Caspases are the main executioners of Fas-mediated apoptosis, irrespective of the ceramide signalling pathway.
Tumor necrosis factor alpha (TNF) or cytotoxic anti-Fas antibodies lead to the activation of apoptotic proteases (caspases) and to sphingomyelinase-mediated ceramide generation. Caspases and ceramide are both known to induce apoptosis on its own, but their relative contribution to Fas- and TNF-induced cell death is not well established. We report here that rapid apoptosis induced by TNF in U937 cells or anti-Fas in Jurkat cells, in the presence of cycloheximide, induced only a very low increase (<20%) in the cell ceramide content. Neither treatment with inhibitors of sphingomyelinases nor incubation of cells with fumonisin B1, which inhibits de novo ceramide synthesis, prevented TNF and Fas-mediated apoptosis. Increasing or depleting the cell ceramide content by prolonged culture in the presence of monensin or fumonisin B1, respectively, did not prevent TNF and Fas-mediated apoptosis. Treatment of cells with sphingomyelinase inhibitors did not affect to the activation of CPP32 (caspase-3) induced by TNF or anti-Fas antibodies. Chromatin condensation and fragmentation in cells treated with anti-Fas or TNF was abrogated by peptide inhibitors of caspases, which also inhibited Fas-, but not TNF-induced cell death. These results indicate that while ceramide does not seem to act as a critical mediator of TNF and Fas-induced apoptosis, it is generated as a consequence of CPP32 activation and could contribute to the spread of the intracellular death signal.
Gamen
S
S
Departamento de Bioquímica y Biología Molecular y Celular, Facultad de Ciencias, Universidad de Zaragoza, 50009 Zaragoza, Spain.
Anel
A
A
Piñeiro
A
A
Naval
J
J
D017209:Apoptosis / Q000187:drug effects / Q000276:immunology* / Q000502:physiology*; D002264:Carboxylic Acids / Q000494:pharmacology; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism / Q000502:physiology*; D002518:Ceramides / Q000502:physiology*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D037341:Fumonisins; D006801:Humans; D019169:Jurkat Cells; D008985:Monensin / Q000494:pharmacology; D015398:Signal Transduction; D013108:Sphingomyelin Phosphodiesterase / Q000037:antagonists & inhibitors; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D020298:U937 Cells; D019014:fas Receptor / Q000502:physiology*
D002264:Carboxylic Acids; D002518:Ceramides; D015853:Cysteine Proteinase Inhibitors; D004791:Enzyme Inhibitors; D037341:Fumonisins; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor; C056933:fumonisin B1; D008985:Monensin; D013108:Sphingomyelin Phosphodiesterase; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases