ABSTRACT
A number of regulatory factors including dietary iron levels can dramatically alter the expression of the intestinal iron transporter DMT1. Here we show that Caco-2 cells exposed to iron for 4h exhibited a significant decrease in plasma membrane DMT1 protein, though total cellular DMT1 levels were unaltered. Following biotinylation of cell surface proteins, there was a significant increase in intracellular biotin-labelled DMT1 in iron-exposed cells. Furthermore, iron-treatment increased levels of DMT1 co-localised with LAMP1, suggesting that the initial response of intestinal epithelial cells to iron involves internalisation and targeting of DMT1 transporter protein towards a late endosomal/lysosomal compartment.
Subject(s)
Cation Transport Proteins/metabolism , Intestinal Mucosa/metabolism , Iron-Binding Proteins/metabolism , Iron/pharmacology , Biotinylation , Caco-2 Cells , Cation Transport Proteins/analysis , Cation Transport Proteins/genetics , Cell Membrane/chemistry , Cell Membrane/metabolism , Endosomes/chemistry , Endosomes/metabolism , Humans , Intestinal Mucosa/drug effects , Intestinal Mucosa/physiology , Iron/metabolism , Iron-Binding Proteins/analysis , Iron-Binding Proteins/genetics , Lysosomal Membrane Proteins , Lysosomes/chemistry , Lysosomes/metabolism , Membrane Glycoproteins/analysis , Membrane Glycoproteins/metabolismABSTRACT
TNFalpha has dramatic effects on iron metabolism contributing to the generation of hypoferraemia in the anaemia of chronic disease. Interestingly, TNFalpha is also synthesised and released within the intestinal mucosa, suggesting that this pro-inflammatory cytokine may play a role in regulating dietary iron absorption. To investigate this possibility, we stimulated intestinal Caco-2 cells with TNFalpha (10 ng/ml). In TNFalpha-treated cells, apical iron uptake was significantly decreased and this was accompanied by a reduction in divalent metal transporter protein and mRNA expression. Our data suggest that TNFalpha could regulate dietary iron absorption and that the apical transport machinery is the target for these actions.
Subject(s)
Biological Transport/drug effects , Epithelial Cells/drug effects , Gene Expression Regulation/drug effects , Intestines/cytology , Iron/metabolism , Membrane Transport Proteins/genetics , Receptors, Transferrin/metabolism , Tumor Necrosis Factor-alpha/pharmacology , Caco-2 Cells , Cation Transport Proteins/genetics , Epithelial Cells/metabolism , Ferritins/metabolism , Humans , Iron-Binding Proteins/genetics , Promoter Regions, Genetic/genetics , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, Transferrin/geneticsABSTRACT
Copper is an essential dietary trace metal, however the mechanisms involved in intestinal copper uptake are unclear. Two putative copper transporters are expressed in Caco-2 cells, the divalent metal transporter (DMT1) and copper transporter (Ctr1). Our data demonstrate that copper could compete with iron for uptake via DMT1 and that DMT1 protein and mRNA expression were decreased following exposure (24 h) to high copper. Expression of Ctr1, which acts as a copper transporter in transfected cell lines, was unaffected by copper treatment. Interestingly, exposure to copper increased iron efflux from Caco-2 cells and up regulated IREG1 (iron-regulated mRNA) expression.
Subject(s)
Cation Transport Proteins/metabolism , Copper/pharmacology , Iron-Binding Proteins/metabolism , Membrane Proteins/metabolism , Caco-2 Cells , Cation Transport Proteins/drug effects , Cation Transport Proteins/genetics , Copper Transporter 1 , Culture Media , Epithelial Cells/drug effects , Epithelial Cells/metabolism , Humans , Iron/metabolism , Iron-Binding Proteins/drug effects , Iron-Binding Proteins/genetics , Membrane Proteins/drug effects , Membrane Proteins/genetics , RNA, Messenger/metabolism , Response Elements/drug effectsABSTRACT
A divalent metal transporter, DMT1, located on the apical membrane of intestinal enterocytes is the major pathway for the absorption of dietary non-haem iron. Using human intestinal Caco-2 TC7 cells, we have shown that iron uptake and DMT1 protein in the plasma membrane were significantly decreased by exposure to high iron for 24 h, in a concentration-dependent manner, whereas whole cell DMT1 protein abundance was unaltered. This suggests that part of the response to high iron involved redistribution of DMT1 between the cytosol and cell membrane. These events preceded changes in DMT1 mRNA, which was only decreased following 72 h exposure to high iron.
