PEROXYNITRITE-INDUCED TYROSINE NITRATION AND PHOSPHORYLATION IN HUMANPLATELETS

Peroxynitrite (ONOO-) induces nitration of tyrosine residues and inhib its tyrosine phosphorylation in cell free systems. We investigated the effect of peroxynitrite on protein tyrosine nitration and phosphoryla tion in resting or thrombin-activated platelets. Peroxynitrite (150 mu M) rapidly induced tyrosine nitration of 187, 164, 113, 89, and 61 kD a proteins in gel-filtered platelets which persisted up to 4.5 h. Repe ated exposure of platelets to peroxynitrite produced increasing levels of nitration. Peroxynitrite also rapidly increased tyrosine phosphory lation of 120, 117, 95, 80-85, and 70 kDa platelet proteins, but this decreased by 5 min. The same pattern of tyrosine phosphorylation, but with higher intensity, was induced by thrombin in control platelets. P retreatment of platelets with peroxynitrite decreased thrombin-induced tyrosine phosphorylation at 0.05 and 1 U/ml thrombin but not at 2 U/m l thrombin. Platelet activation responses such as P-selectin expressio n, serotonin secretion, and aggregation were also decreased by peroxyn itrite treatment at low thrombin concentrations. Peroxynitrite exposur e and tyrosine nitration decreased platelet sensitivity to thrombin bu t did not absolutely prevent tyrosine phosphorylation and other platel et responses. Copyright (C) 1997 Elsevier Science Inc.