“ATP-dependent SAC influx does not participate in ATP modulation of MAPKs in MCF-7 human breast cancer cell line”

SCODELARO BILBAO, PAOLA GABRIELA; BOLAND, RICARDO; SANTILLÁN, GRACIELA

Washington DC, USA

Congreso; American Society for Biochemistry and Molecular Biology Annual Meeting; 2007

American Society of Biology and Molecular Biology

This study investigates the modulation of mitogen-activated protein kinases (MAPKs) ERK1/2, p38 and JNK1/2 and its relationship to changes in intracellular Ca2+ concentration ([Ca2+]i) induced by ATP in breast cancer MCF-7 cells. ATP, UTP but not ADP (5-10 µM) increased [Ca2+]i by cation release from inner stores as determined by spectrofluorometry measurements in a Ca2+ free medium (+0.5 mM EGTA). Slightly different responses were observed in medium with 1.5 mM Ca2+, indicating that Ca2+ release is the main component in this purinergic response. Moreover, ATP or UTP but not ADP stimulation sensitized cells to mechanical stress leading to Ca2+ influx. This mechanical stress activated Ca2+ (SAC) influx was inhibited by 10 mM Gd3+. These data suggest that SAC influx is dependent on P2Y2 receptor activation. Western blot analysis showed that ERK1/2, p38 and JNK1 were rapidly (5 min) phosphorylated by 5 µM ATP, whereas the JNK2 isoform was activated after 15 min treatment with ATP.  Activation of MAPKs was reduced by the use of 1 mM neomycin and 150 mM 2-APB, whereas the use of a Ca2+ free medium (+ 0.5 mM EGTA) or 10 mM Gd3+, a SAC channel inhibitor, had no effects. These results evidence an ATP-dependent SAC influx in MCF-7 cells which, differently to other cell types (Santillán and col., 2006), does not participate in modulation of MAPKs by ATP, effect only dependent on PI-PLC/IP3/Ca2+ release.