Nramp 2 (DCT1/DMT1) Expressed at the Plasma Membrane Transports Iron and Other Divalent Cations into a Calcein-accessible Cytoplasmic Pool

Picard, Virginie; Govoni, Gregory; Jabado, Nada; Gros, Philippe

doi:10.1074/jbc.m005387200

Cited by 177 publications

(156 citation statements)

References 43 publications

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“…S2C), but not Ca 2+ (Fig. S2D), consistent with previous studies with mammalian Nramp2 homologs (9,11,14,21). However, in contrast to DraNramp M230A, M265A did not transport any of the tested metals, similarly to the D86A loss-offunction phenotype, even though both variants expressed as well as WT (Fig.…”

Section: Significancesupporting

confidence: 79%

“…Nramps are generally thought to function as metal-proton symporters (1) and are able to bind and/or transport a wide range of divalent transition metal substrates, including the biologically useful metals Mn 2+ , Fe 2+ , Co 2+ , Ni 2+ , Cu 2+ , and Zn 2+ , as well as the toxic heavy metals Cd 2+ , Pb 2+ , and Hg 2+ (4,(9)(10)(11)(12)(13). Nramps do discriminate against the divalent alkaline earth metal ions Mg 2+ and Ca 2+ (9,14), which are typically several orders of magnitude more abundant than the transition metals (15).…”

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confidence: 99%

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Conserved methionine dictates substrate preference in Nramp-family divalent metal transporters

Bozzi

Bane

Weihofen

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

110

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Natural resistance-associated macrophage protein (Nramp) family transporters catalyze uptake of essential divalent transition metals like iron and manganese. To discriminate against abundant competitors, the Nramp metal-binding site should favor softer transition metals, which interact either covalently or ionically with coordinating molecules, over hard calcium and magnesium, which interact mainly ionically. The metal-binding site contains an unusual, but conserved, methionine, and its sulfur coordinates transition metal substrates, suggesting a vital role in their transport. Using a bacterial Nramp model system, we show that, surprisingly, this conserved methionine is dispensable for transport of the physiological manganese substrate and similar divalents iron and cobalt, with several small amino acid replacements still enabling robust uptake. Moreover, the methionine sulfur's presence makes the toxic metal cadmium a preferred substrate. However, a methionine-to-alanine substitution enables transport of calcium and magnesium. Thus, the putative evolutionary pressure to maintain the Nramp metal-binding methionine likely exists because it-more effectively than any other amino acid-increases selectivity for low-abundance transition metal transport in the presence of high-abundance divalents like calcium and magnesium.transition metals | MntH | divalent metal transporter DMT1 | hard-soft acid-base theory | ion selectivity filters A ll organisms require transition metal ions as cofactors in proteins that perform a variety of essential cellular tasks. Through evolution, organisms have developed mechanisms to acquire, transport, and safely store essential metals such as manganese, iron, cobalt, and zinc. The natural resistance-associated macrophage protein (Nramp) family of metal transporters represents a common transition metal acquisition strategy conserved across all kingdoms of life (1). The first discovered mammalian Nramp (Nramp1) is expressed in phagosomal membranes and likely extracts essential metals to help kill engulfed pathogens (2, 3). Mammals use Nramp2, an essential gene also called DMT1, to absorb dietary iron into the enterocytes that line the small intestine (4) and to extract iron from transferrin-containing endosomes in all tissues. Bacteria express their own Nramp homologs, which they typically use to scavenge manganese and other first row divalent transition metals (5, 6). Last, most plants have several Nramp homologs that take up iron and manganese, the essential cofactor in photosystem II, from the soil or vacuolar stores (7,8).Nramps are generally thought to function as metal-proton symporters (1) and are able to bind and/or transport a wide range of divalent transition metal substrates, including the biologically useful metals Mn 2+ , Fe 2+ , Co 2+ , Ni 2+ , Cu 2+ , and Zn 2+ , as well as the toxic heavy metals Cd 2+ , Pb 2+ , and Hg 2+ (4, 9-13). Nramps do discriminate against the divalent alkaline earth metal ions Mg 2+ and Ca 2+ (9, 14), which are typically several orders of magnitude mor...

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Section: Significancesupporting

confidence: 79%

mentioning

confidence: 99%

Conserved methionine dictates substrate preference in Nramp-family divalent metal transporters

Bozzi

Bane

Weihofen

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

110

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“…Based on homology with members of the LeuT superfamily, Nramp1 and DMT1 most likely have pseudo-symmetry with domains 1-6 and domains 7-12 representing N-and C-terminal halves, respectively (44). The loop between domains 7 and 8 is extracellular and glycosylated (57,62). In a curiosity of nature, DMT1 allelic variants have been found in mice (41) and rats (58) with the same G185R substitution in the predicted fourth transmembrane-spanning domain.…”

Section: D169mentioning

confidence: 99%

Nramp1 and Other Transporters Involved in Metal Withholding during Infection

Wessling‐Resnick

2015

Journal of Biological Chemistry

112

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During the course of infection, many natural defenses are set up along the boundaries of the host-pathogen interface. Key among these is the host response to withhold metals to restrict the growth of invading microbes. This simple act of nutritional warfare, starving the invader of an essential element, is an effective means of limiting infection. The physiology of metal withholding is often referred to as "nutritional immunity," and the mechanisms of metal transport that contribute to this host response are the focus of this review.The rationale for host metal withholding against invading pathogens seems intuitive; restriction of any nutrient required for pathogen survival should combat infection. But why metals? The answer to this question becomes crystal clear when one recognizes that iron is required for nearly all life forms, including pathogenic bacteria, with a few rare exceptions that rely on manganese instead (1-7). Iron is essential for ATP production and other metabolic pathways, but its concentration is limited to avoid production of ROS 2 catalyzed by excess levels of this redox-active metal (8). In some organisms, manganese can substitute for redox-active iron to protect against oxidative stress (9, 10). Moreover, most pathogens require manganese to produce their own superoxide dismutase activity to thwart oxidative killing mechanisms exerted by the host (11-13). Iron and manganese have similar ionic radii, have a divalent charge state under physiological conditions, have similar coordination chemistries, and have cellular concentrations in the micromolar range. Consequently, it comes as no surprise that these two metals also share membrane transporters, despite their different cellular functions and distributions. Restriction of iron and/or manganese provides a broad-spectrum metabolic "antidote" against all pathogenic infections, and the common transport pathways used by these metals present selective targets to limit their availability. Although many other immune responses contribute to the fight against infection, metal withholding represents a major force in host defense with combat controlled through transport.The concept of nutritional immunity through metal withholding is largely based on the observations that the iron content of the human diet profoundly affects infectious diseases such as malaria, brucellosis, and tuberculosis (14, 15). Iron overload states such as sickle cell anemia and ␤-thalassemia increase the risk of infection (16 -18). Hereditary hemochromatosis, an inherited disorder of iron metabolism, is also linked to susceptibility to some pathogens (17, 19 -22). Iron deficiency, on the other hand, is associated with decreased survival of individuals infected with HIV and other agents (23). Still, high iron is positively associated with viral load and mortality in HIV (24 -26), suggesting that an optimal balance of iron is necessary. In general terms, low iron status is protective, whereas elevated iron levels promote infection (27, 28). The complexity of host-pathogen interact...

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“…Studies in cultured mammalian cells have also shown that both isoforms of DMT1 are capable of transporting a variety of divalent metal ions across the plasma membrane [12,13]. Transport of these metal ions was shown to occur at pH 5.5, but not at 7.4 [1].…”

mentioning

confidence: 99%

Structure and topology of the transmembrane domain 4 of the divalent metal transporter in membrane‐mimetic environments

Li¹,

Li²,

Qian³

et al. 2004

European Journal of Biochemistry

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The divalent metal transporter (DMT1) is a 12-transmembrane domain protein responsible for dietary iron uptake in the duodenum and iron acquisition from transferrin in peripheral tissues. The transmembrane domain 4 (TM4) of DMT1 has been shown to be crucial for its biological function. Here we report the 3D structure and topology of the DMT1-TM4 peptide by NMR spectroscopy with simulated annealing calculations in membrane-mimetic environments, e.g. 2,2,2-trifluoroethanol and SDS micelles. The 3D structures of the peptide are similar in both environments, with nonordered and flexible N-and C-termini flanking an ordered helical region. The final set of the 16 lowest energy structures is particularly well defined in the region of residues Leu9-Phe20 in 2,2,2-trifluoroethanol, with a mean pairwise root mean square deviation of 0.23 ± 0.10 Å for the backbone heavy atoms and 0.82 ± 0.17 Å for all heavy atoms. In SDS micelles, the length of the helix is dependent on pH values. In particular, the C-terminus becomes well-structured at low pH (4.0), whereas the N-terminal segment (Arg1-Gly7) is flexible and poorly defined at all pH values studied. The effects of 12-doxylPtdCho spin-label and paramagnetic metal ions on NMR signal intensities demonstrated that both the N-terminus and helical region of the TM4 are embedded into the interior of SDS micelles. Unexpectedly, we observed that amide protons exchanged much faster in SDS than in 2,2, 2-trifluoroethanol, indicating that there is possible solvent accessibility in the structure. The paramagnetic metal ions broaden NMR signals from residues both situated in aqueous phase and in the helical region. From these results we speculate that DMT1-TM4s may self-assemble to form a channel through which metal ions are likely to be transported. These results might provide an insight into the structure-function relationship for the integral DMT1.Keywords: DMT1; membrane; NMR; structure.The divalent metal transporter (DMT1) gene, also known as Nramp2 (natural resistance-associated macrophage protein-2) and DCT1, was identified recently [1,2]. It belongs to a large family of integral membrane proteins highly conserved throughout evolution, from bacteria to human beings [3][4][5][6]. It is the only known cellular iron importer, and is responsible for importing iron from the gut into the enterocytes and also for transporting iron across the endosomal membrane in the transferrin cycle [7][8][9]. The DMT1 consists of 561 amino acids with 12 putative transmembrane domains [1]. The DMT1 gene encodes two messenger RNAs produced by alternative splicing of two 3¢ exons that show different 3¢ untranslated regions containing an iron response element (isoform I) and no iron response element (isoform II), as well as distinct C-terminal protein sequences [7][8][9][10]. Recently, DMT1 mRNA expression has also been detected in the kidney [11].Direct metal transport studies in Xenopus laevis oocytes have demonstrated that DMT1 (isoform I) is a pHdependent divalent metal transporter with broad substrat...

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Nramp 2 (DCT1/DMT1) Expressed at the Plasma Membrane Transports Iron and Other Divalent Cations into a Calcein-accessible Cytoplasmic Pool

Cited by 177 publications

References 43 publications

Conserved methionine dictates substrate preference in Nramp-family divalent metal transporters

Conserved methionine dictates substrate preference in Nramp-family divalent metal transporters

Nramp1 and Other Transporters Involved in Metal Withholding during Infection

Structure and topology of the transmembrane domain 4 of the divalent metal transporter in membrane‐mimetic environments

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