Summary
We studied the factors that determine the differing growth requirements of low-iron-tolerant (LIT) versus high-iron-dependent (HID) cells for extracellular nontransferrin iron. The growth of LIT cells HeLa and THP-1, when transferred from transferrin (5 µg/ml) medium into low-iron (5 µM ferric citrate) medium, was not significantly affected while HID cells Jiyoye and K562 showed nearly no growth. HeLa and THP-1 cells, as well as Jiyoye and K562 cells, do not produce transferrin in sufficient amounts to support their growth in low-iron medium. Surprisingly, similar rates of iron uptake in low-iron medium (0.033 and 0.032 nmol Fe/min and 106 cells) were found for LIT cells HeLa and HID cells K562. Furthermore, the intracellular iron level (4.64 nmol/106 cells) of HeLa cells grown in low-iron medium was much higher than iron levels (0.15 or 0.20 nmol/106 cells) of HeLa or K562 cells grown in transferrin medium. We demonstrated that the activity (ratio activated/total) of the iron regulatory protein (IRP) in HID cells Jiyoye and K562 increased more than twofold (from 0.32 to 0.79 and from 0.47 to 1.12, respectively) within 48 h after their transfer into low-iron medium. In the case of LIT cells HeLa and THP-1, IRP activity stayed at similar or slightly decreased levels (0.86–0.73 and 0.58–0.55, respectively). Addition of iron chelator deferoxamine (50 µM, i.e., about half-maximal growth-inhibitory dose) resulted in significantly increased activity of IRP also in HeLa and THP-1 cells. We hypothesize that the relatively higher bioavailability of nontransferrin iron in LIT cells, over that in HID cells, determines the differing responses observed under low-iron conditions.
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References
Aisen, P.; Listowsky, I. Iron transport and storage proteins. Annu. Rev. Biochem. 49:357–393; 1980.
Basset, P.; Quesneau, Y.; Zwiller, J. Iron-induced L1210 cell growth: evidence of a transferrin-independent iron transport. Cancer Res. 46:1644–1647; 1986.
Brissot, P.; Wright, T. L.; Ma, W. L., et al. Efficient clearance of non-transferrin-bound iron by rat liver. Implications for hepatic iron loading in iron overload states. J. Clin. Invest. 76:1463–1470; 1985.
Brock, J. H.; Stevenson, J. Replacement of transferrin in serum-free cultures of mitogen-stimulated mouse lymphocytes by a lipophilic iron chelator. Immunol. Lett. 15:23–25; 1987.
Carmichael, J.; DeGraff, W. G.; Gazdar, A. F., et al. Evaluation of a tetrazolium-based semiautomated colorimetric assay: assessment of chemosensitivity testing. Cancer Res. 47:936–942; 1987.
Casey, J. L.; Hentze, M. W.; Koeller, D. M., et al. Iron-responsive elements: regulatory RNA sequences that control mRNA levels and translation. Science 240:924–928; 1988.
Chen, T. R. In situ detection of mycoplasma contamination in cell cultures by fluorescent Hoechst 33258 stain. Exp. Cell Res. 104:255–262; 1977.
Chitambar, C. E.; Matthaeus, W. G.; Antholine, W. E., et al. Inhibition of leukemic HL-60 cell growth by transferrin-gallium: effects on ribonucleotide reductase and demonstration of drug synergy with hydroxyurea. Blood 72:1930–1936; 1988.
Dezza, L.; Cazzola, M.; Danova, M., et al. Effects of desferrioxamine on normal and leukemic human hematopoietic cell growth: in vitro and in vivo studies. Leukemia 3:104–107; 1989.
Hansen, M. B.; Nielsen, S. E.; Berg, K. Re-examination and further development of a precise and rapid dye method for measuring cell growth/cell kill. J. Immunol. Methods 119:203–210; 1989.
Hedley, D. W.; Tripp, E. H.; Slowiaczek, P., et al. Effect of gallium on DNA synthesis by human T-cell lymphoblasts. Cancer Res. 48:3014–3018; 1988.
Hoffbrand, A. V.; Ganeshaguru, K.; Hooton, J. W. L., et al. Effect of iron deficiency and desferrioxamine on DNA synthesis in human cells. Br. J. Haematol. 33:517–526; 1976.
Inman, R. S.; Wessling-Resnick, M. Characterization of transferrin-independent iron transport in K562. J. Biol. Chem. 268:8521–8528; 1993.
Kaplan, J.; Jordan, I.; Sturrock, A. Regulation of the transferrin-independent iron transport system in cultured cells. J. Biol. Chem. 266:2997–3004; 1991.
Kaplinsky, C.; Estrov, Z.; Freedman, M. H., et al. Effect of deferoxamine on DNA synthesis, DNA repair, cell proliferation, and differentiation of HL-60 cells. Leukemia 1:437–441; 1987.
Klausner, R. D.; Rouault, T. A.; Harford, J. B. Regulating the fate of mRNA: the control of cellular iron metabolism. Cell 72:19–28; 1993.
Koeller, D. M.; Casey, J. L.; Hentze, M. W., et al. A cytosolic protein binds to structural elements within the iron regulatory region of the transferrin receptor mRNA. Proc. Natl. Acad. Sci. USA 86:3574–3578; 1989.
Kovar, J. Growth-stimulating effect of ferric citrate on hybridoma cells: characterization and relation to transferrin function. Hybridoma 7:255–263; 1988.
Kovar, J.; Franek, F. Iron compounds at high concentrations enable hybridoma growth in a protein-free medium. Biotechnol. Lett. 9:259–264; 1987.
Kovar, J.; Franek, F. Growth-stimulating effect of transferrin on a hybridoma cell line: relation to transferrin iron-transporting function. Exp. Cell Res. 182:358–369; 1989.
Kovar, J.; Neumannova, V.; Kriegerbeckova, K. Iron and cell growth in vitro. In Vitro Cell. Dev. Biol. 29A:115A (Abstract); 1993.
Kriegerbeckova, K.; Kovar, J.; Neumannova, V. Production of transferrin by human cell lines in a defined protein-free system: detection by an ELISA. Immunol. Cell Biol. 71:303–309; 1993.
Kühn, L. C.; Hentze, M. W. Coordination of cellular iron metabolism by post-transcriptional gene regulation. J. Inorg. Biochem. 47:183–195; 1992.
Kühn, L.; Schulman, M.; Ponka, P. Iron-transferrin requirements and transferrin receptor expression in proliferating cells. In: Ponka, P.; Schulman, H. M.; Woodworth, R. C., eds. Iron transport and storage. Boca Raton: CRC Press; 1990:149–191.
Laskey, J.; Webb, I.; Schulman, H. M., et al. Evidence that transferrin supports cell proliferation by supplying iron for DNA synthesis. Exp. Cell Res. 176:87–95; 1988.
Leibold, E. A.; Munro, H. N. Cytoplasmic protein binds in vitro to a highly conserved sequence in the 5′ untranslated region of ferritin heavy- and light-subunit mRNAs. Proc. Natl. Acad. Sci. USA 85:2171–2175; 1988.
May, W. S.; Cuartecasas, P. Transferrin receptor: its biological significance. J. Membr. Biol. 88:205–215; 1985.
Mosmann, T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assay. J. Immunol. Methods 65:55–63; 1983.
Müllner, E. W.; Neupert, B.; Kühn, L. C. A specific mRNA-binding factor regulates the iron-dependent stability of cytoplasmic transferrin receptor mRNA. Cell 58:373–382; 1989.
Neumannova, V.; Richardson, D. R.; Kriegerbeckova, K., et al. Growth of human tumor cell lines in transferrin-free, low-iron medium. In Vitro Cell. Dev. Biol. 31:625–632; 1995.
Nocka, K. H.; Pelus, L. M. Cell cycle specific effects of deferoxamine on human and murine hematopoietic progenitor cells. Cancer Res. 48:3571–3575; 1988.
Qian, Z. M.; Morgan, E. H. Effect of metabolic inhibitors on uptake of non-transferrin-bound iron by reticulocytes. Biochim. Biophys. Acta 1073:456–462; 1991.
Rothenberger, S.; Müllner, E. W.; Kühn, L. C. The mRNA-binding protein which controls ferritin and transferrin receptor expression is conserved during evolution. Nucleic Acids Res. 18:1175–1179; 1990.
Seligman, P. A.; Kovar, J.; Schleicher, R. B., et al. Transferrin-independent iron uptake supports B lymphocyte growth. Blood 78:1526–1531; 1991.
Sturrock, A.; Alexander, J.; Lamb, J., et al. Characterization of a transferrin-independent uptake system for iron in HeLa cells. J. Biol. Chem. 265:3139–3145; 1990.
Trowbridge, I. S.; Shackelford, D. A. Structure and function of transferrin receptors and their relationship to cell growth. Biochem. Soc. Symp. 51:117–129; 1985.
Tsao, M. S.; Sanders, G. S. H.; Grisham, J. W. Regulation of growth of cultured hepatic epithelial cells by transferrin. Exp. Cell Res. 171:52–62; 1987.
Vostrejs, M.; Moran, P. L.; Seligman, P. A. Transferrin synthesis by small cell lung cancer cells acts as an autocrine regulator of cellular proliferation. J. Clin. Invest. 82:331–339; 1988.
Walden, W. E.; Patino, M. M.; Gaffield, L. Purification of a specific repressor of ferritin mRNA translation from rabbit liver. J. Biol. Chem. 264:13765–13769; 1989.
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Kovář, J., Kühn, L.C., Richardson, V. et al. The inability of cells to grow in low iron correlates with increasing activity of their iron regulatory protein (IRP). In Vitro Cell.Dev.Biol.-Animal 33, 633–639 (1997). https://doi.org/10.1007/s11626-997-0114-2
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DOI: https://doi.org/10.1007/s11626-997-0114-2