Evidence on the operation of ATP-induced capacitative calcium entry in breast cancer cells and its blockade by 17beta-estradiol.
Little is known about the regulation of cytosolic calcium Ca(2+) levels ([Ca(2+)](i)) in breast cancer cells. We investigated the existence of capacitative calcium entry (CCE) in the tumorigenic cell line MCF-7 and its responsiveness to ATP. MCF-7 cells express purinergic receptors as well as estrogen receptors (ER). Depletion of calcium stores with thapsigargin (TG, 500 nM) or ATP (10 microM) in the absence of extracellular Ca(2+), resulted in a rapid and transient elevation in [Ca(2+)](i). After recovery of basal levels, Ca(2+) readmission (1.5 mM) to the medium increased Ca(2+) influx (twofold over basal), reflecting pre-activation of a CCE pathway. Cells pretreated with TG were unable to respond to ATP, thus indicating that the same Ca(2+) store is involved in their response. Moreover, IP(3)-dependent ATP-induced calcium mobilization and CCE were completely blocked using compound U-73122, an inhibitor of phospholipase C. Compound 2-APB (75 microM) and Gd(3+) (10 microM), antagonists of the CCE pathway, completely prevented ATP-stimulated capacitative Ca(2+) entry. CCE in MCF-7 cells was highly permeable to Mn(2+) and to the Ca(2+) surrogate Sr(2+). Mn(2+) entry sensitivity to Gd(3+) matched that of the Ca(2+) entry pathway. 17Beta-estradiol blocked ATP-induced CCE, but was without effect on TG-induced CCE. Besides, the estrogen blockade of the ATP-induced CCE was completely abolished by preincubation of the cells with an ER monoclonal antibody. ER alpha immunoreactivity could also be detected in a purified plasma membrane fraction of MCF-7 cells. These results represent the first evidence on the operation of a ATP-responsive CCE pathway in MCF-7 cells and also indicate that 17beta-estradiol interferes with this mechanism by acting at the cell surface level.
Rossi
Ana M
AM
Departamento de Biología, Bioquímica y Farmacia, Universidad Nacional del Sur, Bahia Blanca, Argentina.
Picotto
Gabriela
G
de Boland
Ana R
AR
Boland
Ricardo L
RL
D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors* / Q000494:pharmacology*; D001896:Boron Compounds / Q000494:pharmacology; D001943:Breast Neoplasms / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D045744:Cell Line, Tumor; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004958:Estradiol / Q000494:pharmacology*; D016257:Fura-2 / Q000737:chemistry / Q000378:metabolism; D005682:Gadolinium / Q000737:chemistry / Q000494:pharmacology; D006801:Humans; D019216:Metals, Heavy / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence / Q000379:methods; D019284:Thapsigargin / Q000494:pharmacology
D001896:Boron Compounds; D002121:Calcium Channel Blockers; D004791:Enzyme Inhibitors; D019216:Metals, Heavy; D004958:Estradiol; D019284:Thapsigargin; D000255:Adenosine Triphosphate; D005682:Gadolinium; C109986:2-aminoethoxydiphenyl borate; D002118:Calcium; D016257:Fura-2