Selective dephosphorylation of the threonine(183) residue of ERK2 upon (alpha)llb(beta)3 engagement in platelets.
Thrombin-induced extracellular signal-regulated kinase 2 (ERK2) activation is negatively regulated in conditions of all bP3 integrin engagement and platelet aggregation. Here we show by Western blotting with antibodies against mono- and biphosphorylated forms of ERK2 that the dephosphorylation of ERK2 by alpha llb beta 3 engagement affects threonine183 and not tyrosine185. Addition of a potent serine/threonine phosphatase inhibitor, okadaic acid (OA), restored thrombin-induced threonine phosphorylation of ERK2 in conditions of platelet aggregation, whereas OA had no effect in the absence of alpha llb beta 3 engagement. These observations are consistent with alpha llb beta 3 engagement acting via at least one serine/threonine phosphatase,which dephosphorylates the phosphothreonine183 residue of ERK2. Moreover, a small amount (14%) of ERK2 was translocated to the alpha llb beta 3-dependent cytoskeleton, mostly ina monophosphorylated (i.e. inactive) form, suggesting that cytoskeleton-associated ERK2 plays only a minor role, if any. Finally, we show that negative regulation (i.e. dephosphorylation)occurs primarily or totally in the cytosol and that the alpha llb beta 3-dependent ERK2 Thr183-specific phosphatase is different from phosphatase 1 (PP1) or PP2A. We conclude that all alpha llb beta 3 engagement down-regulates ERK2 through selective dephosphorylation of the phosphothreonine183 residue by a cytosolic serine/threonine phosphatase different from known platelet phosphatases.
Pawlowski
Marc
M
U348 INSERM, IFR-6 Circulation Lariboisière, Hôpital Lariboisière, Paris, France.
Ragab
Ashraf
A
Rosa
Jean-Philippe
JP
Bryckaert
Marijke
M
D000199:Actins / Q000378:metabolism; D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D002417:Cattle; D003599:Cytoskeleton / Q000378:metabolism; D003902:Detergents; D004789:Enzyme Activation; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D017830:Octoxynol; D019319:Okadaic Acid / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D010769:Phosphothreonine / Q000378:metabolism; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism*; D054645:Protein Phosphatase 1; D012995:Solubility; D013347:Subcellular Fractions; D013912:Threonine / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism / Q000494:pharmacology
D000199:Actins; D003902:Detergents; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D010769:Phosphothreonine; D019319:Okadaic Acid; D013912:Threonine; D017830:Octoxynol; D019950:Mitogen-Activated Protein Kinase 1; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D013917:Thrombin