P2X7 receptor activates extracellular signal-regulated kinases ERK1 and ERK2 independently of Ca2+ influx

Abstract

P2X7 nucleotide receptors modulate a spectrum of cellular events in various cells including epithelia, such as exocrine pancreas. Although the pharmacology and channel properties of the P2X7 receptors have been studied intensively, signal transduction pathways are relatively unknown. In this study we applied a heterologous expression system of rat P2X7 receptors in HEK-293 cells. We followed the receptor expression and function using the enhanced green fluorescent protein (EGFP) tag, activation of intracellular proteins and increases in cellular Ca2+. EGFP-P2X7 receptors localized to the plasma membrane, clusters within the membrane and intracellularly. Stimulation of P2X7 receptors in HEK-293 cells led to an activation of extracellular signal-regulated kinases ERK1 and ERK2 and this activation was seen after just 1 min of stimulation with ATP. Using C- and N-terminal P2X7-receptor mutants we show that the N-terminus is important in activation of ERKs, whereas deletion of the last 230 amino acids in the C-terminus did not effect ERK activation. On the other hand, Ca2+ entry was impaired in C-terminal but not in N-terminal mutants. In cell suspensions prepared from rat pancreas we show that P2X7 receptors also activate ERK1 and ERK2, indicating that these signalling pathways are also turned on in native epithelium.

A Role for Mitogen-activated Protein KinaseErk1/2 Activation and Non-selective Pore Formation in P2X7 Receptor-mediated Thymocyte Death

“…These two events are independent of the signaling pathway leading to Erk1/2 activation, whose control may depend on the N-terminal region of P2X7R (55). To the best of our knowledge, this is the first report implicating a role for both activation of MAP kinase Erk1/2 and non-selective pore opening in P2X7R-mediated thymocyte death.…”

“…Moreover, chelation of intracellular calcium with 1,2-bis(2-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetic acid tetrakis not only did not prevent death but instead increased it (data not shown). Erk1/2 phosphorylation is also Ca 2ϩ -independent in rat P2X7R-transfected HEK 293 cells (55) and parotid acinar cells (59). Calcium influx is, on the other hand, needed for Erk1/2 activation in astrocytoma (32) and Jurkat cells (54).…”

“…Calcium influx is, on the other hand, needed for Erk1/2 activation in astrocytoma (32) and Jurkat cells (54). Using various deletion mutants of rat P2X7R, Amstrup et al (55) have concluded that the N-terminal domain of P2X7R is essential for triggering Erk1/2 phosphorylation, whereas the C terminus domain controls Ca 2ϩ entry, suggesting that the two events may be dissociable in many cell types.…”

Purinergic receptors in the endocrine and exocrine pancreas

“…3b) 4 receptors, have an effect on secretion, it may be by means of up-regulating secretion rather than initiating it. The intracellular mechanisms stimulated by P2X receptors in pancreatic ducts are unknown, but they may involve Ca 2+ or Na + influx and an independent activation of mitogenactivated protein kinases, ERK1/2 [112]. Interestingly, in guinea pig pancreatic ducts that were cultured overnight injection of ATP into the lumen (or UTP) increased secretion [103].…”

Purinergic signaling in the lumen of a normal nephron and in remodeled PKD encapsulated cysts

“…Inhibitors of the extracellular signal-regulated kinases (ERK) slow cyst growth and expansion more dramatically. Both P2Y and P2X receptors can trigger ERK activation [156][157][158], as can growth factor receptors [159]. It is well known that purinergic receptor systems can transactivate growth factor receptor systems and cytokine receptor systems [134,160].…”