title: Thrombin and thrombin receptor agonist peptide induce tyrosine phosphorylation and tyrosine kinases in the platelet cytoskeleton. Translocation of pp60c-src and integrin alpha IIb beta 3 (glycoprotein IIb/IIIa) is not required for aggregation, but is dependent on formation of large aggregate structures.
abstract: The maximal aggregation of platelets induced by alpha-thrombin or by the receptor agonist peptide thrombin-(42-47)-peptide (TRP42/47) rapidly increased the pp60c-src associated with the cytoskeleton fraction. There was good correlation between the tyrosine kinase activity and the mass of pp60c-src. Tyrosine kinase activity associated with the cytoskeleton phosphorylated several endogenous cytoskeleton-associated proteins, as revealed by immunoblotting with anti-phosphotyrosine antibody following incubation with ATP in vitro. However, with the exception of pp60c-src, few phosphotyrosine-containing proteins were retained in the cytoskeleton in intact platelets when compared with total platelet lysates. Translocation of pp60c-src to the cytoskeleton induced by alpha-thrombin and TRP42/47 is dependent on glycoprotein IIb/IIIa (GPIIb/IIIa)-fibrinogen-mediated aggregation, but does not occur when ristocetin/von Willebrand factor produces GPIb-mediated platelet aggregation. The translocation of GPIIb/IIIa and pp60c-src to the cytoskeleton is not necessary for aggregation, as it is not seen when clearly visible small to moderate-sized aggregates are initially formed after exposure to thrombin. The linkage of these proteins to the cytoskeleton occurs only after later extensive formation of large aggregates. Translocation of GPIIa/IIIa to the cytoskeleton is not sufficient for the cytoskeletal association of pp60c-src, as the former occurs independently in platelets stimulated with concanavalin A in the absence of aggregation. Linkage of the integrin GPIIb/IIIa and pp60c-src to the internal cytoskeleton structure, and the corresponding tyrosine phosphorylation of certain proteins upon formation of large aggregates, may be an example of mechanochemical transduction by integrin receptors and may represent a structure with the requisite tensile strength to stabilize large platelet aggregates against high shear stresses.
authors.affiliation: Department of Pharmacology, School of Medicine, University of Connecticut Health Center, Farmington 06030.
authors.forename: K M
authors.initials: KM
authors.lastname: Pumiglia
authors.forename: M B
authors.initials: MB
authors.lastname: Feinstein
mesh_terms: D000595:Amino Acid Sequence; D001692:Biological Transport; D001792:Blood Platelets / Q000201:enzymology / Q000378:metabolism*; D003599:Cytoskeleton / Q000201:enzymology / Q000378:metabolism*; D004790:Enzyme Induction; D006801:Humans; D016023:Integrins / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000494:pharmacology; D010766:Phosphorylation; D010974:Platelet Aggregation; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000096:biosynthesis*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000187:drug effects*; D018179:Receptors, Thrombin; D012310:Ristocetin / Q000494:pharmacology; D013917:Thrombin / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*; D014841:von Willebrand Factor / Q000494:pharmacology
chemical_list: D016023:Integrins; D010446:Peptide Fragments; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; D018179:Receptors, Thrombin; C082835:thrombin receptor peptide (42-47); D014841:von Willebrand Factor; D012310:Ristocetin; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D016392:Proto-Oncogene Proteins pp60(c-src); D013917:Thrombin