Protein kinase B activation by reactive oxygen species is independent of tyrosine kinase receptor phosphorylation and requires SRC activity.
Reactive oxygen species (ROS) participate as second messengers in the mitogenic signal transduction. Most of the experimental data supporting the role of ROS as signaling molecules have been obtained by using H2O2. Exposure of cells to H2O2 rapidly increases tyrosine phosphorylation of tyrosine kinase receptors (TKRs) in the absence of growth factor binding, thus inducing the activation of downstream signaling cascades, like that of protein kinase B (AKT). Another molecule able to induce an increase of intracellular ROS levels is diethylmaleate (DEM), which acts by depleting the ROS scavenger reduced glutathione (GSH). A comparison of the effects exerted by H2O2 and DEM shows that the latter induces redox modifications milder than those generated by H2O2. We also demonstrated that DEM-induced redox modifications are not accompanied by platelet-derived growth factor-receptor (PDGF-R) and epidermal growth factor-receptor Tyr phosphorylation, although they are able to activate ERKs and AKT, with kinetics different from those observed following H2O2 treatment. The activation of these two pathways is not blocked by AG1296, a selective inhibitor of PDGF-R Tyr kinase, thus confirming that the effects of DEM are not mediated by the TKR phosphorylation. On the contrary, PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazole[3,4-d]pyrimidine), an inhibitor of Src kinase, completely prevents DEM- and H2O2-induced AKT activation but has no effect on the pathway of ERKs. Finally, nitration of Tyr residues in PDGF-R is observed in DEM-treated cells, thus suggesting that ROS-induced modifications different from Tyr phosphorylation can occur at the growth factor-receptor level and can be involved in the regulation of signaling pathways.
Esposito
Franca
F
Dipartimento di Biochimica e Biotecnologie Mediche, Universitá di Napoli Federico II, Via Pansini 5, Naples 80131, Italy. espositof@dbbm.unina.it
Chirico
Giuseppa
G
Montesano Gesualdi
Nicola
N
Posadas
Inmaculada
I
Ammendola
Rosario
R
Russo
Tommaso
T
Cirino
Giuseppe
G
Cimino
Filiberto
F
D000818:Animals; D002478:Cells, Cultured; D005347:Fibroblasts / Q000166:cytology; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007668:Kidney / Q000166:cytology; D008298:Maleates / Q000494:pharmacology; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016877:Oxidants / Q000494:pharmacology; D010084:Oxidation-Reduction; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D019061:src-Family Kinases / Q000378:metabolism*
D008298:Maleates; D016877:Oxidants; D011518:Proto-Oncogene Proteins; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; D020794:Receptor Protein-Tyrosine Kinases; D019061:src-Family Kinases; C494918:AKT1 protein, human; C494920:Akt1 protein, rat; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D020928:Mitogen-Activated Protein Kinases; C014476:diethyl maleate