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Functional expression in Xenopus oocytes reveals that human ferroportin is an iron exporter shared with zinc
Author(s) -
Mitchell Colin J,
Shawki Ali,
Nemeth Elizabeta,
Ganz Tomas,
Mackenzie Bryan
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.1017.3
Subject(s) - ferroportin , efflux , chemistry , nitrilotriacetic acid , hepcidin , dmt1 , transporter , xenopus , ceruloplasmin , zinc , intracellular , transferrin receptor , biochemistry , microbiology and biotechnology , metabolism , iron homeostasis , transferrin , chelation , gene , biology , inflammation , immunology , organic chemistry
Iron homeostasis is achieved by regulating the intestinal absorption of the metal and its recycling by macrophages. Export of iron from enterocytes or macrophages to the blood is thought to be mediated by ferroportin (Fpn) under the control of hepcidin. Some mutations in Fpn result in macrophage iron loading whereas others, as a result of insensitivity to hepcidin, cause systemic iron overload. Whereas Fpn was identified a decade ago, there remains a paucity of information about how it works. We expressed GFP‐tagged human Fpn in RNA‐injected Xenopus oocytes and observed using confocal microscopy exclusive plasma‐membrane localization. As a first step in its characterization, we established an assay to detect functional expression of Fpn by microinjecting oocytes with 55 Fe and measuring the 55 Fe efflux over 30 min. Expression of Fpn increased the first‐order rate constants describing 55 Fe efflux between 4‐fold and 100‐fold over control. Fpn‐mediated 55 Fe efflux was maximal when the injectate contained nitrilotriacetic acid (NTA), and when we provided apotransferrin, bathophenanthroline disulfonate (BPS) and NTA in the incubation medium, whereas inclusion of ceruloplasmin had only a modest effect. Fpn‐mediated 55 Fe was accelerated in pH 8.2 medium compared with pH 7.2 or pH 6.2 ( P < 0.001). Fpn expression also stimulated the efflux of 65 Zn ( P < 0.001) but not of 54 Mn, 64 Cu or 109 Cd ( P > 0.11). We conclude that Fpn mediates cellular iron efflux and that Zn 2+ counts among its narrow substrate profile.