TUBULIN IS NOT PHOSPHORYLATED IN RESTING AND THROMBIN-ACTIVATED PLATELETS

We have investigated tubulin phosphorylation in human platelets, in order to evaluate whether it might be involved in the microtubular marginal band reorganization during platelet activation, Tubulin was identified with the use of specific monoclonal antibodies directed against alpha and beta subunits of tubulin, After metabolic P-32-labeling of platelets and analysis of separated proteins from whole cells, no phosphorylation of tubulin could be detected on autoradiography of platelet proteins either in resting platelets or during thrombin-induced activation, We also analyzed tubulin-enriched cytoskeletal fractions of resting or thrombin-stimulated platelets prepared in the presence of taxol, in comparison with tubulin-deprived cytoskeletal fractions prepared in the absence of this microtubule-stabilizing drug, Neither polymeric tubulin, assembled in microtubules and belonging to the platelet cytoskeleton, nor dimeric soluble tubulin showed significant P-32 labeling, Finally, no tubulin was recovered among tyrosine-phosphorylated platelet proteins immunoprecipitated with a specific anti-phosphotyrosine protein monoclonal antibody, Thus, human platelet tubulin is not phosphorylated either in unstimulated platelets or in thrombin-stimulated platelets, The fact that both alpha and beta subunits are involved appears to be a unique feature of platelets in comparison with other cells, Microtubule-associated proteins are more likely to be involved in the unbundling of the platelet marginal band,