Activation of src tyrosine kinases by peroxynitrite.
In this study, we demonstrate that the phosphorylation activity of five tyrosine kinases of the src family from both human erythrocytes (lyn, hck and c-fgr) and bovine synaptosomes (lyn and fyn) was stimulated by treatment with 30-250 microM peroxynitrite. This effect was not observed with syk, a non-src family tyrosine kinase. Treatment of kinase immunoprecipitates with 0.01-10 microM peroxynitrite showed that the interaction of these enzymes with the oxidant also activated the src kinases. Higher concentrations of peroxynitrite inhibited the activity of all kinases, indicating enzyme inactivation. The addition of bicarbonate (1.3 mM CO2) did not modify the upregulation of src kinases but significantly protected the kinases against peroxynitrite-mediated inhibition. Upregulation of src kinase activity by 1 microM peroxynitrite was 3.5-5-fold in erythrocytes and 1.2-2-fold in synaptosomes, but this could be the result, at least in part, of the higher basal level of src kinase activity in synaptosomes. Our results indicate that peroxynitrite can upregulate the tyrosine phosphorylation signal through the activation of src kinases.
Mallozzi
C
C
Laboratorio di Biologia Cellulare, Istituto Superiore di Sanità, Roma, Italy.
Di Stasi
A M
AM
Minetti
M
M
D000818:Animals; D002245:Carbon Dioxide / Q000494:pharmacology; D002417:Cattle; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000187:drug effects / Q000378:metabolism; D004912:Erythrocytes / Q000187:drug effects / Q000201:enzymology*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D009566:Nitrates / Q000494:pharmacology*; D009856:Oncogene Proteins, Viral / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000187:drug effects / Q000378:metabolism; D011506:Proteins / Q000187:drug effects / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000187:drug effects / Q000378:metabolism; D051076:Proto-Oncogene Proteins c-fyn; D051572:Proto-Oncogene Proteins c-hck; D015398:Signal Transduction; D000072377:Syk Kinase; D013574:Synaptosomes / Q000187:drug effects / Q000201:enzymology; D015854:Up-Regulation; D019061:src-Family Kinases / Q000187:drug effects / Q000378:metabolism*
D004792:Enzyme Precursors; D047908:Intracellular Signaling Peptides and Proteins; D009566:Nitrates; D009856:Oncogene Proteins, Viral; D011506:Proteins; D011518:Proto-Oncogene Proteins; D002245:Carbon Dioxide; C039642:peroxynitric acid; D011505:Protein-Tyrosine Kinases; C494957:FYN protein, human; C496631:HCK protein, human; D051076:Proto-Oncogene Proteins c-fyn; D051572:Proto-Oncogene Proteins c-hck; C000606896:SYK protein, human; D000072377:Syk Kinase; C060902:lyn protein-tyrosine kinase; C053102:proto-oncogene proteins c-fgr; D019061:src-Family Kinases