Effects of alterations in cellular iron on biosynthesis of the transferrin receptor in K562 cells.

Abstract

Treatment of K562 cells, a human erythroleukemia cell line, with desferrioxamine raised the levels of the receptor for transferrin (Tf) two-to threefold over that of the control cells. The levels of receptor were reduced by at least 50 and 35% of that of the control in cells treated with diferric Tf and ferric ammonium citrate, respectively. These changes were of total cellular receptors with no alteration in the proportion of receptors found on the cell surface. The half-lives of the receptor were identical in cells treated with desferrioxamine, diferric Tf, or ferric ammonium citrate. Cells metabolically labeled with [35S]methionine showed a 2.5-fold increase in the rate of receptor synthesis when treated with desferrioxamine and a 35 and 65% decrease when treated with ferric ammonium citrate and diferric Tf, respectively. In vitro translations of. polyadenylated mRNA isolated from cells incubated with desferrioxamine showed a 2.5-fold increase in translatable mRNA for the receptor, whereas treatment of cells with ferric ammonium citrate and diferric Tf resulted in a 25 and 50% reduction, respectively, in translatable mRNA for this receptor.Iron is an essential constituent of all cells. It is required for the normal functioning of a wide array of cellular enzymes, including those of the respiratory pathway (2, 4, 6). Cells receive iron via the receptor-mediated endocytosis of irontransferrin (Tf). Large amounts of iron can be stored in cells in ferritin and as hemosiderin. The iron needs of cells vary and are high in fetal, erythropoietic, and rapidly proliferating cells. Concomitant with these needs is a high level of expression of Tf receptors in placental cells, reticulocytes, and proliferating cells (1,8,10). It is not known exactly how cells regulate their iron uptake, but clearly the modulation of the number of Tf receptors can play a role in such regulation. Recently, Bridges et al. (Fed. Proc. 42:2193 and Mattia et al. (9) showed that K562 cells can alter Tf receptor numbers in response to intracellular iron chelation by desferrioxamine. The locus of this regulation was shown to be at the level of mRNA for the receptor (9). Ward et al. (12) reported a down regulation of Tf receptors in HeLa cells that were treated with iron salts. In this paper we report the extension of our studies on the regulation of Tf receptor biosynthesis in K562 cells. We show that delivering iron to cells results in a twofold decrease in specific synthesis of the Tf receptor. A decrease in the levels of mRNA is responsible for this drop. Furthermore, we compare the efficacy of Tf with that of ferric ammonium citrate in lowering receptor biosynthesis and establish the range of receptor regulation achievable with iron depletion or iron introduction without causing observable cellular toxicity.

Transferrin Receptor 2: Evidence for Ligand-induced Stabilization and Redirection to a Recycling Pathway

“…TfR1 expression is ubiquitous, consistent with its role in cellular iron delivery. The stability of TfR1 mRNA is negatively regulated by intracellular iron levels through iron-responsive elements (IREs) in the 3Ј untranslated region (Mattia et al, 1984;Ward et al, 1984;Rao et al, 1985;Sciot et al, 1987;Owen and Kuhn, 1987;Casey et al, 1988;Mullner and Kuhn, 1988;Lu et al, 1989;Mullner et al, 1989).…”

“…TfR1 expression is ubiquitous, consistent with its role in cellular iron delivery. The stability of TfR1 mRNA is negatively regulated by intracellular iron levels through iron-responsive elements (IREs) in the 3Ј untranslated region (Mattia et al, 1984;Ward et al, 1984;Rao et al, 1985;Sciot et al, 1987;Owen and Kuhn, 1987;Casey et al, 1988;Mullner and Kuhn, 1988;Lu et al, 1989;Mullner et al, 1989).TfR2 differs from TfR1 in notable ways. TfR2 binds Tf in a pH-dependent manner, but its affinity for Fe 2 Tf (K D ϳ30 nM; Kawabata et al, 2000;West et al, 2000) is significantly lower than that of TfR1 (K D ϳ1 nM, Tsunoo and Sussman, 1983;Enns et al, 1991;Richardson and Ponka, 1997).…”

Transcriptional regulation by iron of the gene for the transferrin receptor.

“…The receptor-Tf complex is internalized by a process of receptor-mediated endocytosis in which the iron is dissociated from the Tf (3). Investigators have previously shown that the levels of intracellular iron regulate the level of expression of the Tf receptor (6, 8,9,14). Raising the levels of intracellular iron by treatment of K562 cells (a human erythroleukemic cell line) with either human diferric Tf or ferric ammonium citrate results in decreased levels of the receptor.…”

“…These changes in cellular content of the Tf receptor through manipulation of intracellular iron levels are caused by altered rates of biosynthesis of the receptor. Corresponding changes in the levels of translatable mRNA for the receptor were measured by in vitro translation (9).…”

“…After the incubation, EDTA and SDS were added to achieve final concentrations of 10 mM and 0.2%, respectively. After extraction with phenol-chloroform, the RNA was precipitated with 2 (6,9). To examine whether this increase was due to increases in the steadystate level of the mRNA for the receptor, we isolated poly(A)+ mRNA from K562 cells that had been treated with 50 ,uM desferrioxamine, a concentration sufficient to give the maximal effect of the chelator.…”