Journal
JOURNAL OF BIOLOGICAL CHEMISTRY
Volume 289, Issue 22, Pages 15449-15462Publisher
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M113.543942
Keywords
Alcohol; AMP-activated Kinase (AMPK); Lipid Peroxidation; Liver; Oxidative Stress
Categories
Funding
- Center for Research Resources, National Institutes of Health Colorado Clinical and Translational Sciences Institute [UL1 RR025780]
- University of Colorado Denver-Anchutz Medical Campus Proteomic Mass Spectrometry Facility
- Colorado Clinical Translational Science Institute
- University of Colorado Cancer Center Grants Clinical and Translational Science Award [UL1 RR025780]
- University of Colorado Cancer Center [P30 CA046934]
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The production of reactive aldehydes including 4-hydroxy-2-nonenal (4-HNE) is a key component of the pathogenesis in a spectrum of chronic inflammatory hepatic diseases including alcoholic liver disease (ALD). One consequence of ALD is increased oxidative stress and altered -oxidation in hepatocytes. A major regulator of -oxidation is 5 AMP protein kinase (AMPK). In an in vitro cellular model, we identified AMPK as a direct target of 4-HNE adduction resulting in inhibition of both H2O2 and 5-aminoimidazole-4-carboxyamide ribonucleoside (AICAR)-induced downstream signaling. By employing biotin hydrazide capture, it was confirmed that 4-HNE treatment of cells resulted in carbonylation of AMPK/, which was not observed in untreated cells. Using a murine model of alcoholic liver disease, treatment with high concentrations of ethanol resulted in an increase in phosphorylated as well as carbonylated AMPK. Despite increased AMPK phosphorylation, there was no significant change in phosphorylation of acetyl CoA carboxylase. Mass spectrometry identified Michael addition adducts of 4-HNE on Cys(130), Cys(174), Cys(227), and Cys(304) on recombinant AMPK and Cys(225) on recombinant AMPK. Molecular modeling analysis of identified 4-HNE adducts on AMPK suggest that inhibition of AMPK occurs by steric hindrance of the active site pocket and by inhibition of hydrogen peroxide induced oxidation. The observed inhibition of AMPK by 4-HNE provides a novel mechanism for altered -oxidation in ALD, and these data demonstrate for the first time that AMPK is subject to regulation by reactive aldehydes in vivo.
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