title: Ethanol metabolism activates cell cycle checkpoint kinase, Chk2.
abstract: Chronic ethanol abuse results in hepatocyte injury and impairs hepatocyte replication. We have previously shown that ethanol metabolism results in cell cycle arrest at the G2/M transition, which is partially mediated by inhibitory phosphorylation of the cyclin-dependent kinase, Cdc2. To further delineate the mechanisms by which ethanol metabolism mediates this G2/M arrest, we investigated the involvement of upstream regulators of Cdc2 activity. Cdc2 is activated by the phosphatase Cdc25C. The activity of Cdc25C can, in turn, be regulated by the checkpoint kinase, Chk2, which is regulated by the kinase ataxia telangiectasia mutated (ATM). To investigate the involvement of the regulators of Cdc2 activity, VA-13 cells, which are Hep G2 cells modified to efficiently express alcohol dehydrogenase, were cultured in the presence or absence of 25 mM ethanol. Immunoblots were performed to determine the effects of ethanol metabolism on the activation of Cdc25C, Chk2, and ATM. Ethanol metabolism increased the active forms of ATM and Chk2, as well as the phosphorylated form of Cdc25C. Additionally, inhibition of ATM resulted in approximately 50% of the cells being rescued from the G2/M cell cycle arrest and ameliorated the inhibitory phosphorylation of Cdc2. Our findings demonstrated that ethanol metabolism activates ATM. ATM can activate the checkpoint kinase Chk2, resulting in phosphorylation of Cdc25C and ultimately in the accumulation of inactive Cdc2. This may, in part, explain the ethanol metabolism-mediated impairment in hepatocyte replication, which may be important in the initiation and progression of alcoholic liver injury.
authors.affiliation: Research Service, Veterans Administration Medical Center, 4101 Woolworth Avenue, Omaha, NE 68105, USA. dclemens@unmc.edu
authors.forename: Dahn L
authors.initials: DL
authors.lastname: Clemens
authors.forename: Katrina J Mahan
authors.initials: KJ
authors.lastname: Schneider
authors.forename: Robert F
authors.initials: RF
authors.lastname: Nuss
mesh_terms: D000426:Alcohol Dehydrogenase / Q000235:genetics; D064007:Ataxia Telangiectasia Mutated Proteins; D016203:CDC2 Protein Kinase / Q000378:metabolism; D059447:Cell Cycle Checkpoints; D018797:Cell Cycle Proteins / Q000378:metabolism*; D064447:Checkpoint Kinase 2; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004789:Enzyme Activation; D000431:Ethanol / Q000378:metabolism*; D015870:Gene Expression; D056945:Hep G2 Cells; D006801:Humans; D008108:Liver Diseases, Alcoholic / Q000201:enzymology; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics; D025521:Tumor Suppressor Proteins / Q000378:metabolism*; D020687:cdc25 Phosphatases / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D025521:Tumor Suppressor Proteins; D000431:Ethanol; D000426:Alcohol Dehydrogenase; D064447:Checkpoint Kinase 2; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C578634:CHEK2 protein, human; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase; D020687:cdc25 Phosphatases