16896059

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1

Mitochondrial dysfunction and internucleosomal DNA fragmentation are well-recognized features of acetaminophen (AAP)-induced hepatocyte cell death. However, the endonucleases responsible for this effect have not been identified. Apoptosis-inducing factor (AIF) and endonuclease G are nucleases located in the intermembrane space of mitochondria. AIF is thought to trigger chromatin condensation and induce cleavage of DNA into high molecular weight fragments (50-300 kb), and endonuclease G can produce oligonucleosomal DNA fragments. Therefore, the objective of this investigation was to test the hypothesis that endonuclease G and AIF could be involved in AAP-induced nuclear DNA fragmentation. Using immunofluorescence microscopy, it was shown that in primary cultured mouse hepatocytes, endonuclease G and AIF translocated to the nucleus between 3 and 6 h after exposure to 5 mM AAP. In contrast, other mitochondrial intermembrane proteins such as cytochrome c or the second mitochondria-derived activator of caspases (Smac) did not accumulate in the nucleus. The translocation of AIF and endonuclease G correlated with mitochondrial dysfunction as indicated by the progressive loss of the mitochondrial membrane potential (measured with the JC-1 assay) and the appearance of nuclear DNA fragments in the cytosol (determined by an anti-histone ELISA). Pretreatment with 20mM N-acetylcysteine prevented mitochondrial dysfunction, the nuclear translocation of endonuclease G and AIF, and the nuclear DNA fragmentation. The data support the conclusion that endonuclease G and AIF translocate to the nucleus in response to AAP-induced mitochondrial dysfunction and may be responsible, at least in part, for the initial DNA fragmentation during AAP hepatotoxicity.

eng

IM

MEDLINE

1096-6080

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217-25

pubmed-meshheading:16896059-Acetaminophen, pubmed-meshheading:16896059-Acetylcysteine, pubmed-meshheading:16896059-Active Transport, Cell Nucleus, pubmed-meshheading:16896059-Animals, pubmed-meshheading:16896059-Apoptosis Inducing Factor, pubmed-meshheading:16896059-Benzimidazoles, pubmed-meshheading:16896059-Carbocyanines, pubmed-meshheading:16896059-Carrier Proteins, pubmed-meshheading:16896059-Cell Nucleus, pubmed-meshheading:16896059-Cell Survival, pubmed-meshheading:16896059-Cells, Cultured, pubmed-meshheading:16896059-Cytochromes c, pubmed-meshheading:16896059-DNA Fragmentation, pubmed-meshheading:16896059-Endodeoxyribonucleases, pubmed-meshheading:16896059-Hepatocytes, pubmed-meshheading:16896059-Immunohistochemistry, pubmed-meshheading:16896059-Male, pubmed-meshheading:16896059-Membrane Potential, Mitochondrial, pubmed-meshheading:16896059-Mice, pubmed-meshheading:16896059-Mice, Inbred C3H, pubmed-meshheading:16896059-Microscopy, Fluorescence, pubmed-meshheading:16896059-Mitochondria, pubmed-meshheading:16896059-Mitochondrial Proteins

Nuclear translocation of endonuclease G and apoptosis-inducing factor during acetaminophen-induced liver cell injury.

Journal Article, Research Support, N.I.H., Extramural