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The first step in intestinal iron absorption is mediated by the H + -coupled Fe 2+ transporter called divalent cation transporter 1/divalent metal ion transporter 1 (DCT1/DMT1) (also known as natural resistance-associated macrophage protein 2). DCT1/DMT1 mRNA levels in the duodenum strongly increase in response to iron depletion. To study the mechanism of iron-dependent DCT1/DMT1 mRNA regulation, we investigated the endogenous expression of DCT1/DMT1 mRNA in various cell types. We found that only the iron responsive element (IRE)-containing form, which corresponds to one of two splice forms of DCT1/DMT1, is responsive to iron treatment and this responsiveness was cell type specific. We also examined the interaction of the putative 3P P-UTR IRE with iron responsive binding proteins (IRP1 and IRP2), and found that IRP1 binds to the DCT1/ DMT1-IRE with higher affinity compared to IRP2. This differential binding of IRP1 and IRP2 was also reported for the IREs of transferrin receptors, erythroid 5-aminolevulinate synthase and mitochondrial aconitase. We propose that regulation of DCT1/DMT1 mRNA by iron involves post-transcriptional regulation through the binding of IRP1 to the transporter's IRE, as well as other as yet unknown factors. ß 2001 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.

We performed a genome-wide association study (GWAS) of Kawasaki disease in Japanese subjects using data from 428 individuals with Kawasaki disease (cases) and 3,379 controls genotyped at 473,803 SNPs. We validated the association results in two independent replication panels totaling 754 cases and 947 controls. We observed significant associations in the FAM167A-BLK region at 8p22-23 (rs2254546, P = 8.2 × 10(-21)), in the human leukocyte antigen (HLA) region at 6p21.3 (rs2857151, P = 4.6 × 10(-11)) and in the CD40 region at 20q13 (rs4813003, P = 4.8 × 10(-8)). We also replicated the association of a functional SNP of FCGR2A (rs1801274, P = 1.6 × 10(-6)) identified in a recently reported GWAS of Kawasaki disease. Our findings provide new insights into the pathogenesis and pathophysiology of Kawasaki disease.

NRAMP2 (natural resistance-associated macrophage protein 2)/DMT1 (divalent metal transporter 1) is a divalent metal transporter conserved from prokaryotes to higher eukaryotes that exhibits an unusually broad substrate range, including Fe 2؉ , Zn 2؉ , Mn 2؉ , Cu 2؉, Cd 2؉ , Co 2؉ , Ni 2؉ , and Pb 2؉ , and mediates active proton-coupled transport. Recently, it has been shown that the microcytic anemia (mk) mouse and the Belgrade (b) rat, which have inherited defects in iron transport that result in iron deficiency anemia, have the same missense mutation (G185R) in Nramp2. These findings strongly suggested that NRAMP2 is the apical membrane iron transporter in intestinal epithelial cells and the endosomal iron transporter in transferrin cycle endosomes of other cells. To investigate the cellular functions of NRAMP2, we generated a polyclonal antibody against the N-terminal cytoplasmic domain of human NRAMP2. The affinity-purified anti-NRAMP2 N-terminal antibody recognized a 90 -116-kDa membrane-associated protein, and this band was shifted to 50 kDa by deglycosylation with peptide N-glycosidase F. Subcellular fractionation revealed that NRAMP2 co-sedimented with the late endosomal and lysosomal membrane proteins and LAMP-1 (lysosome-associated membrane protein 1), but not with the transferrin receptor in early endosomes. The intracellular localization of endogenous NRAMP2 and recombinant green fluorescent protein (GFP)-NRAMP2 was examined by immunofluorescence staining and by native fluorescence of GFP, respectively. Both endogenous and GFP-NRAMP2 were detected in vesicular structures and were colocalized with LAMP-2, but not with EEA1 (early endosome antigen 1) or the transferrin receptor. These results indicated that NRAMP2 is localized to the late endosomes and lysosomes, where NRAMP2 may function to transfer the endosomal free Fe 2؉ into the cytoplasm in the transferrin cycle.Iron is indispensable for life, serving as a metal cofactor for many enzymes, including both non-heme and heme proteins.The normal human adult has 35-45 mg of iron/kg of body weight. More than two-thirds of the body's iron content is incorporated into hemoglobin in developing erythroid precursors and mature red cells (1). Nutritional iron absorption (both heme and non-heme iron) occurs primarily in the intestine. Heme iron constitutes only a small fraction of the available dietary iron, but it is highly available for absorption. On the other hand, the absorption of non-heme iron is low and markedly regulated in the first part of the duodenum, in which the acidic pH promotes solubilization of iron transformed to Fe 2ϩ by ferrireductase and ascorbate. In non-intestinal cells, iron is taken into the cell by receptor-mediated endocytosis by transferrin (Tf).1 Specific receptors (Tf receptor (TfR)) on the outer face of the plasma membrane bind diferric Tf with high affinity (2). Once internalized into the cells, the Tf⅐TfR complex is delivered to endosomes, which are acidified to pH 5.5-6.0 through the action of an ATP-dependent proton pump. E...

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