Transferrin iron uptake is stimulated by ascorbate via an intracellular reductive mechanism

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• 作者：Darius J.R. Lane ; ¹ ; ^{darius.lane@sydney.edu.au} ; Sherin Chikhani ; Vera Richardson ; Des R. Richardson ; ¹ ; ^{d.richardson@med.usyd.edu.au}
• 关键词：BCA ; bicinchoninic acid ; BPS ; bathophenanthroline disulfonate ; BSA ; bovine serum albumin ; CHO ; Chinese hamster ovary ; CHX ; cycloheximide ; Dcytb ; duodenal cytochrome b ; DHA ; dehydroascorbate ; DFO ; desferrioxamine ; DMT1 ; divalent metal transporter ; isoform
• 刊名：Biochimica et Biophysica Acta (BBA)/Molecular Cell Research
• 出版年：2013
• 出版时间：June, 2013
• 年：2013
• 卷：1833
• 期：6
• 页码：1527-1541
• 全文大小：2177 K

文摘

Although ascorbate has long been known to stimulate dietary iron (Fe) absorption and non-transferrin Fe uptake, the role of ascorbate in transferrin Fe uptake is unknown. Transferrin is a serum Fe transport protein supplying almost all cellular Fe under physiological conditions. We sought to examine ascorbate's role in this process, particularly as cultured cells are typically ascorbate-deficient. At typical plasma concentrations, ascorbate significantly increased ⁵⁹Fe uptake from transferrin by 1.5-2-fold in a range of cells. Moreover, ascorbate enhanced ferritin expression and increased ⁵⁹Fe accumulation in ferritin. The lack of effect of cycloheximide or the cytosolic aconitase inhibitor, oxalomalate, on ascorbate-mediated ⁵⁹Fe uptake from transferrin indicate increased ferritin synthesis or cytosolic aconitase activity was not responsible for ascorbate's activity. Experiments with membrane-permeant and -impermeant ascorbate-oxidizing reagents indicate that while extracellular ascorbate is required for stimulation of ⁵⁹Fe uptake from ⁵⁹Fe-citrate, only intracellular ascorbate is needed for transferrin ⁵⁹Fe uptake. Additionally, experiments with l-ascorbate analogs indicate ascorbate's reducing ene-diol moiety is necessary for its stimulatory activity. Importantly, neither N-acetylcysteine nor buthionine sulfoximine, which increase or decrease intracellular glutathione, respectively, affected transferrin-dependent ⁵⁹Fe uptake. Thus, ascorbate's stimulatory effect is not due to a general increase in cellular reducing capacity. Ascorbate also did not affect expression of transferrin receptor 1 or ¹²⁵I-transferrin cellular flux. However, transferrin receptors, endocytosis, vacuolar-type ATPase activity and endosomal acidification were required for ascorbate's stimulatory activity. Therefore, ascorbate is a novel modulator of the classical transferrin Fe uptake pathway, acting via an intracellular reductive mechanism.