ABSTRACT
Menadione (2-methyl-1,4-naphthoquinone) has been used extensively as an oxidant stressor at the cellular level. However, the mechanism of cytotoxicity of this compound still remains controversial. This study deals with the role of intracellular glutathione in the resistance of the yeast Saccharomyces cerevisiae to menadione. Incubation with 0.5 mM menadione resulted in a decrease of total glutathione concentration in yeast cells, intracellular formation of menadione S-glutathione conjugate and export of the conjugate from cells. GSH-deficient mutants showed lower stimulation of superoxide and hydrogen peroxide production upon exposure to menadione and were more resistant to menadione than wild-type isogenic strains. These results indicate that in yeast cells the formation of S-glutathione conjugate is a major pathway of menadione metabolism and that this reaction leads to redox activation of menadione but permits its removal from the cells.
Subject(s)
Glutathione/metabolism , Hemostatics/pharmacokinetics , Hemostatics/toxicity , Saccharomyces cerevisiae/drug effects , Saccharomyces cerevisiae/metabolism , Vitamin K/pharmacokinetics , Vitamin K/toxicity , Biotransformation , Disulfides/metabolism , Hydrogen Peroxide/metabolism , Oxidation-Reduction , Oxidative Stress , Sensitivity and Specificity , Superoxides/metabolismABSTRACT
A hypothesis of the flippase nature of the glutathione S-conjugate transport is presented. Experimental premises for this hypothesis include interaction of glutathione S-conjugates with the membrane, as demonstrated by their effects on membrane fluidity, quenching of 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene fluorescence and induction of echinocytosis by 2,4-dinitrophenyl-S-glutathione (DNP-SG). This hypothesis can rationalize, i. a., observations of the enhancement of DNP-SG transport by butanol and stimulation of erythrocyte membrane Mg(2+)-ATPase activity by albumin-coupled DNP-SG.
Subject(s)
Carrier Proteins/blood , Membrane Proteins/blood , Phospholipid Transfer Proteins , 1-Butanol/pharmacology , Biological Transport/drug effects , Ca(2+) Mg(2+)-ATPase/blood , Cell Size/drug effects , Dinitrophenols/blood , Dinitrophenols/pharmacology , Erythrocyte Membrane/enzymology , Glutathione/analogs & derivatives , Glutathione/blood , Glutathione/pharmacology , Humans , Membrane Transport Proteins , Serum Albumin, Bovine/pharmacologyABSTRACT
The content of reactive carbonyls and of glutathione-protein mixed disulfides, two indices of oxidative stress, were compared in wild-type Saccharomyces cerevisiae and in strains deficient in superoxide dismutase and catalase, and of decreased glutathione level. Both indices were higher in stationary than in logarithmic cultures and were not increased in antioxidant-deficient strains. Oxidation of dichlorofluorescin, an estimate of peroxide production, measured in the presence of exogenous peroxidase, was higher in antioxidant-deficient strains. These results corroborate our previous results on compensatory antioxidant mechanisms in the mutant yeast strains.
Subject(s)
Acatalasia , Antioxidants/metabolism , Fungal Proteins/metabolism , Oxidative Stress/physiology , Saccharomyces cerevisiae/metabolism , Superoxide Dismutase/deficiency , Disulfides/metabolism , Glutathione/metabolism , Oxidation-Reduction , Saccharomyces cerevisiae/enzymologyABSTRACT
The effect of chosen compounds on the total glutathione (GSH) level in stationary cultures of S. cerevisiae was compared. 1-Chloro-2,4-dinitrobenzene, 1-fluoro-2,4-dinitrobenzene, maleimide, iodacetamide and allyl alcohol (1 mM), and menadione (0.5 mM) caused an almost complete GSH depletion during several minutes. Bromobenzoic acid and chloramine T (I mM), and daunomycin (60 mu M) induced a slower GSH decrease, down to 30-70% after 60 min. Paraquat (1 mM), CuSO(4) (0.5 mM) and cadmium acetate (1 mM) decreased glutathione level down to ca 70%. Diamide (0.5 mM), phenazine methosulphate, phenylhydrazine, acetylphenylhydrazine and H(2)O(2) (1 mM), and t-butyl hydroperoxide (2 mM) did not affect total GSH during 60-min exposure. There was no clear-cut dependence between the ability of various chemicals to deplete cellular GSH and their increased toxicity to a glutathione-poor mutant.
