title: Redox-regulation of Erk1/2-directed phosphatase by reactive oxygen species: role in signaling TPA-induced growth arrest in ML-1 cells.
abstract: Extracellular signal-regulated kinase (Erk)1/2 activity signals myeloid cell differentiation induced by 12-O-tetradecanoyl-phorbol-13-acetate (TPA). Previously, we reported that Erk1/2 activation (phosphorylation) induced by TPA required reactive oxygen species (ROS) as a second messenger. Here, we hypothesized that ROS generated in response to TPA inhibit Erk1/2-directed phosphatase activity, which leads to an increase phosphorylation of Erk1/2 to signal p21(WAF1/Cip1)-mediated growth arrest in ML-1 cells. Incubation of ML-1 cells with TPA resulted in a marked accumulation of phosphorylated Erk1/2, and is subsequent to H2O2 generation. Interestingly, post-TPA-treatment with N-acetylcysteine (NAC) stimulated a marked and a rapid dephosphorylation of Erk1/2, suggesting a regeneration of Erk1/2-directed phospahatase activity by NAC. ROS generation in ML-1 cells induced by TPA was suggested to occur in the mitochondrial electron transport chain (METC) based on the following observations: (i) undifferentiated ML-1 cells not only lack p67-phox and but also express a low level of p47-phox key components required for NADPH oxidase enzymatic activity, (ii) pretreatment with DPI, an inhibitor of NADH- and NADPH-dependent enzymes, or rhein, an inhibitor of complex I, blocked the ROS generation, and (iii) examination of the microarray analysis data and Western blot analysis data revealed an induction of MnSOD expression at both mRNA and protein levels in response to TPA. MnSOD is a key member of the mitochondrial defense system against mitochondrial-derived superoxide. Together, this study suggested that TPA stimulated ROS generation as a second messenger to activate Erk1/2 via a redox-mediated inhibition of Erk1/2-directed phosphatase in ML-1 cells.
authors.affiliation: Department of Environmental Health Sciences, Johns Hopkins Medical Institutions, Baltimore, Maryland 21205, USA. ktraore@jhsph.edu
authors.forename: Kassim
authors.initials: K
authors.lastname: Traore
authors.forename: Rajni
authors.initials: R
authors.lastname: Sharma
authors.forename: Rajesh K
authors.initials: RK
authors.lastname: Thimmulappa
authors.forename: Walter H
authors.initials: WH
authors.lastname: Watson
authors.forename: Shyam
authors.initials: S
authors.lastname: Biswal
authors.forename: Michael A
authors.initials: MA
authors.lastname: Trush
mesh_terms: D002273:Carcinogens / Q000378:metabolism; D002460:Cell Line; D049109:Cell Proliferation; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D004789:Enzyme Activation; D004790:Enzyme Induction; D004791:Enzyme Inhibitors / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D020869:Gene Expression Profiling; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D016877:Oxidants / Q000378:metabolism; D010084:Oxidation-Reduction; D010617:Phenanthridines / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D013482:Superoxide Dismutase / Q000235:genetics / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000378:metabolism*
chemical_list: D002273:Carcinogens; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D016877:Oxidants; D010617:Phenanthridines; D010750:Phosphoproteins; D017382:Reactive Oxygen Species; C079789:neutrophil cytosol factor 67K; C058428:hydroethidine; D006861:Hydrogen Peroxide; D013482:Superoxide Dismutase; D011493:Protein Kinase C; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D010749:Phosphoprotein Phosphatases; D013755:Tetradecanoylphorbol Acetate