Protease activation in apoptosis induced by MAL.
The proteolytic caspase cascade plays a central role in the signaling and execution steps of apoptosis. This study investigated the activation of different caspases in apoptosis induced by MAL (a folding variant of human alpha-lactalbumin) isolated from human milk. Our results show that the caspase-3-like enzymes, and to a lesser extent the caspase-6-like enzymes, were activated in Jurkat and A549 cells exposed to MAL. Activated caspases subsequently cleaved several protein substrates, including PARP, lamin B, and alpha-fodrin. A broad-range caspase inhibitor, zVAD-fmk, blocked the caspase activation, the cleavage of proteins, and DNA fragmentation, indicating an important role for caspase activation in MAL-induced apoptosis. Since an antagonistic anti-CD95 receptor antibody, ZB4, did not influence the MAL-induced killing, we conclude that this process does not involve the CD95-mediated pathway. While MAL did not directly activate caspases in the cytosol, it colocalized with mitochondria and induced the release of cytochrome c. Thus, these results demonstrate that caspases are activated and involved in apoptosis induced by MAL and that direct interaction of MAL with mitochondria leads to the release of cytochrome c, suggesting that this release is an important step in the initiation and/or amplification of the caspase cascade in these cells.
Köhler
C
C
Institute of Environmental Medicine, Division of Toxicology, Karolinska Institutet, Stockholm, S-171 77, Sweden.
Håkansson
A
A
Svanborg
C
C
Orrenius
S
S
Zhivotovsky
B
B
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D053178:Caspase 6; D020169:Caspases / Q000378:metabolism; D002457:Cell Extracts / Q000737:chemistry; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism; D003593:Cytoplasm / Q000201:enzymology; D010450:Endopeptidases / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D005260:Female; D006801:Humans; D019169:Jurkat Cells; D007768:Lactalbumin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D000590:Amino Acid Chloromethyl Ketones; D002457:Cell Extracts; D015853:Cysteine Proteinase Inhibitors; D003574:Cytochrome c Group; D004792:Enzyme Precursors; D016326:Extracellular Matrix Proteins; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D007768:Lactalbumin; D010450:Endopeptidases; C505407:CASP3 protein, human; C505514:CASP6 protein, human; D053148:Caspase 3; D053178:Caspase 6; D020169:Caspases