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Zinc Deficiency Induced in Swiss 3T3 Cells by a Low-Zinc Medium Impairs Calcium Entry and Two Mechanisms of Entry Are Involved


Zinc deficiency in 3T3 cells induced by the use of diethylenetriaminepentaacetate (DTPA) has been shown to impair calcium entry associated with failure of proliferation when the cells are stimulated with polypeptide growth factors (GF). These functions of zinc have been evaluated here in the same clone of cells by simple depletion using a low-zinc medium (0.05 μmol/L zinc) without chelator. Confluent cells were maintained for 1 day in the low-zinc medium without GF, then loaded with Fluo-4, and stimulated with GF. Calcium entry was measured by the increase in sustained fluorescence. It was preceded by the release of stored calcium as observed in the previous study using DTPA. Zinc deprivation decreased calcium entry when calcium was added at 0 or 0.05 mmol/L but not when 0.1 mmol/L or higher. Cell proliferation reflected similar effects of zinc and calcium concentrations. In a newly acquired clone of 3T3 cells, GF did not induce internal calcium release but thapsigargin (TG) did. When added in a low-calcium medium, both agonists stimulated calcium entry when external calcium was added, suggesting that two different mechanisms of entry were impaired by zinc deficiency. Zinc deficiency produced by DTPA in the newer clones gave similar results, decreasing calcium entry induced by both agonists. The effects of GF and TG were not additive. The results confirm the earlier observation that zinc deficiency impairs calcium entry into 3T3 cells when stimulated by GF and show that the cells can take up calcium by either store-operated or receptor-operated mechanisms.

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Bovine serum albumin






Epidermal growth factor


Growth factors


Insulin-like growth factor-I


Inositol 1,4,5-trisphosphate


Low-zinc, low-calcium medium


Platelet-derived growth factor


Receptor-operated calcium entry


Store-operated calcium entry


Stromal interaction molecule 1, a calcium sensing protein




Transient receptor potential canonical


  1. Chesters JK, Petrie L, Vint H (1989) Specificity and timing of the Zn2+ requirement for DNA synthesis by 3T3 cells. Exp Cell Res 184:499–508

    PubMed  Article  CAS  Google Scholar 

  2. MacDonald RS, Wollard-Biddle LC, Browning JD, Thornton WH, O’Dell BL (1998) Zinc deprivation of murine 3T3 cells by use of diethylenetrinitrilo- pentataacetate impairs DNA synthesis upon stimulation with insulin-like growth factor-I (IGF-I). J Nutr 128:1600–1605

    PubMed  CAS  Google Scholar 

  3. O’Dell BL, Browning JD (2011) Zinc deprivation impairs growth-factor stimulated calcium influx into murine 3T3 cells associated with decreased cell proliferation. J Nutr 141:1036–1040

    PubMed  Article  Google Scholar 

  4. Putney JW, Bird GS (1993) The inositol phosphate-calcium signaling system in nonexcitable cells. Endocr Rev 14:610–631

    PubMed  CAS  Google Scholar 

  5. Lewis RS (2007) The molecular choreography of a store-operated calcium channel. Nature 446:284–287

    PubMed  Article  CAS  Google Scholar 

  6. Blakesley VA, Kalebic T, Helman LJ, Stannard B, Faria TN, Roberts CT, LeRoith D (1996) Tumorigenic and mitogenic capacities are reduced in transfected fibroblasts expressing mutant insulin-like growth factor (IGF-I) receptors. The role of tyrosine residues 1250, 1251, and 1316 in the carboxy-terminus of the IGFR-I receptor. Endocrinology 37:410–417

    Article  Google Scholar 

  7. Rink TJ (1990) Receptor-mediated calcium entry. FEBS 268:381–385

    Article  CAS  Google Scholar 

  8. Waymouth C (1974) “Feeding the baby”-designing the culture milieu to enhance cell stability. J Natl Cancer Institute 53:1443–1448

    CAS  Google Scholar 

  9. Dernison MM, Almirza WHMA, Kusters JMAM, van Meerwijk WPM, Gielen CCAM, van Zoelen EJJ, Theuenet APR (2010) Growth-dependent modulation of capacitative calcium entry in normal rat kidney fibroblasts. Cell Signal 22:1044–1053

    PubMed  Article  CAS  Google Scholar 

  10. Berridge MJ (1993) Inositol trisphosphate and calcium signaling. Nature 361:315–325

    PubMed  Article  CAS  Google Scholar 

  11. Wang Y, Deng X, Hewavitharana T, Soboloff J, Gill DL (2008) STIM, ORAI and TRPC channels in the control of calcium entry signals in smooth muscle. Clin Exp Pharmacol Physiol 35:1127–1133

    PubMed  Article  CAS  Google Scholar 

  12. Lee KP, Yuan JP, So I, Worley PF, Muallem S (2010) STIM1-dependent and STIM1-independent function of transient receptor potential canonical (TRPC) channels tune their store-operated mode. J Biol Chem 285:38666–38673

    PubMed  Article  CAS  Google Scholar 

  13. Salido GM, Jardin I, Rosado JA (2011) The TRPC ion channel: association with Orai1 and STIM1 proteins and participation in capacitative and non-capacitative calcium entry (Review). Adv Exp Biol Med 704:413–433

    Article  CAS  Google Scholar 

  14. Putney JW (2007) Recent breakthroughs in the molecular mechanism of capacitative calcium entry (with thoughts on how we got there). Cell Calcium 42:103–110

    PubMed  Article  CAS  Google Scholar 

  15. Venkatachalam K, Montell C (2007) TRP Channels. Annu Rev Biochem 76:387–417

    PubMed  Article  CAS  Google Scholar 

  16. Worley PF, Zeng W, Huang GN, Yuan JP, Kim JY, Lee MG, Muallem S (2007) TRPC channels as STIM1-regulated store-operated channels. Cell Calcium 42:205–211

    PubMed  Article  CAS  Google Scholar 

  17. Lee KP, Yuan JP, Hong JH, So I, Worley PF, Muallem S (2010) An endoplasmic/plasma membrane junction: STIM1/Orai1/TRPCs. FEBS Lett 584:2022–2027

    PubMed  Article  CAS  Google Scholar 

  18. Beck B, Zholos A, Sydorenko V, Roudbaraki M, Lehen’kyi V, Bordat P, Prevarskaya N, Skryma R (2006) TRPC7 is a receptor-operated DAG-activated channel in human keratinocytes. J Invest Dermatol 126:1982–1993

    PubMed  Article  CAS  Google Scholar 

  19. Gamberucci A, Giurisato E, Pizzo P, Tassi M, Giunti R (2002) Diacylglycerol activates the influx of extracellular cations in T-lymphocytes independently of intracellular calcium-store depletion and possibly involving endogenous gene products. Biochem J 364:245–254

    PubMed  CAS  Google Scholar 

  20. Tu P, Kunert-Keil C, Lucke S, Brinkmeier H, Bouron (2009) Diacylglycerol analogues activate second messenger-operated calcium channels exhibiting TRPC-like properties in cortical neurons. J Neurochem 108:126–138

    PubMed  Article  CAS  Google Scholar 

  21. Salmon MD, Ahluwalia J (2011) J. Pharmacology of receptor operated calcium entry in human. Intl Immunopharmacol 11:145–148

    Article  CAS  Google Scholar 

  22. O’Dell BL, Emery MP (1991) Compromised zinc status in rats adversely affects calcium metabolism in platelets. J Nutr 121:1763–1768

    PubMed  Google Scholar 

  23. Xia J, O’Dell BL (1995) Zinc deficiency in rats decreases thrombin-stimulated platelet aggregation by lowering protein kinase C secondary to calcium uptake. J Nutr Biochem 6:661–666

    Article  CAS  Google Scholar 

  24. Browning JD, O’Dell BL (1994) Low zinc status in guinea pigs impairs calcium uptake by brain synaptosomes. J Nutr 124:436–443

    PubMed  CAS  Google Scholar 

  25. Emery MP, Browning JD, O’Dell BL (1990) Impaired homeostasis platelet function in rats fed low zinc diets based on egg white protein. J Nutr 120:1062–1067

    PubMed  CAS  Google Scholar 

  26. O’Dell BL, Conley-Harrison J, Besch-Williford C, Browning JD, O’Brien D (1990) Zinc status and peripheral nerve function in guinea pigs. FASEB J 4:2919–2923

    PubMed  Google Scholar 

  27. Jankowski-Hennig MA, Clegg MS, Daston GP, Rogers JM, Keen CL (2000) Zinc-deficient rat embryos have increased caspase 3-like activity and apoptosis. Biochem Biophys Res Commun 271:250–256

    PubMed  Article  CAS  Google Scholar 

  28. Clegg MS, Hanna LA, Niles BJ, Momma TY, Keen CL (2005) Zinc deficiency-induced cell death. IUBMB Life 57:661–669

    PubMed  Article  CAS  Google Scholar 

  29. Bray TM, Bettger WJ (1990) The physiological role of zinc as an antioxidant. Free Radic Biol Med 8:281–291

    PubMed  Article  CAS  Google Scholar 

  30. McKenzie GG, Keen CL, Oteiza PI (2002) Zinc status of human IMR-32 neuroblastoma cells influences their susceptibility to iron-induced oxidative stress. Dev Neurosci 24:125–133

    Article  Google Scholar 

  31. King PI, Osati-Ashtiani F, Fraker PJ (2002) Apoptosis plays a distinct role in the loss of precursor lymphocytes during zinc deficiency in mice. J Nutr 132:974–979

    PubMed  CAS  Google Scholar 

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Correspondence to Boyd L. O’Dell.

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This study was supported by the University of Missouri Food for the 21st Century Program.

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O’Dell, B.L., Browning, J.D. Zinc Deficiency Induced in Swiss 3T3 Cells by a Low-Zinc Medium Impairs Calcium Entry and Two Mechanisms of Entry Are Involved. Biol Trace Elem Res 152, 98–104 (2013).

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  • Zinc deficiency
  • Calcium entry
  • Cell proliferation
  • Zinc–calcium interaction