Nuclear translocation of endonuclease G and apoptosis-inducing factor during acetaminophen-induced liver cell injury.
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.
Bajt
Mary Lynn
ML
Liver Research Institute, University of Arizona, College of Medicine, Tucson, Arizona 85724, USA.
Cover
Cathleen
C
Lemasters
John J
JJ
Jaeschke
Hartmut
H
D000082:Acetaminophen / Q000633:toxicity*; D000111:Acetylcysteine / Q000494:pharmacology; D021581:Active Transport, Cell Nucleus; D000818:Animals; D051033:Apoptosis Inducing Factor / Q000378:metabolism*; D051017:Apoptosis Regulatory Proteins; D001562:Benzimidazoles / Q000378:metabolism; D002232:Carbocyanines / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D002467:Cell Nucleus / Q000187:drug effects* / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D045304:Cytochromes c / Q000378:metabolism; D053938:DNA Fragmentation / Q000187:drug effects; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D007150:Immunohistochemistry / Q000379:methods; D008297:Male; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D051379:Mice; D008809:Mice, Inbred C3H; D008856:Microscopy, Fluorescence / Q000379:methods; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D024101:Mitochondrial Proteins / Q000378:metabolism
D051033:Apoptosis Inducing Factor; D051017:Apoptosis Regulatory Proteins; D001562:Benzimidazoles; D002232:Carbocyanines; D002352:Carrier Proteins; C499605:Diablo protein, mouse; D024101:Mitochondrial Proteins; C068624:5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine; D000082:Acetaminophen; D045304:Cytochromes c; D004706:Endodeoxyribonucleases; C052391:endonuclease G; D000111:Acetylcysteine