A serine protease is involved in the initiation of DNA damage-induced apoptosis.
Caspases are considered to be the key effector proteases of apoptosis. Initiator caspases cleave and activate downstream executioner caspases, which are responsible for the degradation of numerous cellular substrates. We studied the role of caspases in apoptotic cell death of a human melanoma cell line. Surprisingly, the pancaspase inhibitor zVAD-fmk was unable to block cleavage of poly(ADP-ribose) polymerase (PARP) after treatment with etoposide, while it did prevent DEVDase activity. It is highly unlikely that caspase-2, which is a relatively zVAD-fmk-resistant caspase, is mediating etoposide-induced PARP cleavage, as a preferred inhibitor of this caspase could not prevent cleavage. In contrast, caspase activation and PARP degradation were blocked by pretreatment of the cells with the serine protease inhibitor 4-(2-aminoethyl)benzenesulfonyl fluoride (AEBSF). We therefore conclude that a serine protease regulates an alternative initiation mechanism that leads to caspase activation and PARP cleavage. More importantly, while zVAD-fmk could not rescue melanoma cells from etoposide-induced death, the combination with AEBSF resulted in substantial protection. This indicates that this novel pathway fulfills a critical role in the execution of etoposide-induced programmed cell death.
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics*; D015153:Blotting, Western; D053143:Caspase 2; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D045744:Cell Line, Tumor / Q000187:drug effects / Q000528:radiation effects; D003374:Coumarins / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004249:DNA Damage; D005047:Etoposide / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D008858:Microscopy, Phase-Contrast; D009842:Oligopeptides / Q000378:metabolism / Q000494:pharmacology; D010447:Peptide Hydrolases / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D051381:Rats; D012697:Serine Endopeptidases / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D013450:Sulfones / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D003374:Coumarins; D015853:Cysteine Proteinase Inhibitors; D009842:Oligopeptides; D015842:Serine Proteinase Inhibitors; D013450:Sulfones; D014409:Tumor Necrosis Factor-alpha; C115495:aspartyl-glutamyl-valyl-aspartyl-7-amino-4-trifluoromethylcoumarin; C427308:benzoylcarbonyl-valyl-aspartyl-valyl-alanyl-aspartyl-fluoromethyl ketone; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C002010:4-(2-aminoethyl)benzenesulfonylfluoride; D005047:Etoposide; D011065:Poly(ADP-ribose) Polymerases; D010447:Peptide Hydrolases; D012697:Serine Endopeptidases; C505407:CASP3 protein, human; C505410:Casp3 protein, rat; D053143:Caspase 2; D053148:Caspase 3; D020169:Caspases; C108711:DEVDase