Table; caspase-1 ,-2 ,-3 ,-4 ,-5 ,-6 ,-7 ,-8 ,-9 ,-10



By use of a PCR approach that uses degenerate oligonucleotides encoding the highly conserved pentapeptides QACRG and GSWFI that are present in all known caspases, a novel apoptotic gene was cloned from human Jurkat T lymphocytes. This new gene encodes an approximately 34-kDa protein that is highly homologous to human caspase-3, the Caenorhabditis elegans cell death protein CED-3, caspase-2 and caspase-1. Because of its high homology to the C. elegans ced-3 gene, this new gene was first named "mammalian Ced-3 homologue - Mch2". Two Mch2 transcripts (Mch2 alpha, 1.7 kb; Mch2 beta, 1.4 kb) were detected in Jurkat T lymphocytes and other cell lines. Mch2 alpha transcript encodes the full-length Mch2, whereas the Mch2 beta transcript encodes a shorter Mch2 isoform, probably as a result of alternative splicing. Like caspase-1 and -3, recombinant Mch2 alpha, but not Mch2 beta, possesses protease activity, as determined by its ability to cleave the fluorogenic peptide DEVD-AMC. Caspase-3 and Mch2 alpha can also cleave poly(ADP-ribose) polymerase in vitro, suggesting that these enzymes participate in poly(ADP-ribose) polymerase cleavage observed during cellular apoptosis. In addition, overexpression of recombinant Mch2 alpha, but not Mch2 beta, induces apoptosis in Sf9 insect cells (Fernandes Alnemri et al., 1995).


Activation of pro-caspase-6 by caspase-3 results in an active enzyme that is capable of cleaving an artificially introduced lamin cleavage site (VEID¯ N) (Srinivasula et al., 1996). These data suggest that caspase-3 is activated prior to caspase-6, and may be responsible for, the activation of caspase-6. However, these results are in apparent conflict with a study which identified and purified from hamster liver a homologue of caspase-6 that is capable of activating caspase-3 (Liu et al., 1996). In addition, caspase-6 processes pro-caspase-3 at the I172ETD¯ S176 site between the large and small subunits, and this cleavage is blocked when Asp-175 is mutated to Ala (Srinivasula et al., 1996). Thus activation of caspase-3 can result in activation of pro-caspase-6 but, similarly, activation of pro-caspase-6 can also result in activation of caspase-3; resulting in a protease amplification cycle (Srinivasula et al., 1996).


Some caution must be exercised in the interpretation of such in vitro experiments, as purified or partially purified proteases may cleave substrates that they do not cleave in vivo. Either because the ratio of enzyme to substrate would never reach such a level in the cell or because of so-called space/time constraints meaning that the enzyme may be located in a different subcellular environment and/or is expressed at a different time point than the substrate.