DIFFERENTIAL ACTIVATION OF CYTOSOLIC PHOSPHOLIPASE A(2) (CPLA(2)) BY THROMBIN AND THROMBIN RECEPTOR AGONIST PEPTIDE IN HUMAN PLATELETS - EVIDENCE FOR ACTIVATION OF CPLA(2) INDEPENDENT OF THE MITOGEN-ACTIVATED PROTEIN-KINASES ERK1 2/

The thrombin receptor agonist peptide SFLLRN was less effective than t hrombin in eliciting the liberation of arachidonic acid and the genera tion of thromboxane A(2) by human platelets, We found that while SFLLR N evokes an initial transient increase in cytosolic free calcium conce ntration ([Ca2+](i)) of similar magnitude as that caused by thrombin, the SFLLRN-induced elevation of [Ca2+](i) declines more rapidly to nea r resting levels than that evoked by thrombin, suggesting that dispara te levels of [Ca2+](i) may contribute to the attenuated arachidonic ac id release. Furthermore, we observed that SFLLRN is less effective tha n thrombin in mediating the ''activating'' phosphorylation of cytosoli c phospholipase A(2) (cPLA(2)). Both thrombin and SFLLRN rapidly and t ransiently activated kinases that phosphorylate the 21-residue synthet ic peptide Thr(669) derived from the epidermal growth factor receptor, but the maximal activation of proline-directed kinases by SFLLRN was less pronounced than that by thrombin. MonoQ chromatography and immuno blot analysis of extracts from stimulated platelets revealed that whil e thrombin induced a prominent activation of the mitogen-activated pro tein kinases ERK1 and ERK2, SFLLRN completely failed to do so. On the other hand, SFLLRN, like thrombin, stimulated the activity of a prolin e-directed kinase distinct from ERK1/2, but the activation of this kin ase was less pronounced following stimulation of platelets with SFLLRN compared with thrombin. We conclude 1) that the partial activation of cPLA(2) and the subsequent attenuated mobilization of arachidonic aci d in response to SFLLRN may be the consequence of a less prolonged ele vation of [Ca2+](i) and insufficient activation of proline-directed ki nase(s) by SFLLRN and 2) that the ability of SFLLRN to mediate the act ivating phosphorylation of cPLA(2) in the absence of ERK1/2 stimulatio n suggests that, at least in human platelets, proline-directed kinases other than ERK1/2 may phosphorylate and activate cPLA(2).