Pronounced activation of protein kinase C, ornithine decarboxylase and c-jun proto-oncogene by paraquat-generated active oxygen species in WI-38 human lung cells.
Paraquat (methyl viologen, PQ) is a widely used herbicide that produces oxygen-derived free radicals and severely injures human lungs. In this study we examined the effects of PQ on the protein kinase C (PKC), ornithine decarboxylase (ODC) and c-jun oncogene expression in WI-38 human lung cells. Exposure of cells to 25-200 microM PQ resulted in an increase of [3H]phorbol dibutyrate (PDBu) binding and PKC redistribution in a dose-dependent manner. Interestingly, a superoxide dismutase mimic, 4-hydroxyl-2,2,6,6-tetramethyl-piperidine-1-oxyl (Tempol, 2.5 mM) and catalase (400 micrograms/ml) could significantly reduce the PQ-stimulated increase of phorbol ester binding and particular PKC phosphorylating activity, but dimethylsulfoxide (DMSO, 1.5%), an effective .OH trapping agent, failed to prevent this stimulation. In addition, an endogenous substrate of PKC, 80 kDa protein, was found to be highly phosphorylated in intact WI-38 cells treated with 50 microM PQ. The increase of phosphorylated proteins could be completely or partly abolished by Tempol or catalase, but only the phosphorylation of 80 kDa protein was diminished by protein kinase C inhibitor, 1-(5-isoquinolinyl-sulfonyl)-2-methylpiperazine (H-7). A maximal peak of ODC activity was observed at 6 h of treatment with 50 microM PQ. PQ induced activity was reduced at the following rates, Tempol 85%, DMSO 80% and catalase 45%, but H-7 failed to do so. Furthermore, we found that the level of c-jun mRNA was transiently increased by PQ and the peak appeared at 1 h of treatment. When correlated with the PKC result, Tempol, catalase and H-7 all effectively blocked PQ-elicited c-jun transcript expression, but DMSO only exhibited a weakly inhibitory effect. We therefore propose that superoxide anion (O2- and H2O2 generated by PQ could activate PKC and lead to induction of c-jun gene expression; on the other hand, O2- and .OH might trigger other kinase pathways to elevate ODC activity. Finally, the sequential expression of c-jun oncogene and ODC may cooperate to relieve the oxidative damages elicited by PQ.
Kuo
M L
ML
Institute of Toxicology, college of Medicine National Taiwan University, Taipei, Republic of China.
Lee
K C
KC
Lin
J K
JK
Huang
T S
TS
D002460:Cell Line; D004789:Enzyme Activation; D015870:Gene Expression / Q000187:drug effects; D016758:Genes, jun; D006801:Humans; D007700:Kinetics; D008168:Lung / Q000187:drug effects* / Q000378:metabolism; D009955:Ornithine Decarboxylase / Q000378:metabolism*; D010269:Paraquat / Q000633:toxicity*; D011493:Protein Kinase C / Q000378:metabolism*; D000090063:Proto-Oncogene Mas; D017382:Reactive Oxygen Species / Q000378:metabolism*
C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; D017382:Reactive Oxygen Species; D011493:Protein Kinase C; D009955:Ornithine Decarboxylase; D010269:Paraquat