title: Selenocompounds induce a redox modulation of protein kinase C in the cell, compartmentally independent from cytosolic glutathione: its role in inhibition of tumor promotion.
abstract: Since selenite and other redox-active selenocompounds can modify protein kinase C (PKC) in the test tube, we have determined whether or not this redox regulation occurs inside the cell despite having high concentrations of GSH and the role of this regulation in the inhibition of tumor promotion. By using phorbol ester-promoted JB6 epidermal cell transformation assay, the concentrations of selenite, selenocystine, and selenodiglutathione which are optimal for chemopreventive activity were determined. At such concentrations (0.5 to 2 microM) in the cells treated with these agents, only a slight but transient decrease in PKC activity was observed when measured with a low (5 microM), but not with a high (100 microM) concentration of ATP. However, when the cells were serum starved or pretreated with 2-deoxyglucose, there was a pronounced but transient inactivation of PKC when assayed with both low and high concentrations of ATP. The inactivation was reversed in the cell by an endogenous mechanism or by treatment with thiol agents in the test tube. In spite of a substantial (90%) depletion of GSH in the cells by pretreatment with buthionine sulfoximine, there was no further increase in the redox modification of PKC by selenite as well as no change in the inhibitory effect of selenite on the phorbol ester-stimulated induction of ornithine decarboxylase, which is an intermediate marker related to cell transformation. While GSH is known to influence certain actions of selenium, it may not be required to mediate the effects of selenite tested in this study. The water-soluble cytosolic GSH did not interfere with the redox modification of PKC probably due to the shielding of the cysteine-rich region of the enzyme by a weak hydrophobic association with the membrane. Due to the presence of cofactors in the crude cell extracts, PKC was more sensitive to selenite than in the purified form and was inactivated by low concentrations of selenite (IC50 = 0.05 microM). This modification was reversed by thiol agents as well as by NADPH. A protein disulfide reductase, which can regenerate PKC, was present in the homogenate. Conceivably, selenite and other selenocompounds induce a redox modification of cellular PKC, compartmentally independent from the cytosolic GSH, but intimately connected to a NADPH-dependent reductase system, to mediate, at least in part, some of the cancer-preventive actions.
authors.affiliation: Department of Cell and Neurobiology, School of Medicine, University of Southern California, Los Angeles 90033, USA. rgopalak@hsc.usc.edu
authors.forename: R
authors.initials: R
authors.lastname: Gopalakrishna
authors.forename: Z H
authors.initials: ZH
authors.lastname: Chen
authors.forename: U
authors.initials: U
authors.lastname: Gundimeda
mesh_terms: D000818:Animals; D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D002273:Carcinogens / Q000633:toxicity; D002460:Cell Line; D002471:Cell Transformation, Neoplastic / Q000187:drug effects*; D002848:Chromatography, DEAE-Cellulose; D003553:Cystine / Q000031:analogs & derivatives / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D005978:Glutathione / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D019803:Glutathione Disulfide / Q000378:metabolism; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D016566:Organoselenium Compounds / Q000494:pharmacology*; D010084:Oxidation-Reduction; D015240:Phorbol 12,13-Dibutyrate / Q000633:toxicity; D011493:Protein Kinase C / Q000302:isolation & purification / Q000378:metabolism*; D018038:Sodium Selenite / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000633:toxicity; D014407:Tumor Cells, Cultured
chemical_list: D016588:Anticarcinogenic Agents; D002273:Carcinogens; D007527:Isoenzymes; D016566:Organoselenium Compounds; C009226:selenocystine; C017710:selenodiglutathione; D015240:Phorbol 12,13-Dibutyrate; D003553:Cystine; D011493:Protein Kinase C; D005978:Glutathione; D018038:Sodium Selenite; D013755:Tetradecanoylphorbol Acetate; D019803:Glutathione Disulfide