Role of MAPK phosphatase-1 in sustained activation of JNK during ethanol-induced apoptosis in hepatocyte-like VL-17A cells

Senthil K. Venugopal, Jenny Chen, Yanhong Zhang, Dahn Clemens, Antonia Follenzi, Mark A Zern

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Abstract

Ethanol metabolism plays a central role in activating the mitogen-activated protein kinase (MAPK) cascade leading to inflammation and apoptosis. Sustained activation of c-Jun N-terminal kinase (JNK), one of the MAPKs, has been shown to induce apoptosis in hepatocytes. MAPK phosphatase-1 (MKP-1) has been shown to dephosphorylate MAPKs in several cells. The aim of the study is to evaluate the role of MKP-1 in sustained JNK activation as a mechanism to explain ethanol-induced hepatocyte apoptosis. VL-17A cells (HepG2 cells overexpressing alcohol dehydrogenase and cytochrome P450-2E1) were exposed to ethanol for different time periods. Western blots were performed for MKP-1, phospho-JNK, phosphotyrosine, and protein kinase Cδ(PKCδ). Electrophoretic mobility shift assays for AP-1 were performed. Apoptosis was measured by caspase-3 activity assay, TUNEL, and 4′,6-diamidino-2-phenylindole staining. Reactive oxygen species were neutralized by overexpressing both superoxide dismutase-3 and catalase genes using lentiviral vectors in VL-17A cells. Ethanol incubation markedly decreased the MKP-1 protein levels to 15% of control levels and was associated with sustained phosphorylation of p46 JNK and p54 JNK, as well as increased apoptosis. VL-17A cells overexpressing superoxide dismutase-3 and catalase, treatment with a tyrosine kinase inhibitor, or incubation of the cells with PKCδ small interference RNAs significantly inhibited the ethanol-induced MKP-1 degradation and apoptosis. Ethanol-induced oxidative stress enhanced the tyrosine phosphorylation of PKCδ, which in turn caused the proteasomal degradation of MKP-1, leading to sustained JNK activation and increased apoptosis in VL-17A cells.

Original languageEnglish (US)
Pages (from-to)31900-31908
Number of pages9
JournalJournal of Biological Chemistry
Volume282
Issue number44
DOIs
StatePublished - Nov 2 2007

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Mitogen-Activated Protein Kinase Phosphatases
Dual Specificity Phosphatase 1
JNK Mitogen-Activated Protein Kinases
Hepatocytes
Ethanol
Chemical activation
Apoptosis
Protein Kinase C
Phosphorylation
Protein-Tyrosine Kinases
Catalase
Superoxide Dismutase
Assays
Cytochrome P-450 CYP2E1
Degradation
Electrophoretic mobility
Phosphotyrosine
Oxidative stress
Alcohol Dehydrogenase
Level control

ASJC Scopus subject areas

  • Biochemistry

Cite this

Role of MAPK phosphatase-1 in sustained activation of JNK during ethanol-induced apoptosis in hepatocyte-like VL-17A cells. / Venugopal, Senthil K.; Chen, Jenny; Zhang, Yanhong; Clemens, Dahn; Follenzi, Antonia; Zern, Mark A.

In: Journal of Biological Chemistry, Vol. 282, No. 44, 02.11.2007, p. 31900-31908.

Research output: Contribution to journalArticle

Venugopal, Senthil K. ; Chen, Jenny ; Zhang, Yanhong ; Clemens, Dahn ; Follenzi, Antonia ; Zern, Mark A. / Role of MAPK phosphatase-1 in sustained activation of JNK during ethanol-induced apoptosis in hepatocyte-like VL-17A cells. In: Journal of Biological Chemistry. 2007 ; Vol. 282, No. 44. pp. 31900-31908.
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