Selective Activation of the c-Jun N-Terminal Protein Kinase Pathway during 4-Hydroxynonenal-Induced Apoptosis of PC12 Cells
- Yunjo Soh1,2,1,
- Kyu-Shik Jeong1,3,1,
- Insong James Lee4,1,
- Myung-Ae Bae1,
- Yong-Chul Kim2 and
- Byoung J. Song1
- 1Laboratory of Membrane Biochemistry and Biophysics, National Institute on Alcohol Abuse and Alcoholism, Rockville, Maryland (Y.S., K.-S.J., I.J.L., M.-A.B., B.J.S.); and 2Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, Maryland (Y.-C.K.)
Abstract
The by-product of lipid peroxidation, 4-hydroxynonenal (HNE), was shown to cause apoptosis in PC12 cells. In this study, we investigated the molecular mechanism of HNE-induced apoptosis in these cells. Specifically, we determined the effect of HNE on the activities of mitogen-activated protein (MAP) kinases involved in early signal transduction. Within 15 to 30 min after HNE treatment, c-Jun N-terminal protein kinase (JNK) was maximally activated, before it returned to control level at 1 h post-treatment. In contrast, activities of extracellular signal-regulated kinase and p38 MAP kinase remained unchanged from their baseline levels. Stress-activated protein kinase kinase (SEK1), an upstream kinase of JNK, was also activated within 5 min after HNE treatment and remained activated for up to 60 min. Marked activation of the JNK pathway through SEK1 and apoptosis signal-regulating kinase 1 (ASK1), an upstream kinase of SEK1, was demonstrated by the transient transfection of cDNA for wild-type SEK1 or ASK1 together with JNK into COS-7 cells. Furthermore, significant reductions in JNK activation and HNE-induced cell death were observed when either of the dominant negative mutant of SEK1 or ASK1 was cotransfected with JNK. Pretreatment of PC12 cells with a survival-promoting agent, 8-(4-chlorophenylthio)-cAMP, prevented both the HNE-induced JNK activation and apoptosis. Nonaldehyde, a nontoxic aldehyde, neither caused apoptosis nor JNK activation. Pretreatment of PC12 cells with SB203580, a specific inhibitor of p38 MAP kinase, had no effect on HNE-induced apoptosis. All these data suggest that the selective JNK activation by HNE is critical for the apoptosis of PC12 cells and that the HNE-mediated apoptosis is likely to be mediated through the activation of the ASK1-SEK1-JNK pathway without activation of extracellular signal-regulated kinase or p38 MAP kinase.
Footnotes
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Send reprint requests to: Byoung J. Song, Laboratory of Membrane Biochemistry and Biophysics, National Institute on Alcohol Abuse and Alcoholism, 12420 Parklawn Dr., Rm. 425, Rockville, MD 20852. E-mail: bs7t{at}nih.gov
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↵1 Both authors contributed equally to the present work.
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↵2 Current address: Department of Neuroscience, Graduate School of East-West Medical Science, Kyung-Hee University, Seoul, Korea.
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↵3 Current address: Korea Research Institute of Bioscience and Biotechnology, Taejon, Korea.
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↵4 Current address: EpiGenesis Pharmaceuticals, Inc., 2005 Eastpark Blvd., Cranbury, NJ.
- Abbreviations:
- HNE
- 4-hydroxy-2-nonenal
- ERK
- extracellular signal-regulated protein kinase
- SAPK
- stress-activated protein kinase
- JNK
- c-Jun N-terminal protein kinase
- p38 MAP kinase
- p38 mitogen-activated protein kinase
- SEK1
- SAPK kinase
- MAP kinase
- mitogen-activated protein kinase
- MEKK
- MAP kinase kinase kinase
- ASK1
- apoptosis signal-regulating kinase 1
- HA
- hemagglutinin
- DAPI
- 4,6-diamidino-2-phenylindole
- CPT-cAMP
- 8-(4-chlorophenylthio)-adenosine 3′:5′-cyclic monophosphate
- wt
- wild type
- ATF-2
- activator of transcription factor-2
- MTS
- 3-[4,5-dimethylthiazol-2-yl]-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2-tetrazolium
- MKK
- MAP kinase kinase
- MTT
- 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide
- AP-1
- activator protein 1
- GST
- glutathione S-transferase
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- Received December 10, 1999.
- Accepted May 30, 2000.
- U.S. Government
