Abstract
The calcium-sensing receptor (CaR) is expressed in epithelial ducts of both normal human breast and breast cancer tissue, as well as in the MCF-7 cell line as assessed by immunohistochemistry and Western blot analysis. However, to date, there are no data regarding the transduction pathways of CaR in breast cancer cells. In this study, we show that a CaR agonist, spermine, and increased extracellular Ca2+ ([Ca2+]o) sequentially activate two inward currents at –80 mV. The first was highly permeable to Ca2+ and inhibited by 2-aminophenyl borate (2-APB). In contrast, the second was more sensitive to Na+ and Li+ than to Ca2+ and insensitive to 2-APB. Furthermore, intracellular dialysis with high Mg2+, flufenamic acid or amiloride perfusion was without any effect on the second current. Both currents were inhibited by La3+. Calcium imaging recordings showed that both [Ca2+]o and spermine induced an increase in intracellular calcium ([Ca2+]i) and that removal of extracellular Ca2+ or perfusion of 2-APB caused a decline in [Ca2+]i. It is well known that stimulation of CaR by an increase in [Ca2+]o or with spermine is associated with activation of phospholipase C (PLC). Inhibition of PLC reduced the [Ca2+]o-stimulated [Ca2+]i increase. Lastly, reverse-transcriptase polymerase chain reaction showed that MCF-7 cells expressed canonical transient receptor potential (TRPCs) channels. Our results suggest that, in MCF-7 cells, CaR is functionally coupled to Ca2+-permeable cationic TRPCs, for which TRPC1 and TRPC6 are the most likely candidates for the highly selective Ca2+ current. Moreover, the pharmacology of the second Na+ current excludes the involvement of the more selective Na+ transient receptor potential melastatin (TRPM4 and TRPM5) and the classical epithelial Na+ channels.
Similar content being viewed by others
References
Beech D.J. 2005. TRPC1: Store-operated channel and more. Pfluegers Arch. 451:53–60
Boring C.C., Squire T.S., Tong T., Montgomery S. 1994. Cancer statistics. CA Cancer J. Clin. 44:7–26
Broad L.M., Braun F.J., Lievremont J.P., Bird G.S., Kurosaki T., Putney J.W., Jr. 2001. Role of the phospholipase C-inositol 1,4,5-trisphosphate pathway in calcium release-activated calcium current and capacitative calcium entry. J. Biol. Chem. 276:15945–15952
Brown E.M., Enyedi P., LeBoff M., Rotberg J., Preston J., Chen C. 1987. High extracellular Ca2+ and Mg2+ stimulate accumulation of inositol phosphates in bovine parathyroid cells. FEBS Lett. 218:113–138
Brown E.M., Gamba G., Riccardi D., Lombardi M., Butters R., Kifor O., Sun A., Hediger M.A., Lytton J., Hebert S.C. 1993. Cloning and characterization of an extracellular Ca2+-sensing receptor from bovine parathyroid. Nature 366:575–580
Brown E.M., MacLeod R.J. 2001. Extracellular calcium sensing and extracellular calcium signalling. Physiol. Rev. 81:239–297
Buchs N., Manen D., Bonjour J.P., Rizzoli R. 2000. Calcium stimulates parathyroid hormone-related protein production in Leydig tumor cells through a putative cation-sensing mechanism. Eur. J. Endocrinol. 142:500–505
Chattopadyay N., Evliyaoglu C., Heese O., Carroll R., Sanders J., Black P., Brown E.M. 2000. Regulation of secretion of PTHrP by Ca2+-sensing receptor in human astrocytes, astrocytomas, and meningiomas. Am. J. Physiol. 279:C691-C699
Cheng I., Klingensmith M.E., Chattopadhyay N., Kifor O., Butters R.R., Soybel D.I., Brown E.M. 1998. Identification and localization of the extracellular calcium-sensing receptor in human breast. J. Clin. Endocrinol. Metab. 83:703–707
Falzon M., Du P. 2000. Enhanced growth of MCF-7 breast cancer cells overexpressing parathyroid hormone-related peptide. Endocrinology 141:1882–1892
Fatherazi S., Belton C.M., Cai S., Zarif S., Goodwin P.C., Lamont R.J., Izutsu K.T. 2004. Calcium receptor message, expression and function decrease in differentiating keratinocytes. Pfluegers Arch. 448:93–104
Guise T.A., Yin T.A., Taylor S.D., Kumagai Y., Dallas M., Boyce B.F., Yoneda T., Mundy G.R. 1996. Evidence for a causal role of parathyroid hormone-related protein in the pathogenesis of human breast cancer-mediated osteolysis. J. Clin. Invest. 98:1544–1549
Huang C., Handlogten M.E., Miller R.T. 2002. Parallel activation of phosphotidylinositol 4-kinase and phospholipase C by the extracellular calcium-sensing receptor. J. Biol. Chem. 277:20293–20300
Jiang Y.F., Zhang Z., Kifor O., Lane C.R., Suinn S.J., Bai M. 2002. Protein kinase C (PKC) phosphorylation of the Ca2+-sensing receptor (CaR) modulates functional interaction of G proteins with the CaR cytoplasmic tail. J. Biol. Chem. 277:50543–50549
Journe F., Dumon J.C., Kheddoumi N., Fox J., Laios I., Leclercq G., Body J.J. 2004. Extracellular calcium downregulates estrogen receptor alpha and increases its transcriptional activity through calcium-sensing receptor in breast cancer cells. Bone 35:479–488
Kraft R., Harteneck C. 2005. The mammalian melastatin-related transient receptor potential cation channels: An overview. Pfluegers Arch. 451:204–211
Li S., Huang S., Peng S.B. 2005. Overexpression of G protein-coupled receptors in cancer cells: Involvement in tumor progression. Int. J. Oncol. 27:1329–1338
Liapis H., Crouch E.C., Grosso L.E., Kitazawa S., Wick M.R. 1993. Expression of parathyroidlike protein in normal, proliferative, and neoplastic human breast tissues. Am. J. Pathol. 174:1169–1178
Mundy G.R. 1997. Mechanisms of bone metastasis. Cancer 80:1546–1556
Nemeth E.F., Scarpa A. 1987. Rapid mobilization of cellular Ca2+ in bovine parathyroid cells evoked by extracellular divalent cations. Evidence for a cell surface calcium receptor. J. Biol. Chem. 262:5188–5196
Ouadid-Ahidouch H., Roudbaraki M., Delcourt P., Ahidouch A., Joury N., Prevarskaya N. 2004. Functional and molecular identification of intermediate-conductance Ca2+-activated K+ channels in breast cancer cells: Association with cell cycle progression. Am. J. Physiol. 287:C125-C134
Parekh A.B., Putney J.W., Jr. 2005. Store-operated calcium channels. Physiol. Rev. 85:757–810
Parkash J., Chaudhry M.A., Rhoten W.B. 2004. Calbindin-D28k and calcium sensing receptor cooperate in MCF-7 human breast cancer cells. Int. J. Oncol. 24:1111–1119
Pedersen S.F., Owsianik G., Nilius B. 2005. TRP channels: An overview. Cell Calcium 38:233–252
Rodland K.D. 2004. The role of the calcium-sensing receptor in cancer. Cell Calcium 35:291–295
Sanders J.L., Chattopadhyay N., Kifor O., Yamaguchi T., Butter R.R., Brown E.M. 2000. Extracellular calcium-sensing receptor expression and its potential role in regulating parathyroid hormone-related peptide secretion in human breast cancer cell lines. Endocrinology 141:4357–4364
Sanders J.L., Chattopadhyay N., Kifor O., Yamaguchi T., Butter R.R., Brown E.M. 2001. Ca2+-sensing receptor expression and PTHrP secretion in PC-3 human prostate cancer cells. Am. J. Physiol. 281:E1267-E1274
Shen X., Qian L., Falzon M. 2004. PTH-related protein enhances MCF-7 breast cancer cell adhesion, migration, and invasion via an intracrine pathway. Exp. Cell. Res. 294:420–433
Silver I.A., Murrills R.J., Etherington D.J. 1988. Microelectrode studies on the acid microenvironment beneath adherent macrophages and osteoclast. Exp. Cell. Res. 175:266–276
Ullrich N.D., Voets T., Prenen J., Vennekens R., Talavera K., Droogmans G., Nilius B. 2005. Comparison of functional properties of the Ca2+-activated cation channels TRPM4 and TRPM5 from mice. Cell Calcium 37:267–278
VanHouten J.N. 2005. Calcium sensing by the mammary gland. Neoplasia 10:129–139
VanHouten J.N., Danna P., McGeoch G., Brown E.M., Krapcho K., Neville M., Wysolmerski J.J. 2004. The calcium-sensing receptor regulates mammary gland parathyroid hormone-related protein production and calcium transport. J. Clin. Invest. 113:598–608
Yamaguchi T., Ye C., Chattopadhyay N., Sanders J.L., Vassilev P.M., Brown E.M. 2000. Enhanced expression of extracellular calcium sensing receptor in monocyte-differentiated versus undifferentiated HL-60 cells: Potential role in regulation of a nonselective cation channel. Calcif. Tissue Int. 66:375–382
Ye C.P., Kanazirskia M., Quinn S., Brown E.M., Vassilev P.M. 1996a. Modulation by polycationic Ca2+-sensing receptor agonists of nonselective cation channels in rat hippocampal neurons. Biochem. Biophys. Res. Commun. 224:271–280
Ye C.P., Rogers K., Bai M., Quinn S., Seidman C.E., Seidman J.G., Brown E.M., Vassilev P.M. 1996b. Agonists of the Ca2+-sensing receptor (CaR) activate nonselective cation channels in HEK293 cells stably transfected with the human CaR. Biochem. Biophys. Res. Commun. 226:272–279
Ye C.P., Ho-Pao C.L., Kanazirskia M., Quinn S., Rogers K., Seidman C.E., Seidman J.G., Brown E.M., Vassilev P.M. 1997. Amyloid-beta proteins activate Ca2+-permeable channels through calcium-sensing receptors. J. Neurosci. Res. 47:547–554
Zitt C., Halaszovich C.R., Luckhoff A. 2002. The TRP family of cation channels: Probing and advancing the concepts on receptor-activated calcium entry. Prog. Neurobiol. 66:243–264
Acknowledgments
We thank Jean François Lefebvre and Philippe Delcourt for their excellent technical assistance. This work was supported by the Ministère de l’Education Nationale, the Ligue Nationale Contre le Cancer, the Association pour la Recherche Contre le Cancer and the Region Picardie, France and by grants from Morocco.
Author information
Authors and Affiliations
Corresponding author
Additional information
This work concretizes the scientific cooperation between Université de Picardie Jules Verne and Université Ibn-Zohr.
Rights and permissions
About this article
Cite this article
Hiani, Y.E., Ahidouch, A., Roudbaraki, M. et al. Calcium-Sensing Receptor Stimulation Induces Nonselective Cation Channel Activation in Breast Cancer Cells. J Membrane Biol 211, 127–137 (2006). https://doi.org/10.1007/s00232-006-0017-2
Received:
Revised:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00232-006-0017-2