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Divalent metal-ion transporter DMT1 mediates both H+ -coupled Fe2+ transport and uncoupled fluxes

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Abstract

The H+ -coupled divalent metal-ion transporter DMT1 serves as both the primary entry point for iron into the body (intestinal brush-border uptake) and the route by which transferrin-associated iron is mobilized from endosomes to cytosol in erythroid precursors and other cells. Elucidating the molecular mechanisms of DMT1 will therefore increase our understanding of iron metabolism and the etiology of iron overload disorders. We expressed wild type and mutant DMT1 in Xenopus oocytes and monitored metal-ion uptake, currents and intracellular pH. DMT1 was activated in the presence of an inwardly directed H+ electrochemical gradient. At low extracellular pH (pHo), H+ binding preceded binding of Fe2+ and its simultaneous translocation. However, DMT1 did not behave like a typical ion-coupled transporter at higher pHo, and at pHo 7.4 we observed Fe2+ transport that was not associated with H+ influx. His272 → Ala substitution uncoupled the Fe2+ and H+ fluxes. At low pHo, H272A mediated H+ uniport that was inhibited by Fe2+. Meanwhile H272A-mediated Fe2+ transport was independent of pHo. Our data indicate (i) that H+ coupling in DMT1 serves to increase affinity for Fe2+ and provide a thermodynamic driving force for Fe2+ transport and (ii) that His-272 is critical in transducing the effects of H+ coupling. Notably, our data also indicate that DMT1 can mediate facilitative Fe2+ transport in the absence of a H+ gradient. Since plasma membrane expression of DMT1 is upregulated in liver of hemochromatosis patients, this H+ -uncoupled facilitative Fe2+ transport via DMT1 can account for the uptake of nontransferrin-bound plasma iron characteristic of iron overload disorders.

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Acknowledgments

We are grateful to Hitomi Takanaga and Jonathan Sabbagh for their help in the laboratory, and to William A. Stein for advice regarding mathematical fitting. This study was supported by NIH grants R01-DK057782 (to M.A.H.) and R01-DK056218 (to M.F.R.), and a pilot/feasibility award (to B.M.) from the Harvard Digestive Diseases Center, funded by NIH center grant P30-DK034854.

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Correspondence to Matthias A. Hediger.

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Mackenzie, B., Ujwal, M.L., Chang, MH. et al. Divalent metal-ion transporter DMT1 mediates both H+ -coupled Fe2+ transport and uncoupled fluxes. Pflugers Arch - Eur J Physiol 451, 544–558 (2006). https://doi.org/10.1007/s00424-005-1494-3

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