Extracellular ATP activates MAP kinase cascades through a P2Y purinergic receptor in the human intestinal Caco-2 cell line

Background: ATP exerts diverse effects on various cell types via specific purinergic P2Y receptors. Intracellular signaling cascades are the main routes of communication between P2Y receptors and regulatory targets in the cell. Methods and results: We examined the role of ATP in the modulation of ERK1/2, JNK1/2, and p38 MAP kinases (MAPKs) in human colon cancer Caco-2 cells. Immunoblot analysis showed that ATP induces the phosphorylation of MAPKs in a time- and dose-dependent manner, peaking at 5 min at 10 PM ATP. Moreover, ATP gamma S, UTP, and UDP but not ADP or ADP beta S increased phosphorylation of MAPKs, indicating the involvement of, at least, P2Y(2)/P2Y(4) and P2Y(6) receptor subtypes. RT-PCR studies and PCR product sequencing supported the expression of P2Y(2) and P2Y(4) receptors in this cell line. Spectrofluorimetric measurements showed that cell stimulation with ATP induced transient elevations in intracellular calcium concentration. In addition, ATP-induced phosphorylation of MAPKs in Caco-2 cells was dependent on Src family tyrosine kinases, calcium influx, and intracellular Ca2+ release and was partially dependent on the cAMP/PKA and PKC pathways and the EGFR. General significance: These findings provide new molecular basis for further understanding the mechanisms involved in ATP functions, as a signal transducer and activator of MAP kinase cascades, in colon adenocarcinoma Caco-2 cells. (C) 2009 Elsevier B.V. All rights reserved.