Extracellular ATP stimulates an inhibitory pathway towards growth factor-induced cRaf-1 and MEKK activation in astrocyte cultures

ATP, acting via P2Y, G protein-coupled receptors (GPCRs), is a mitogenic si gnal and also synergistically enhances fibroblast growth factor-2 (FGF-2)-i nduced proliferation in astrocytes. Here, we have examined the effects of A TP and FGF-2 cotreatment on the main components of the extracellular-signal regulated protein kinase (ERK) cascade, cRaf-1, MAP/ERK kinase (MEK) and E RK, key regulators of cellular proliferation. Surprisingly, ATP inhibited a ctivation of cRaf-1 by FGF-2 in primary cultures of rat cortical astrocytes . The inhibitory effect did not diminish MEK and ERK activation; indeed, co treatment resulted in a greater initial activation of ERK. ATP inhibition o f cRaf-1 activation was not mediated by an increase in cyclic AMP levels or by protein kinase C activation. ATP also inhibited the activation of cRaf- 1 by other growth factors, epidermal growth factor and platelet-derived gro wth factor, as well as other MEK1 activators stimulated by FGF-2, MEK kinas e 1 (MEKK1) and MEKK2, Serotonin, an agonist of another GPCR coupled to ERK , did not inhibit FGF-2-induced cRaf-1 activation, thereby indicating speci ficity in the ATP-induced inhibitory cross-talk. These findings suggest tha t ATP stimulates an inhibitory activity that lays upstream of MEK activator s and inhibits growth factor-induced activation of cRaf-1 and MEKKs. Such a mechanism might serve to integrate the actions of receptor tyrosine kinase s and P2Y-GPCRs.