Abstract
To identify essential components of the Fas-induced apoptotic signaling pathway, Jurkat T lymphocytes were chemically mutagenized and selected for clones that were resistant to Fas-induced apoptosis. We obtained five cell lines that contain mutations in the adaptor FADD. All five cell lines did not express FADD by immunoblot analysis and were completely resistant to Fas-induced death. Complementation of the FADD mutant cell lines with wild-type FADD restored Fas-mediated apoptosis. Fas activation of caspase-2, caspase-3, caspase-7, and caspase-8 and the proteolytic cleavage of substrates such as BID, protein kinase Cdelta, and poly(ADP-ribose) polymerase were completely defective in the FADD mutant cell lines. In addition, Fas activation of the stress kinases p38 and c-Jun NH2 kinase and the generation of ceramide in response to Fas ligation were blocked in the FADD mutant cell lines. These data indicate that FADD is essential for multiple signaling events downstream of Fas.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Adaptor Proteins, Signal Transducing*
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Calcium-Calmodulin-Dependent Protein Kinases / metabolism
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Carrier Proteins / genetics
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Carrier Proteins / metabolism*
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Carrier Proteins / physiology
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Caspase 8
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Caspase 9
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Caspases / metabolism
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Ceramides / biosynthesis
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Enzyme Activation
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Fas-Associated Death Domain Protein
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Genetic Complementation Test
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Humans
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JNK Mitogen-Activated Protein Kinases
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Jurkat Cells
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MAP Kinase Signaling System*
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Mitogen-Activated Protein Kinases / metabolism
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Mutagenesis
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fas Receptor / metabolism*
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p38 Mitogen-Activated Protein Kinases
Substances
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Adaptor Proteins, Signal Transducing
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Carrier Proteins
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Ceramides
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FADD protein, human
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Fas-Associated Death Domain Protein
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fas Receptor
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Calcium-Calmodulin-Dependent Protein Kinases
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JNK Mitogen-Activated Protein Kinases
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Mitogen-Activated Protein Kinases
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p38 Mitogen-Activated Protein Kinases
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CASP8 protein, human
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CASP9 protein, human
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Caspase 8
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Caspase 9
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Caspases