ABSTRACT Methionine adenosyltransferase (MAT) is an essential cellular enzyme that catalyzes the formation of S- adenosylmethionine (SAMe), the principal biological methyl donor. Mammals express MAT1A and MAT2A, which encode for α1 and α2, respectively, with MAT1A expressed mainly in normal liver. Patients with alcohol- associated liver disease (ALD) have decreased MAT1A expression and activity. In addition to the cytosol, we showed MATα1 is also in the nucleus to modulate gene expression via epigenetics and as a transcription co- factor. Our works from the past funding cycle showed MATα1 is also present in the mitochondrial matrix to regulate mitochondrial function. MATα1 interacts with CYP2E1 leading to CYP2E1 R379 methylation and proteasomal degradation. We further showed MATα1 mitochondrial targeting is impaired in the murine NIAAA model of ALD and human ALD. This is due to activation of casein kinase 2 in ALD, which phosphorylates MATα1 at serine 114 and this allows peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (PIN1) to bind and block MATα1 mitochondrial targeting. Our unpublished data show that MATα1 is also hyper-sumoylated in ALD at K48 (K47 in human) and this prevents PIN1 binding but in return, PIN1 is required for MATα1 to be sumoylated. Additionally, the same MATα1 post-translational modifications result in selective depletion of MATα1 nuclear content. In ALD, there is also increased interaction of PIN1 with Forkhead Box M1 (FOXM1), an oncoprotein that exerts a negative reciprocal regulation with MAT1A. Indeed, FOXM1 expression is induced in ALD, treatment with a small molecule inhibitor of FOXM1 and hepatocyte-specific Foxm1 knockout mice are protected from ALD. This renewal application will allow us to define three novel intertwined areas linked to MATα1 and test the central hypothesis that alcohol increases MATα1 sumoylation, phosphorylation, and interaction with PIN1, leading to mitochondrial and nuclear MATα1 depletion that drives ALD in part through FOXM1. Altered PIN1 interactome further contributes to this process. Here we follow up these novel findings in three specific aims: 1) examine the mechanisms for lower nuclear MATα1 content in ALD and its consequences. We will examine MATα1’s nuclear import and export and whether these processes are dysregulated in ALD, nuclear MATα1’s influence on the epigenome and identify its target genes, and whether raising nuclear MATα1 will protect from ALD. 2) examine the role of hepatocyte FOXM1 in ALD. We will define the FOXM1 phosphosites and signaling event that leads to increased interaction with PIN1, whether this interaction is important in ALD. We will also examine hepatocyte-derived FOXM1’s target genes in different liver cell populations. 3) examine the role of PIN1 in ALD. We will define PIN1 interactome in normal liver vs. ALD, examine the efficacy of small molecule inhibitors of PIN1-MATα1 interaction in ALD. Lastly, we will decipher the crosstalk between PIN1 and sumoy...