Redox Regulation of O-GlcNAcylation Signaling in the Pathogenesis of Alcoholic Fatty Liver Disease

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PROJECT SUMMARY Alcoholic fatty liver disease (AFLD) is a major cause of chronic liver disease. Globally, AFLD accounts for 0.9% of total mortality and 0.6% of disability-adjusted life years and remains a public health problem worldwide. Despite its profound health and economic impact, the AFLD management remains challenging due to the lack of detailed understanding of determinants of its pathogenesis and progression. The goal of this project is to elucidate a novel role of redox-modulated hepatic O-GlcNAc signaling in alcohol-induced liver injury, and thereby identify potential novel preventive, diagnostic and/or therapeutic targets against AFLD. Oxidative stress is implicated to play a central role in many pathways involved in the initiation and progression of AFLD. Hepatic glutathione (GSH) functions in maintaining cellular redox homeostasis. Intriguingly, chronic GSH deficiency in mice harboring a global disruption of the glutamate-cysteine ligase modifier subunit (Gclm) gene confers protection against alcohol-induced steatosis. Molecular and metabolomics studies indicate that the hepatoprotective effect of low GSH is linked to activations of AMPK signaling pathway and NRF2 antioxidant response, and reprogramming of hepatic metabolism that benefits the maintenance of cellular redox and metabolic homeostasis. Multiomics analyses imply that post-transcriptional mechanisms play a significant role in mediating low GSH-elicited metabolic adaptation upon ethanol exposure. O-GlcNAcylation of protein is a prevalent form of post-translational modification (PTM), where a single O-linked N-acetylglucosamine (O- GlcNAc) moiety is added to nuclear, cytoplasmic and mitochondrial proteins. This process is controlled by a pair of O-GlcNAc cycling enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). In the liver, O- GlcNAc signaling has been shown to serve as an important mechanism for nutrient and stress sensing and subsequent regulation of liver cellular homeostasis. Aberrant O-GlcNAcylation has been implicated in hepatic insulin resistance, fatty liver disease and associated fibrosis. To date, little is known about the role of O- GlcNAcylation in AFLD pathogenesis. Our preliminary studies show that, in GSH-deficient Gclm-null mouse livers, chronic-binge ethanol feeding induced beneficial changes in global O-GlcNAylation and associated cellular pathways and signaling proteins. This project will expand upon these provocative findings and test the hypothesis predicting that adaptive O-GlcNAc signaling driven by chronic oxidative stress (due to GSH deficiency) in the liver protects against alcohol-induced liver injury. We propose to (1) utilize the Gclm-null mouse model to examine ethanol-induced changes in O-GlcNAcylation and phosphorylation modifications of the liver proteome and substrate targeting of O-GlcNAc cycling enzymes, and (2) evaluate the functional impacts of top candidate protein-O-GlcNAcylations in ethanol-associated stress response in human HepaRG cells. The proposed study will be the first examination of in vivo interplay between hepatic redox status and O- GlcNAcylation signaling in the context of alcohol induced liver injury.