title: Bcl-xL interrupts oxidative activation of neutral sphingomyelinase.
abstract: Recent studies demonstrate a role for intracellular oxidation in the regulation of neutral sphingomyelinase (N-SMase). Glutathione (GSH) has been shown to regulate N-SMase in vitro and in cells. However, it has not been established whether the effects of GSH in cells are due to direct action on N-SMase. In this study, treatment of human mammary carcinoma MCF-7 cells with diamide, a thiol-depleting agent, caused a decrease in intracellular GSH and degradation of sphingomyelin (SM) to ceramide. The SM pool hydrolyzed in response to diamide belonged to the bacterial SMase-resistant pool of SM. Importantly, pretreatment of MCF-7 cells with GSH, N-acetylcysteine, an antioxidant, or GW69A, a specific N-SMase inhibitor, prevented diamide-induced degradation of SM to ceramide, suggesting that intracellular levels of GSH regulate the extent to which SM is degraded to ceramide and that this probably involves a GW69A-sensitive N-SMase. Unexpectedly, expression of Bcl-xL prevented tumor necrosis factor-alpha-induced SM hydrolysis and ceramide accumulation but not the decrease in intracellular GSH. Furthermore, Bcl-xL inhibited diamide-induced SM hydrolysis and ceramide accumulation but not the decrease in intracellular GSH. These results suggest that the site of action of Bcl-xL is downstream of GSH depletion and upstream of ceramide accumulation, and that GSH probably does not exert direct physiologic effects on N-SMase.
authors.affiliation: Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, 173 Ashley Ave., 29425, USA.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Okamoto
authors.forename: Lina M
authors.initials: LM
authors.lastname: Obeid
authors.forename: Yusuf A
authors.initials: YA
authors.lastname: Hannun
mesh_terms: D003958:Diamide / Q000494:pharmacology; D004789:Enzyme Activation; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D010084:Oxidation-Reduction; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000502:physiology*; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism*; D013109:Sphingomyelins / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D051020:bcl-X Protein
chemical_list: C494811:BCL2L1 protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; D013109:Sphingomyelins; D014409:Tumor Necrosis Factor-alpha; D051020:bcl-X Protein; D003958:Diamide; D013108:Sphingomyelin Phosphodiesterase; D005978:Glutathione