title: PKC downstream of Pl3-kinase regulates peroxynitrite formation for Nrf2-mediated GSTA2 induction.
abstract: The protective adaptive response to electrophiles and reactive oxygen species is mediated by the induction of phase II detoxifying genes including glutathione S-transferases (GSTs). NF-E2-related factor-2 (Nrf2) phosphorylation by protein kinase C (PKC) is a critical event for its nuclear translocation in response to oxidative stress. Previously, we have shown that peroxynitrite plays a role in activation of Nrf2 and Nrf2 binding to the antioxidant response element (ARE) via the pathway of phosphatidylinositol 3-kinase (Pl3-kinase) and that nitric oxide synthase in hepatocytes is required for GSTA2 induction. In view of the importance of PKC and Pl3-kinase in Nrf2-mediated GST induction, we investigated the role of these kinases in peroxynitrite formation for GSTA2 induction by oxidative stress and determined the relationship between PKC and Pl3-kinase. Although PKC activation by phorbol 12-myristate-13-acetate (PMA) did not increase the extents of constitutive and inducible GSTA2 expression, either PKC depletion by PMA or PKC inhibition by staurosporine significantly inhibited GSTA2 induction by tert-butylhydroquinone (t-BHQ) a prooxidant chemical. Therefore, the basal PKC activity is requisite for GSTA2 induction. 3-Morpholinosydnonimine (SIN-1), which decomposes and yields peroxynitrite, induced GSTA2, which was not inhibited by PKC depletion, but slightly enhanced by PKC activation, suggesting that PKC promotes peroxynitrite formation for Nrf2-mediated GSTA2 induction. Treatment of cells with S-nitroso-N-acetyl-penicillamine (SNAP), an exogenous NO donor, in combination with t-BHQ may produce peroxynitrite. GSTA2 induction by SNAP + t-BHQ was not decreased by PKC depletion, but rather enhanced by PKC activation, showing that the activity of PKC might be required for peroxynitrite formation. LY294002 a Pl3-kinase inhibitor blocked GSTA2 induction by t-BHQ, which was reversed by PMA-induced PKC activation. These results provide evidence that PKC may play a role in formation of peroxynitrite that activates Nrf2 for GSTA2 induction and that PKC may serve an activator for GSTA2 induction downstream of Pl3-kinase.
authors.affiliation: College of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Seoul 151-742, Korea. sgk@snu.ac.kr
authors.forename: Sang Geon
authors.initials: SG
authors.lastname: Kim
authors.forename: Sun Ok
authors.initials: SO
authors.lastname: Kim
mesh_terms: D000818:Animals; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000502:physiology*; D004789:Enzyme Activation / Q000187:drug effects; D005982:Glutathione Transferase / Q000096:biosynthesis*; D022781:Hepatocytes / Q000378:metabolism; D006873:Hydroquinones / Q000494:pharmacology; D015151:Immunoblotting; D008981:Molsidomine / Q000031:analogs & derivatives* / Q000494:pharmacology; D009025:Morpholines / Q000494:pharmacology; D051267:NF-E2-Related Factor 2; D030421:Peroxynitrous Acid / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D015534:Trans-Activators / Q000502:physiology*
chemical_list: D002867:Chromones; D004268:DNA-Binding Proteins; D006873:Hydroquinones; D009025:Morpholines; D051267:NF-E2-Related Factor 2; C495637:Nfe2l2 protein, rat; D017382:Reactive Oxygen Species; D015534:Trans-Activators; D030421:Peroxynitrous Acid; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C002385:linsidomine; C018855:2-tert-butylhydroquinone; D008981:Molsidomine; D005982:Glutathione Transferase; D019869:Phosphatidylinositol 3-Kinases; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate