To understand this paradoxical result, we applied a capillary isoelectric focusing (IEF) method to determine the pattern of FOXO3 posttranslational modifications (PTMs) induced by HCV and alcohol. We observed the presence of multiple different nuclear and cytosolic species of FOXO3 and used antiphosphoserine, find more acetyl-lysine, methylarginine, and ubiquitin antibodies to identify the PTM patterns present in each species. HCV caused

multiple changes including phosphorylation of FOXO3 at S-574, a novel c-Jun N-terminal kinase (JNK) site, which promoted nuclear translocation and transcription. Ethanol suppressed arginine-methylation of FOXO3 promoting nuclear export and degradation of the JNK phosphorylated form. Human liver biopsy samples showed the presence of the HCV-specific form of FOXO3 in HCV-infected

livers but not in normal liver or nonalcoholic steatohepatitis. Conclusion: The development of this novel IEF method for the simultaneous quantification of differently modified FOXO3 species allowed us to demonstrate how HCV and alcohol combine to modify a complex pattern of FOXO3 PTMs that contribute to pathogenesis. This approach will allow further dissection of the role of protein PTMs in viral liver disease. (Hepatology 2014;58:58–70) Hepatitis C virus (HCV) and alcohol

MCE公司 ABT-737 concentration each cause liver injury that results from a combination of immune-mediated cytotoxicity and alterations in adaptive signaling pathways within hepatocytes. While these two disease-causing agents produce liver injury by themselves, there is considerable evidence that when present in combination HCV and alcohol have effects that do not occur with either stimulus alone. In epidemiological studies, the alcohol-HCV combination results in rapid fibrosis progression, impaired viral clearance, and enhanced carcinogenesis.[1] In cell culture, synergistic effects include induction of cell death pathways, mitochondrial reactive oxygen species (ROS) production, and suppression of antioxidant protein expression.[2] Recent studies have shown that the function of FOXO transcription factors is altered as a consequence of HCV infection, potentially contributing to insulin resistance and impaired activation of starvation-induced autophagy.[3] FOXO transcription factors control expression of proteins responsible for longevity, antioxidant response, cell cycle arrest, insulin sensitivity, apoptosis, and autophagy.[4, 5] FOXO3 is also a tumor suppressor.[4, 6] FOXO proteins are regulated by a complex series of posttranslational modifications (PTMs) that have collectively been suggested to constitute a “FOXO code.