INTERLEUKIN 1-BETA-CONVERTING ENZYME RELATED PROTEASES CASPASES ARE INVOLVED IN TRAIL-INDUCED APOPTOSIS OF MYELOMA AND LEUKEMIA-CELLS/

The Fas/APO-1/CD95 ligand (CD95L) and the recently cloned TRAIL ligand belong to the TNF-family and share the ability to induce apoptosis in sensitive target cells, Little information is available on the degree of functional redundancy between these two ligands in terms of target selectivity and intracellular signalling pathway(s). To address these issues, we have expressed and characterized recombinant mouse TRAIL. Specific detection with newly developed rabbit anti-TRAIL antibodies s howed that the functional TRAIL molecule released into the supernatant of recombinant baculovirus-infected Sf9 cells is very similar to that associated with the membrane fraction of Sf9 cells. CD95L resistant m yeloma cells were found to be sensitive to TRAIL, displaying apoptotic features similar to those of the CD95L- and TRAIL-sensitive T leukemi a cells Jurkat. To assess if IL-1 beta-converting enzyme (ICE) and/or ICE-related proteases (IRPs) (caspases) are involved in TRAIL-induced apoptosis of both cell types, peptide inhibition experiments were perf ormed. The irreversible IRP/caspase-inhibitor Ac-YVAD-cmk and the reve rsible IRP/caspase-inhibitor Ac-DEVD-CHO blocked the morphological cha nges, disorganization of plasma membrane phospholipids, DNA fragmentat ion, and loss of cell viability associated with TRAIL-induced apoptosi s. In addition, cells undergoing TRAIL-mediated apoptosis displayed cl eavage of poly(ADP)-ribose polymerase (PARP) that was completely block ed by Ac-DEVD-CHO. These results indicate that TRAIL seems to compleme nt the activity of the CD95 system as it allows cells, otherwise resis tant, to undergo apoptosis triggered by specific extracellular ligands . Conversely, however, induction of apoptosis in sensitive cells by TR AIL involves IRPs/caspases in a fashion similar to CD95L. Thus, differ ential sensitivity to CD95L and TRAIL seems to map to the proximal sig naling events associated with receptor triggering.