Inhibition of hepatic mitochondrial aldehyde dehydrogenase by carbon tetrachloride through JNK-mediated phosphorylation

The aim of this study was to investigate the mechanism of inhibition of mitochondrial aldehyde dehydrogenase (ALDH2) by carbon tetrachloride (CCl₄). CCl₄ administration caused marked hepatocyte ballooning and necrosis in the pericentral region. CCl₄ also inhibited hepatic ALDH2 activity in a time-dependent manner without altering the protein level, suggesting ALDH2 inhibition through covalent modifications such as phosphorylation by JNK. To demonstrate phosphorylation, the isoelectric point (pI) of ALDH2 in CCl₄-exposed rats was compared to that of untreated controls. Immunoblot analysis revealed that immunoreactive ALDH2 bands in CCl₄-exposed rats were shifted to acidic pI ranges on two-dimensional electrophoresis (2-DE) gels. Incubation with alkaline phosphatase significantly restored the suppressed ALDH2 activity with a concurrent alkaline pI shift of the ALDH2 spots. Both JNK and activated JNK were translocated to mitochondria after CCl₄ exposure. In addition, incubation with catalytically active JNK led to significant inhibition of ALDH2 activity, with an acidic pI shift on 2-DE gels. Furthermore, immunoprecipitation followed by immunoblot analysis with anti-phospho-Ser–Pro antibody revealed phosphorylation of a Ser residue(s) of ALDH2. These results collectively indicate a novel underlying mechanism by which CCl₄ exposure activates JNK, which translocates to mitochondria and phosphorylates ALDH2, contributing to inhibition of ALDH2 activity accompanied by decreased cellular defense capacity and increased lipid peroxidation.