Use of inhibitors of ion transport to differentiate iron transporters in erythroid cells

Donna Savigni, Evan Morgan

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Iron uptake by rabbit reticulocytes and mature erythrocytes was investigated using 4 incubation systems: 1. Fe-transferrin in NaCl at pH 7.4, 2. Fe-transferrin in sucrose at pH 5.9, 3. Fe(II)-sucrose in sucrose at pH 6.5, and 4. Fe(II)-sucrose in KCl at pH 7.0. These systems were compared with respect to their magnitude and response to many membrane transport inhibitors and modifying agents. Iron uptake eia the first 3 systems had many similar features that were quite distinct from those of iron uptake in the fourth system. On the basis of these results, it is concluded that erythroid cells contain two iron transport mechanisms, one with a high affinity and relatively low capacity for iron transport, which can be studied using incubation systems 1-3, and the other of low affinity but high capacity (incubation system 4). High-affinity transport is present only in immature erythroid cells, is relatively sensitive to inhibition by N-ethylmaleimide (NEM), N,N-1-dicyclohexylcarbodiimide (DCCD), and 7-chloro-4-nitrobenz-2-oxa-1,3 diazole (NBD), and is probably the mechanism by which iron, released from transferrin within endosomes, is transported across the endosomal membrane into the cytosol. DCCD and NBD are also inhibitors of the endosomal H+-ATPase, which is in keeping with the hypothesis that this ATPase functions as the iron transporter in endosomal membranes. However, the more specific inhibitor of this enzyme, bafilomycin A(1), inhibited iron uptake only in incubation system 1, where its action can be attributed to inhibition of endosomal acidification. Hence, it is unlikely that the ATPase also functions as the iron transporter. The low-affinity uptake mechanism is sensitive to inhibition by amiloride, valinomycin, quinidine, imipramine, quercetin, and diethylstilbestrol (to all of which high-affinity transport is relatively resistant), and is present in mature erythrocytes as well as reticulocytes.
Original languageEnglish
Pages (from-to)371-378
JournalBiochemical Pharmacology
Volume52
DOIs
Publication statusPublished - 1996

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