Induction of microsomal epoxide hydrolase by sulfur amino acid deprivation via the pathway of C-Jun N-terminal kinase and its extracellular exposure during cell death.
Microsomal epoxide hydrolase (mEH), an epoxide detoxifying enzyme and putative cell surface autoantigen, is inducible by xenobiotics and by certain pathophysiological conditions (e.g., tumorigenesis and protein-calorie malnutrition). The present study was designed to determine mEH expression in H4IIE cells during cell death initiated by sulfur amino acid deprivation (SAAD) and to identify the signaling pathway for the enzyme induction. SAAD induced cell death at 48-72 h with translocation of Bax to mitochondria and increased mitochondrial permeability with cytochrome c release, both of which were prevented by SB203580 or by dominant-negative JNK1 [JNK1(-)] stable transfection. Caspase-3 activity was only marginally increased by SAAD. Neither genomic DNA fragmentation nor poly(ADP-ribose) polymerase cleavage was observed during SAAD-induced cell death. Thus, SAAD induced cell death independent of caspase activation. This was supported by the observation that benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone, a general caspase inhibitor, did not prevent cell death. The levels of mEH mRNA and protein were notably increased in cells under SAAD for 48-72 h. The induction of mEH occurred in parallel with cell death. Whereas SAAD-induced cell death resulted from both JNK1 and p38 kinase activation, mEH induction was decreased only by JNK1(-) transfection. Immunocytochemistry revealed that mEH protein was intensely stained in dying cells, cellular fragments and cell debris. Furthermore, the number of cells positive for surface mEH substantially increased by SAAD, as evidenced by flow cytometry analysis. These results demonstrated that SAAD induced nonapoptotic cell death with Bax translocation to mitochondria and mitochondrial cytochrome c release, but not through caspase-3 activation, and that mEH was induced by SAAD via the pathway of JNK1, but not ERK1/2 or p38 kinase, in parallel with cell death.
Kang
Keon Wook
KW
National Research Laboratory (MDT), College of Pharmacy, Seoul National University, Sillim-dong Kwanak-gu, Seoul 151-742, South Korea.
Novak
Raymond F
RF
Lee
Chang Ho
CH
Kim
Sang Geon
SG
D000603:Amino Acids, Sulfur / Q000172:deficiency* / Q000494:pharmacology; D000818:Animals; D015152:Blotting, Northern; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D016923:Cell Death / Q000502:physiology; D003574:Cytochrome c Group / Q000378:metabolism; D004790:Enzyme Induction; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004851:Epoxide Hydrolases / Q000096:biosynthesis*; D005455:Fluorescent Antibody Technique; D007124:Immunoenzyme Techniques; D020287:In Situ Nick-End Labeling; D048031:JNK Mitogen-Activated Protein Kinases; D008861:Microsomes / Q000201:enzymology*; D008928:Mitochondria / Q000201:enzymology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D051381:Rats; D015398:Signal Transduction / Q000502:physiology*; D013778:Tetrazolium Salts; D013844:Thiazoles; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D051028:bcl-2-Associated X Protein; D048051:p38 Mitogen-Activated Protein Kinases
D000603:Amino Acids, Sulfur; C494828:Bax protein, rat; D061945:Caspase Inhibitors; D003574:Cytochrome c Group; D004791:Enzyme Inhibitors; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D013778:Tetrazolium Salts; D013844:Thiazoles; D051028:bcl-2-Associated X Protein; D011065:Poly(ADP-ribose) Polymerases; D048031:JNK Mitogen-Activated Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D004851:Epoxide Hydrolases; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; C022616:thiazolyl blue