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Apoptosis is orchestrated by a family of cysteine proteases known as the caspases. Fourteen mammalian caspases have been identified, three of which (caspase-3, -6, and -7) are thought to coordinate the execution phase of apoptosis by cleaving multiple structural and repair proteins. However, the relative contributions that the "executioner" caspases make to the demolition of the cell remains speculative. Here we have used cell-free extracts immuno-depleted of either caspase-3, -6, or -7 to examine the caspase requirements for apoptosis-associated proteolysis of 14 caspase substrates as well as nuclear condensation, chromatin margination, and DNA fragmentation. We show that caspase-3 is the primary executioner caspase in this system, necessary for cytochrome c/dATP-inducible cleavage of fodrin, gelsolin, U1 small nuclear ribonucleoprotein, DNA fragmentation factor 45 (DFF45)/inhibitor of caspase-activated DNase (ICAD), receptor-interacting protein (RIP), X-linked inhibitor of apoptosis protein (X-IAP), signal transducer and activator of transcription-1 (STAT1), topoisomerase I, vimentin, Rb, and lamin B but not for cleavage of poly(ADP-ribose) polymerase (PARP) or lamin A. In addition, caspase-3 was also essential for apoptosis-associated chromatin margination, DNA fragmentation, and nuclear collapse in this system. Surprisingly, although caspase-6 and -7 are considered to be important downstream effector caspases, depletion of either caspase had minimal impact on any of the parameters investigated, calling into question their precise role during the execution phase of apoptosis.

Original publication




Journal article


The Journal of biological chemistry

Publication Date





7320 - 7326


Molecular Cell Biology Laboratory, Department of Genetics, The Smurfit Institute, Trinity College, Dublin 2, Ireland.


Jurkat Cells, Cell Nucleus, Chromatin, Cell-Free System, Animals, Humans, Vimentin, Microfilament Proteins, Cytochrome c Group, Caspases, DNA Topoisomerases, Type I, Proteins, Carrier Proteins, Gelsolin, Ribonucleoprotein, U1 Small Nuclear, DNA-Binding Proteins, Retinoblastoma Protein, Trans-Activators, Nuclear Proteins, Lamins, Lamin Type A, Lamin Type B, Adenosine Triphosphate, Apoptosis, DNA Fragmentation, Models, Biological, Time Factors, STAT1 Transcription Factor, Apoptosis Regulatory Proteins, X-Linked Inhibitor of Apoptosis Protein, Caspase 3, Caspase 6, Caspase 7, Receptor-Interacting Protein Serine-Threonine Kinases