Identification of 5′ AMP activated kinase as a target of reactive aldehydes during chronic ingestion of high concentrations of ethanol [Molecular Bases of Disease]

April 10th, 2014 by Shearn, C. T., Backos, D. S., Orlicky, D. J., Smathers-McCullough, R. L., Petersen, D. R.

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 β-oxidation in hepatocytes. A major regulator of β-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 H2O2 and AICAR-induced downstream signaling. By employing biotin hydrazide capture, it was confirmed that 4-HNE treatment of cells resulted in carbonylation of AMPKαβ 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 AMPKα. Despite increased AMPK phosphorylation, there was no significant change in phosphorylation of ACC. Mass spectrometry identified Michael addition adducts of 4-HNE on Cys130, Cys174, Cys227, and Cys304 on rAMPKα and Cys225 on rAMPKβ. Molecular modeling analysis of 4-HNE adducted to Cys130, Cys174 , Cys 227 , or Cys 304 on AMPKα suggest that inhibition of AMPK occurs by steric hindrance of the active site pocket and by inhibition of hydrogen peroxide induced oxidation. The inhibition of AMPK by 4-HNE provides a novel mechanism for altered β-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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