Transferrin receptor-independent uptake of differic transferrin by human hepatoma cells with antisense inhibition of receptor expression.
The hepatic uptake of transferrin-bound iron by a nontransferrin receptor (NTR)-mediated process was investigated using the human hepatoma cell line HuH7. Because HuH7 cells also acquire iron from transferrin by a receptor (TR)-mediated process, TR expression was inhibited by transfecting the cells with a plasmid containing human TR complementary DNA in antisense orientation relative to a human cytomegalovirus promoter/enhancer element. Cell clones were obtained that expressed a 50% to 60% reduction in cell surface TR, leading to a corresponding decrease in transferrin and iron uptake compared with wild-type cells. Uptake of transferrin by a second process was nonsaturable and not inhibited by a 100-fold excess of unlabeled transferrin. The amounts of transferrin taken up by the wild-type and antisense cells by this process were similar, showing that it did not involve TR. The proteolytic enzyme Pronase reduced the uptake of transferrin, suggesting that the NTR-mediated process entailed the nonsaturable binding of transferrin to plasma membrane proteins. This process, like the TR-mediated one, involved the internalization and recycling of transferrin, leading to accumulation of iron with time. Iron uptake mediated by NTR process was saturable and displaced by 100-fold excess unlabeled transferrin and reduced by weak bases and metabolic inhibitors. Therefore, the NTR-mediated process entailed transferrin adsorption to membrane-bound proteins, internalization, and release of iron from transferrin by a pH-dependent step followed by the intracellular transport of iron into ferritin and heme by a saturable carrier-mediated mechanism.
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Department of Physiology and Biophysics, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Zak
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Aisen
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D001693:Biological Transport, Active; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism*; D016373:DNA, Antisense / Q000235:genetics / Q000494:pharmacology; D015870:Gene Expression; D006801:Humans; D007501:Iron / Q000378:metabolism; D007700:Kinetics; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism*; D011990:Receptors, Transferrin / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D014168:Transferrin / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D016373:DNA, Antisense; D011990:Receptors, Transferrin; D014168:Transferrin; C028283:diferric transferrin; D007501:Iron