Protein tyrosine phosphorylation induced by lysophosphatidic acid in rat- 1 fibroblasts. Evidence that phosphorylation of map kinase is mediated by the G(i)-p21(ras) pathway

Lysophosphatidic acid (LPA) is a platelet-derived phospholipid that serves as a mitogen for fibroblasts. LPA activates its own G protein-coupled receptor(s) leading to stimulation of phospholipase C and inhibition of adenylate cyclase. Furthermore, LPA rapidly activates p21(ras) through a pertussis toxin-sensitive pathway. In this study, we have examined LPA- induced protein tyrosine phosphorylation in Rat-1 fibroblasts. LPA action was compared with that of endothelin, which is a stronger activator of phospholipase C than LPA but fails to activate p21(ras) and to stimulate DNA synthesis in these cells. LPA and, more effectively, endothelin rapidly stimulate tyrosine phosphorylation of proteins of 110-130, 95, and 65-75 kDa. The effect of LPA is dose- and time-dependent, being half-maximal at 3-30 nM and peaking after 2-5 min. Among the 110-130-kDa group of phosphotyrosyl proteins is the 125-kDa 'focal adhesion kinase' (p125(FAK)) but not the 120- kDa p21(ras) GTPase-activating protein. Furthermore, LPA, like epidermal growth factor, causes tyrosine phosphorylation and activation of the p42/p44 mitogen-activated protein (MAP) kinases, paralleling p21(ras) activation. In contrast, endothelin fails to phosphorylate MAP kinase. Treatment of the cells with pertussis toxin blocks LPA-induced MAP kinase phosphorylation without affecting the other tyrosine phosphorylations. The kinase inhibitor staurosporine (1 μM) blocks LPA-induced, but not epidermal growth factor- induced, activation of p21(ras) and MAP kinase, consistent with an intermediate protein kinase linking the LPA receptor to p21(ras) activation. The results support a model in which LPA-induced phosphorylation of MAP kinase is mediated by p21(ras), and tyrosine phosphorylation of the other substrates, including p125(FAK), is associated with phospholipase C activation.