Involvement of caspase 3-activated DNase in internucleosomal DNA cleavage induced by diverse apoptotic stimuli.
Degradation of chromosomal DNA into nucleosome-sized fragments is one of the characteristics of apoptotic cell death. Here, we examined whether caspase-activated DNase (CAD) is responsible for the DNA fragmentation that occurs upon exposure to various apoptotic stimuli. When human Jurkat cells were exposed to etoposide, or UV or gamma radiation, a caspase-3-like protease was activated, and nuclear DNA was fragmented. Human TF-1 cells, which are dependent on granulocyte-macrophage colony-stimulating factor (GM-CSF), also underwent apoptosis accompanied by the activation of caspase-3-like protease and DNA fragmentation, when cultured without the cytokine. Both Jurkat and TF-1 cells expressed two forms of ICAD, ICAD-L and ICAD-S, which were cleaved upon exposure to these apoptotic stimuli. Among eight different caspases examined, recombinant caspases 3 and 7 specifically cleaved ICAD synthesized in a cell-free system. An expression plasmid containing mouse ICAD-L mutated at the caspase-3-recognition sites was then introduced into Jurkat and TF-1 cells. When the transformants were induced to undergo apoptosis (by treatment with etoposide, UV or gamma radiation for Jurkat cells, or factor withdrawal for TF-1 cells) they did not show DNA fragmentation, although they still died as a result of these stimuli. These results indicated that CAD, released from ICAD by caspase activation, is involved in the nuclear DNA fragmentation induced by these apoptotic stimuli.
McIlroy
D
D
Department of Genetics, Osaka University Medical School, Suita, Japan.
Sakahira
H
H
Talanian
R V
RV
Nagata
S
S
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000528:radiation effects; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D002460:Cell Line; D004247:DNA / Q000378:metabolism*; D053938:DNA Fragmentation; D003851:Deoxyribonucleases / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects / Q000528:radiation effects; D005047:Etoposide / Q000633:toxicity; D005720:Gamma Rays; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D019169:Jurkat Cells; D007700:Kinetics; D051379:Mice; D009707:Nucleosomes / Q000187:drug effects / Q000378:metabolism*; D010957:Plasmids; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D014466:Ultraviolet Rays
D007527:Isoenzymes; D009707:Nucleosomes; D011994:Recombinant Proteins; D005047:Etoposide; D004247:DNA; D003851:Deoxyribonucleases; C109663:caspase-activated deoxyribonuclease; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases