.H. Barton
Keyword(s):  
Transcriptional Control ◽  
Cell Activation ◽  
Zinc Finger Protein ◽  
Specific Binding ◽  
Divalent Cations ◽  
Natural Resistance ◽  
Finger Protein ◽  
Late Endosomes ◽  
Solute Carrier ◽  
Macrophage Protein

Slc11a1/Nramp1 (solute carrier family 11 member a1/murine natural resistance-associated macrophage protein 1 gene) encodes a divalent cation transporter that resides within lysosomes/late endosomes of macrophages. Nramp1 modulates the cellular distribution of divalent cations in response to cell activation by intracellular pathogens. Nramp1 expression is repressed and activated by the proto-oncogene c-Myc and Miz-1 (c-Myc-interacting zinc finger protein 1) respectively. Here we demonstrate, using a c-Myc mutant (V394D, Val394→Asp) that is incapable of binding Miz-1, that c-Myc repression of Nramp1 transcription is dependent on its interaction with Miz-1. An oligo pull-down assay demonstrates specific binding of recombinant Miz-1 to the Nramp1 Miz-1-binding site or initiator element(s), and Miz-1-dependent c-Myc recruitment.

scholarly journals Host Resistance to Intracellular Infection: Mutation of Natural Resistance-associated Macrophage Protein 1 (Nramp1) Impairs Phagosomal Acidification

1998 ◽  
Vol 188 (2) ◽  
pp. 351-364 ◽  
Author(s):  
David J. Hackam ◽  
Ori D. Rotstein ◽  
Wei-jian Zhang ◽  
Samantha Gruenheid ◽  
Philippe Gros ◽  
...  
Keyword(s):  
Integral Membrane Protein ◽  
Natural Resistance ◽  
Wild Type ◽  
Intracellular Parasites ◽  
Late Endosomes ◽  
Latex Beads ◽  
Buffering Power ◽  
Ratio Imaging ◽  
Macrophage Protein ◽  
Phagosomal Membrane

The mechanisms underlying the survival of intracellular parasites such as mycobacteria in host macrophages remain poorly understood. In mice, mutations at the Nramp1 gene (for natural resistance-associated macrophage protein), cause susceptibility to mycobacterial infections. Nramp1 encodes an integral membrane protein that is recruited to the phagosome membrane in infected macrophages. In this study, we used microfluorescence ratio imaging of macrophages from wild-type and Nramp1 mutant mice to analyze the effect of loss of Nramp1 function on the properties of phagosomes containing inert particles or live mycobacteria. The pH of phagosomes containing live Mycobacterium bovis was significantly more acidic in Nramp1- expressing macrophages than in mutant cells (pH 5.5 ± 0.06 versus pH 6.6 ± 0.05, respectively; P &lt;0.005). The enhanced acidification could not be accounted for by differences in proton consumption during dismutation of superoxide, phagosomal buffering power, counterion conductance, or in the rate of proton “leak”, as these were found to be comparable in wild-type and Nramp1-deficient macrophages. Rather, after ingestion of live mycobacteria, Nramp1-expressing cells exhibited increased concanamycin-sensitive H+ pumping across the phagosomal membrane. This was associated with an enhanced ability of phagosomes to fuse with vacuolar-type ATPase–containing late endosomes and/or lysosomes. This effect was restricted to live M. bovis and was not seen in phagosomes containing dead M. bovis or latex beads. These data support the notion that Nramp1 affects intracellular mycobacterial replication by modulating phagosomal pH, suggesting that Nramp1 plays a central role in this process.

Natural-resistance-associated macrophage protein 1 is an H+/bivalent cation antiporter

2001 ◽  
Vol 354 (3) ◽  
pp. 511-519 ◽  
Author(s):  
Tapasree GOSWAMI ◽  
Arin BHATTACHARJEE ◽  
Paul BABAL ◽  
Susan SEARLE ◽  
Elizabeth MOORE ◽  
...  
Keyword(s):  
Metal Ions ◽  
Metal Ion ◽  
Proton Gradient ◽  
Natural Resistance ◽  
Bivalent Cation ◽  
Bivalent Cations ◽  
Carbonyl Cyanide ◽  
Endosomal Membranes ◽  
Data Points ◽  
Macrophage Protein

In mammals, natural-resistance-associated macrophage protein 1 (Nramp1) regulates macrophage activation and is associated with infectious and autoimmune diseases. Nramp2 is associated with anaemia. Both belong to a highly conserved eukaryote/prokaryote protein family. We used Xenopus oocytes to demonstrate that, like Nramp2, Nramp1 is a bivalent cation (Fe2+, Zn2+ and Mn2+) transporter. Strikingly, however, where Nramp2 is a symporter of H+ and metal ions, Nramp1 is a highly pH-dependent antiporter that fluxes metal ions in either direction against a proton gradient. At pH9.0, oocytes injected with cRNA from wild-type murine Nramp1 with a glycine residue at position 169 (Nramp1G169; P = 3.22×10-6) and human NRAMP1 (P = 3.87×10-5) showed significantly enhanced uptake of radiolabelled Zn2+ compared with water-injected controls. At pH5.5, Nramp1G169 (P = 1.34×10-13) and NRAMP1 (P = 1.09×10-6) oocytes showed significant efflux of Zn2+. Zn2+ transport was abolished when the proton gradient was dissipated using carbonyl cyanide p-trifluoromethoxyphenylhydrazone. Using pre-acidified oocytes, currents of 130±57 nA were evoked by 100µM Zn2+ at pH7.5, and 139±47 nA by 100µM Fe2+ at pH7.0, in Nramp1G169 oocytes; currents of 254±49 nA and 242±26 nA were evoked, respectively, in NRAMP1 oocytes. Steady-state currents evoked by increasing concentrations of Zn2+ were saturable, with apparent affinity constants of approx. 614nM for Nramp1G169 and approx. 562nM for NRAMP1 oocytes, and a curvilinear voltage dependence of transporter activity (i.e. the data points approximate to a curve that approaches a linear asymptote). In the present study we propose a new model for metal ion homoeostasis in macrophages. Under normal physiological conditions, Nramp2, localized to early endosomal membranes, delivers extracellularly acquired bivalent cations into the cytosol. Nramp1, localized to late endosomal/lysosomal membranes, delivers bivalent cations from the cytosol into this acidic compartment where they may directly affect antimicrobial activity.

scholarly journals Localisation of Nramp1 in macrophages: modulation with activation and infection

1998 ◽  
Vol 111 (19) ◽  
pp. 2855-2866 ◽  
Author(s):  
S. Searle ◽  
N.A. Bright ◽  
T.I. Roach ◽  
P.G. Atkinson ◽  
C.H. Barton ◽  
...  
Keyword(s):  
Macrophage Activation ◽  
Leishmania Major ◽  
Polyclonal Antibodies ◽  
Natural Resistance ◽  
Activated Macrophages ◽  
Wild Type ◽  
Late Endosomes ◽  
Gold Labelling ◽  
Macrophage Protein ◽  
Phagosome Fusion

The murine natural resistance-associated macrophage protein, Nramp1, has multiple pleiotropic effects on macrophage activation and regulates survival of intracellular pathogens including Leishmania, Salmonella and Mycobacterium species. Nramp1 acts as an iron transporter, but precisely how this relates to macrophage activation and/or pathogen survival remains unclear. To gain insight into function, anti-Nramp1 monoclonal and polyclonal antibodies are used here to localise Nramp1 following activation and infection. Confocal microscope analysis in uninfected macrophages demonstrates that both the mutant (infection-susceptible) and wild-type (infection-resistant) forms of the protein localise to the membranes of intracellular vesicular compartments. Gold labelling and electron microscopy defines these compartments more precisely as electron-lucent late endosomal and electron-dense lysosomal compartments, with Nramp1 colocalizing with Lamp1 and cathepsins D and L in both compartments, with macrosialin in late endosomes, and with BSA-5 nm gold in pre-loaded lysosomes. Nramp1 is upregulated with interferon-(gamma) and lipopolysaccaride treatment, coinciding with an increase in labelling in lysosomes relative to late endosomes and apparent dispersion of Nramp1-positive vesicles from a perinuclear location towards the periphery of the cytoplasm along the microtubular network. In both control and activated macrophages, expression of the protein is 3- to 4-fold higher in wild-type compared to mutant macrophages. In Leishmania major-infected macrophages, Nramp1 is observed in the membrane of the pathogen-containing phagosomes, which retain a perinuclear localization in resting macrophages. In Mycobacterium avium-infected resting and activated macrophages, Nramp1-positive vesicles migrated to converge, but not always fuse, with pathogen-containing phagosomes. The Nramp1 protein is thus located where it can have a direct influence on phagosome fusion and the microenvironment of the pathogen, as well as in the more general regulation of endosomal/lysosomal function in macrophages.

scholarly journals Natural-resistance-associated macrophage protein 1 is an H+/bivalent cation antiporter

2001 ◽  
Vol 354 (3) ◽  
pp. 511 ◽  
Author(s):  
Tapasree GOSWAMI ◽  
Arin BHATTACHARJEE ◽  
Paul BABAL ◽  
Susan SEARLE ◽  
Elizabeth MOORE ◽  
...  
Keyword(s):  
Natural Resistance ◽  
Bivalent Cation ◽  
Macrophage Protein

Heterologous expression, functional characterization and localization of two isoforms of the monkey iron transporter Nramp2

2000 ◽  
Vol 349 (1) ◽  
pp. 289-297 ◽  
Author(s):  
Li ZHANG ◽  
Timothy LEE ◽  
Yue WANG ◽  
Tuck W. SOONG
Keyword(s):  
Amino Acid ◽  
Subcellular Localization ◽  
Tissue Distribution ◽  
Iron Uptake ◽  
Functional Characterization ◽  
Amino Acid Level ◽  
Natural Resistance ◽  
C Terminus ◽  
Macrophage Protein ◽  
Responsive Element

Natural resistance-associated macrophage protein 2 (Nramp2) has been suggested to be involved in transferrin-independent iron uptake. Two isoforms of the Nramp2 gene generated by alternative splicing of the 3ʹ exons were identified in mouse, rat and human, but it is unclear if they perform distinct functions. To rationalize our previous work, which indicated an increase in iron deposition in a Parkinsonian monkey brain, two monkey Nramp2 isoforms were isolated for a comparative study to assess their relative iron-uptake abilities, tissue distribution and subcellular localization. The monkey Nramp2 isoforms, 2a and 2b, exhibit approx. 98% identity at the amino acid level when compared with the human homologues. The Nramp2a transcript contains a canonical iron-responsive element (IRE), whereas that of Nramp2b lacks the IRE motif in the 3ʹ untranslated region. By reverse transcriptase (RT)-PCR, the mRNAs of both isoforms were detected in all tissues examined. The amino acid differences at the C-terminus neither affected the protein expression levels in HEK-293T and COS-7 cells nor altered the subcellular localization and tissue distribution of the isoforms. Similar levels of iron uptake were detected in the HEK-293T cells transfected with either the Nramp2a or 2b gene, and a reduction of iron from the ferric (Fe3+) to the ferrous (Fe2+) state is necessary before transport can take place. However, this transferrin-independent uptake of iron into the cells is not a Ca2+-dependent process.

The Natural Resistance-Associated Macrophage Protein from the Protozoan ParasitePerkinsus marinusMediates Iron Uptake

Biochemistry ◽  
2011 ◽  
Vol 50 (29) ◽  
pp. 6340-6355 ◽  
Author(s):  
Zhuoer Lin ◽  
José-Antonio Fernández-Robledo ◽  
Mathieu F. M. Cellier ◽  
Gerardo R. Vasta
Keyword(s):  
Iron Uptake ◽  
Natural Resistance ◽  
Macrophage Protein

scholarly journals Nramp1 locus encodes a 65 kDa interferon-γ-inducible protein in murine macrophages

1997 ◽  
Vol 325 (3) ◽  
pp. 779-786 ◽  
Author(s):  
Peter G. P. ATKINSON ◽  
Jenefer M. BLACKWELL ◽  
C. Howard BARTON
Keyword(s):  
Divalent Cation ◽  
Sequence Similarity ◽  
Iron Chelation ◽  
Interferon Γ ◽  
Natural Resistance ◽  
Protein Levels ◽  
Macrophage Cells ◽  
Protein Locus ◽  
Macrophage Protein ◽  
Inducible Protein

The murine Nramp1 (natural-resistance-associated macrophage protein) locus, formerly known as Ity/Lsh/Bcg, was isolated previously on the basis of chromosomal location, and as conferring natural resistance to infection against intracellular macrophage pathogens. The gene encodes a transporter molecule of unknown function. We have prepared polyclonal antisera against the C-terminal 35 amino acids of murine Nramp1. This serum is reactive towards a 65 kDa protein, expressed in murine macrophage cells from resistant or susceptible mice stimulated with interferon-γ and lipopolysaccharide, but not in non-macrophage cells. Evidence indicates that Nramp1 is localized in a subcellular membrane rather than at the cell surface. This evidence includes: the identification of conserved endocytic targeting motifs following inspection of human and murine Nramp sequences; the enrichment of Nramp1, following magnetic selection of phagolysosomal vesicles from activated macrophages that were allowed to phagocytose magnetic, IgG-coated beads; confocal microscopy. These studies place Nramp1 on a membrane in close proximity to obligate intracellular pathogens. A link between Nramp1 and divalent-cation transport is suggested by sequence similarity with yeast SMF1. Evidence showing modulation of Nramp1 protein levels by iron chelation provides a direct link with Nramp1 function and divalent-cation metabolism.

scholarly journals Role of Nramp1 Deletion inChlamydia Infection in Mice

2000 ◽  
Vol 68 (8) ◽  
pp. 4831-4833 ◽  
Author(s):  
Sukumar Pal ◽  
Ellena M. Peterson ◽  
Luis M. de la Maza
Keyword(s):  
Chlamydia Trachomatis ◽  
Chlamydial Infection ◽  
Chlamydia Pneumoniae ◽  
Natural Resistance ◽  
Wild Type ◽  
Intranasal Infection ◽  
Macrophage Protein

ABSTRACT Elicited macrophages from 129sv mice with a functional deletion of the natural-resistance-associated macrophage protein 1 gene (Nramp1) were shown to be as susceptible as wild-type mice to infection with the Chlamydia trachomatis mouse pneumonitis and L3 serovars and to Chlamydia pneumoniae. Furthermore, the two groups of mice were shown to be similarly susceptible to an intranasal infection with these microorganisms. In conclusion, the Nramp1 gene does not appear to play a major role in the regulation of the susceptibility of mice to a chlamydial infection.

Sign in / Sign up

Export Citation Format

Share Document