Abrin triggers cell death by inactivating a thiol-specific antioxidant protein.
Abrin A-chain (ABRA) inhibits protein synthesis by its N-glycosidase activity as well as induces apoptosis, but the molecular mechanism of ABRA-induced cell death has been obscure. Using an ABRA mutant that lacks N-glycosidase activity as bait in a yeast two-hybrid system, a 30-kDa antioxidant protein-1 (AOP-1) was found to be an ABRA(E164Q)-interacting protein. The interaction was further confirmed in vitro by a glutathione S-transferase pull-down assay. The colocalization of endogenous AOP-1 and exogenous ABR proteins in the cell was demonstrated by confocal immunofluorescence. We also demonstrated that ABRA attenuates AOP-1 antioxidant activity in a dose-dependent manner and the intracellular level of reactive oxygen species (ROS) increases in ABR-treated cells. Moreover, ROS scavengers N-acetylcysteine and 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl delayed programmed cell death. This indicates that ROS are important mediators of ABR-induced apoptosis. When ectopically expressed, AOP-1 blocked the release of cytochrome c and prevented apoptosis in ABR-treated cells. These findings suggest that the binding of ABRA to AOP-1 promotes apoptosis by inhibiting the mitochondrial antioxidant protein AOP-1, resulting in the increase of intracellular ROS and the release of cytochrome c from the mitochondria to the cytosol, which activates caspase-9 and caspase-3.
Shih
S F
SF
Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, Taipei 10081, Taiwan, Republic of China.
Wu
Y H
YH
Hung
C H
CH
Yang
H Y
HY
Lin
J Y
JY
D000036:Abrin / Q000493:pharmacokinetics / Q000494:pharmacology*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000975:Antioxidants; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001692:Biological Transport; D016923:Cell Death / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D003001:Cloning, Molecular; D003574:Cytochrome c Group / Q000032:analysis; D006367:HeLa Cells; D006801:Humans; D019169:Jurkat Cells; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry; D010544:Peroxidases / Q000737:chemistry; D061105:Peroxiredoxin III; D054464:Peroxiredoxins; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000493:pharmacokinetics / Q000494:pharmacology; D012441:Saccharomyces cerevisiae; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D000975:Antioxidants; D003574:Cytochrome c Group; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D011994:Recombinant Proteins; D000036:Abrin; D010544:Peroxidases; C516115:PRDX3 protein, human; D061105:Peroxiredoxin III; D054464:Peroxiredoxins