Abstract Despite much progress, alcoholic liver disease (ALD) remains a major health problem worldwide. The disease process is characterized by early steatosis, steatohepatitis, with some individuals ultimately progressing to fibrosis/cirrhosis and liver failure. Unfortunately, there is currently no accepted therapies available to halt or reverse this process in humans. Nicotinamide N-methyltransferase (NNMT) catalyzes SAM-dependent degradation of nicotinamide, a predominant precursor for cellular NAD+ biosynthesis via a salvage pathway. SAM (s-adenosylmethionine) is the first product of methionine metabolism and a universal methyl donor in cellular transmethylation reactions. The critical role of NNNT in regulating both NAD+ and SAM homeostasis make it an emerging novel metabolic regulator. We are the first to report that the liver ATF4 transactivation plays a mechanistic role in mediating NNMT upregulation in the setting of chronic alcohol consumption and adenoviral shRNA knockdown of NNMT is protective against alcoholic fatty liver development, suggesting that NNMT can be an ideal therapeutic choice for ALD treatment. The preliminary data recently obtained from our laboratory uncovered that NNMT inhibition was associated with improved mitochondrial unfolded protein response (UPRmt) and blunted hepatic PPAR-gamma activation upon chronic alcohol exposure. In this proposal, we will further elucidate the mechanistic implication of NNMT in the pathogenesis of ALD. Successful performance of the studies proposed in this proposal will not only shed new light on the pathogenesis of this disease, but also pave the way for novel therapeutic interventions for ALD. The three aims are included in this proposal to test our hypothesis: AIM 1: To delineate mechanism(s) underlying NNMT-associated liver pathologies in ALD. Both animal and cell culture studies will be conducted to elucidate mechanism by which NNMT inhibition improves UPRmt elicitation in the liver and to determine the mechanism whereby NNMT upregulation contributes alcohol-induced liver PPAR-gamma activation. AIM 2: To elucidate mechanism(s) by which chronic alcohol consumption leads to hepatic ATF4 activation and NNMT upregulation. Both hepatocyte- specific Gcn2 knockout mice and hepatocyte-specific arginase-1 overexpressing mice will be fed with isocaloric control or alcohol-diet for 5 weeks. Targeted metabolomics will be conducted to using primary hepatocytes to quantify the effects of alcohol on hepatic arginine metabolism. AIM 3: To determine both preventive (the pathogenic role) and therapeutic potential of NNMT-targeting approach for ALD. Animals with liver-specific NNMT knockout will be exposed to either isocaloric control or alcohol-diet. Both preventive and therapeutic efficacy of NNMT inhibition for ALD will be evaluated.