Regulation of cytosolic PlA 2 activity by PP1/PP2A serine/threonine phosphatases in human platelets

Platelet thromboxane A 2 (TXA 2 ) synthesis is an important pathway of platelet reactivity. We report that in thrombin- stimulated platelets, PP1/PP2A serine/threonine phosphatases regulate phospholipase A 2 (cPLA 2 ) activity, which is required for TXA 2 synthesis. Two mechanisms are involved: (a) constitutively active PP1/PP2A regulate cPLA 2 phosphorylation, and (b) PP1/PP2A activity mediates agonist-induced increase in cytosolic Ca 2+ ([Ca 2+ ] i ). Inhibition of PP1/PP2A with okadaic acid (OA) induces cPLA 2 phosphorylation but reduces Ca 2+ responses: release from intracellular stores and influx through the plasma membrane, particularly that mediated by store-mediated Ca 2+ entry (SMCE). A significant correlation ( r =0.64) exists between OA-regulated [Ca 2+ ] i and TXA 2 synthesis. Okadaic acid- induced decrease in SMCE and the associated TXA 2 synthesis are mediated by a reduction in protein-tyrosine phosphorylation. This reduction is not due to inhibition of tyrosine kinases but rather to an OA-mediated increase in tyrosine phosphatases. This is the first study to report that PP1/PP2A phosphatases are involved in the regulation of the two key elements in eicosanoid synthesis, [Ca 2+ ] i and cPLA 2 phosphorylation. Moreover, PP1/PP2A regulation of [Ca 2+ ] i and tyrosine phosphorylation may be important for other calcium-dependent processes and/or signal transduction mechanisms in platelets.