Premium
Mechanism of Acetaminophen‐Induced Apoptosis in Cultured Cells: Roles of Caspase‐3, DNA Fragmentation Factor, and the Ca 2+ and Mg 2+ Endonuclease DNAS1L3
Author(s) -
Boulares A. Hamid,
Ren Tao
Publication year - 2004
Publication title -
basic and clinical pharmacology and toxicology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.805
H-Index - 90
eISSN - 1742-7843
pISSN - 1742-7835
DOI - 10.1111/j.1742-7843.2004.pto_940105.x
Subject(s) - dna fragmentation , apoptotic dna fragmentation , microbiology and biotechnology , apoptosis , programmed cell death , hela , biology , poly adp ribose polymerase , acetaminophen , cytotoxic t cell , transfection , caspase , cell culture , chemistry , dna , polymerase , biochemistry , in vitro , genetics
We have recently shown that acetaminophen induces many of the apoptotic traits in hepatoma cells and lymphocytes (Boulares et al. (2002d). In an effort to further investigate the mechanism by which non‐metabolized acetaminophen induces apoptosis, we have now examined the roles of caspase‐3, the DNA fragmentation factor, and the poly(ADP‐ribose) polymerase‐1‐regulated Ca 2+ and Mg 2+ ‐dependent endonuclease DNAS1L3 in the induction of such death process. This was achieved with the use of MCF‐7 cells, a caspase‐3‐deficient breast adenocarcinoma cell line, thymocytes isolated from DFF45 (the inhibitory and chaperone subunit of the DNA fragmentation factor subunit, DFF40) deficient mice, and HeLa cells, a DNAS1L3‐deficient cervical carcinoma cell line. MCF‐7 exhibited a marked resistance to acetaminophen treatment. Ectopic expression of human caspase‐3 significantly potentiated the cytotoxic effect of acetaminophen and promoted the release of cytochrome c into the cytosol of treated cells suggesting a direct role for caspase‐3 in acetaminophen‐induced apoptosis. Expression and cleavage of DFF45 were required but not sufficient for acetaminophen‐induced internucleosomal DNA fragmentation. DFF45 gene knockout rendered thymocytes resistant against acetaminophen‐induced generation of both large and internucleosomal DNA fragments. The treatment of HeLa cells with acetaminophen resulted in internuclesomal DNA fragmentation only after transfection of these cells with a plasmid encoding the DNAS1L3 gene suggesting that this endonuclease is required for acetaminophen‐induced internucleosomal DNA fragmentation. DNAS1L3 expression potentiated the cytotoxic effect of acetaminophen in HeLa cells suggesting an active role in the death process induced by this drug. Altogether, these results demonstrate the specific roles of caspase‐3, DNA fragmentation factor, and DNAS1L3 in the process of acetaminophen‐induced apoptosis in cultured cells.