Caspase-3 Is a Component of Fas Death-Inducing Signaling Complex in Lipid Rafts and Its Activity Is Required for Complete Caspase-8 Activation during Fas-Mediated Cell Death

• Published in: The Journal of immunology (1950) Vol. 172; no. 4; pp. 2316 - 2323
• Main Authors: Aouad, Salah M, Cohen, Luchino Y, Sharif-Askari, Ehsan, Haddad, Elias K, Alam, Antoine, Sekaly, Rafick-Pierre
• Format: Journal Article
• Language: English
• Published: United States Am Assoc Immnol 15-02-2004
• Subjects: Apoptosis - immunology; Caspase 3; Caspase 8; Caspases - biosynthesis; Caspases - metabolism; Caspases - physiology; Death Domain Receptor Signaling Adaptor Proteins; death receptors; Enzyme Activation - immunology; Fas antigen; Fas Ligand Protein; fas Receptor - immunology; fas Receptor - metabolism; fas Receptor - physiology; Humans; Hydrolysis; Intracellular Membranes - enzymology; Intracellular Membranes - immunology; Jurkat Cells; Ligands; Membrane Glycoproteins - metabolism; Membrane Glycoproteins - physiology; Membrane Microdomains - enzymology; Membrane Microdomains - physiology; Membrane Potentials - immunology; Mitochondria - enzymology; Mitochondria - immunology; Protein Processing, Post-Translational - immunology; Receptors, Tumor Necrosis Factor - metabolism; Receptors, Tumor Necrosis Factor - physiology; Signal Transduction - immunology; T-Lymphocytes - cytology; T-Lymphocytes - enzymology; T-Lymphocytes - immunology
• ISSN: 0022-1767; 1550-6606
• DOI: 10.4049/jimmunol.172.4.2316

Since its discovery, caspase-8 has been placed at the apex of the proteolytic cascade triggered by death receptor (DR) cross-linking. Because of its capacity to interact with the cytoplasmic portion of DR, it has been suggested that caspase-8 acts independently of other caspases in the initiation of Fas and other DR signaling. In this study, we demonstrate that in Jurkat cells, caspase-3 cleavage is an early step during Fas-induced apoptosis. We show that caspase-3 processing into its p20 occurs rapidly after Fas cross-linking, in the absence of mitochondrial depolarization and caspase-9 activation. Moreover, caspase-3 is present in lipid rafts of untreated Jurkat cells and peripheral T lymphocytes. Caspase-3, caspase-8, and Fas-associated death domain are further recruited to lipid rafts of Jurkat cells following anti-Fas treatment. Fas immunoprecipitation reveals that caspase-3 is a component of the death-inducing signaling complex, suggesting that this cysteine protease is in close proximity to caspase-8. Furthermore, transduction of Jurkat cells with a caspase-3 dominant-negative form inhibits caspase-8 processing and results in inhibition of apoptosis, suggesting that caspase-3 activity is required for caspase-8 activation. Overall, these findings support a model whereby caspase-3 is a component of the death-inducing signaling complex located in lipid rafts, and as such, is involved in the amplification of caspase-8 activity by the mitochondrion.