ROLE OF PROTEIN PHOSPHATASES IN BLOOD PLATELET FUNCTION

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The role of platelets in wound healing and inflammation processes has been well established. In performing these functions, platelets undergo massive biochemical and morphological changes. The molecular mechanism(s), however, by which platelet reactivity is controlled is poorly understood. Thus, why platelets fail to function normally in certain disease states remains an open question. Recognized as being important reactions in stimulusresponse coupling, the role of phosphorylation of platelet proteins has been studied by many laboratories including ours; but the involvement of specific phosphorylation reactions in regulating platelet responses remains unclear. The broad long-term objective of this application is to understand the role of these reactions in platelets. The phosphorylation status of proteins reflects a balance between protein kinase and phosphatase activities. Thus, there are several paths one can pursue to address this problem. The objectives of this research program are to characterize basic properties of platelet phosphatases and their roles. The proposed studies will concentrate on the protein serine/threonine phosphatase, PP1, focussing attention primarily on: (i) The physiochemical properties of this enzyme, which includes an understanding of its structure, subcellular localization, and substrate specificity making use of exogenous phosphoproteins. (ii) Identifying which of the biochemical event that occur in response to a platelet agonist depends on PP1 activity. These studies will make use of okadaic acid, a membrane permeable inhibitor of PP1, in attempt to define the intracellular pathway that requires PP1 to proceed normally. Finally, (iii) attempts will be made to identify the platelet phosphoproteins that are endogenous substrates for PP1. Characterizing PPl-sensitive phosphorylations may help identify proteins that regulated platelet reactivity.