¹Department of Genetics, Osaka University Medical School, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan

²BASF Bioresearch Corporation, Worcester, Massachusetts, MA 01605, USA

Correspondence to: Shigekazu Nagata, Department of Genetics, Osaka University Medical School, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan

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 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 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.

apoptosis; caspase; DNase; anti-cancer drugs