The overall goal of this project is to define the metabolic and molecular basis of fatty liver disease (FLD), a burgeoning health problem with few therapeutic options. In 2004, our group performed the first survey of hepatic triglyceride (TG) content in a multiethnic population-based sample, the Dallas Heart Study (DHS). This survey showed that hepatic steatosis is much more common in Hispanics and less common in African- Americans (AA) relative to European-Americans (EA). To glean insights into the molecular underpinnings of this ethnic disparity we used human genetics to identify the first genetic variation (PNPLA3-148M) associated reproducibly with FLD. The variant accounts for a majority of the ancestry-related differences in HTGC among these 3 ethnic groups. The variant is not only associated with steatosis, but also steatohepatitis, cirrhosis and hepatocellular carcinoma. It confers equivalent risk for progression of alcoholic liver disease. Subsequently we identified a variant in TM6SF2 that also is associated with the full spectrum of FLD despite causing steatosis by a completely different mechanism. More recently we discovered the first variation that protects against progression of FLD. In this application we build on these discoveries to elucidate the pathogenic mechanisms of these variants and provide proof-of-principle experiments for therapeutic intervention.
In Aim 1 we will determine how PNPLA3-148M causes hepatic steatosis and develop strategies to reverse this process. Previously we showed that PNPLA3-148M accumulates on lipid droplets (LD). In this Aim we will determine how accumulation of PNPLA3 on LD promotes hepatic steatosis using a 148M ?knockin? mouse to model the human pathophysiology associated with this variant. Short hairpin(sh) RNAs delivered by adeno-associated virus (AAV), and siRNAs will be used to determine if knocking down PNPLA3-148M reverses steatosis. We will also target the regulatory machinery that controls both PNPLA3 expression and TG synthesis in the liver.
In Aim 2, we will determine the molecular basis of TM6SF2 167K- associated hepatic steatosis. Previously, we showed that TM6SF2 is an ER and Golgi protein that is required for bulk lipidation of VLDL and that Tm6Sf2-/- mice replicate the human phenotype. We will use a TM6SF2 knockout liver cell line to examine the role of TM6SF2 in VLDL assembly, trafficking, and secretion. We will take advantage of lysine-based targeting sequences at the C-terminal end of the protein to define the subcellular site(s) at which TM6SF2 promotes lipidation of nascent VLDL particles. We will test the hypothesis that TM6SF2 serves as a scaffold to coordinate addition of neutral lipids to VLDL in the secretory pathway.
In Aim 3 we will focus on a variant in HSD17B13 that is common in AA and confers resistance to FLD progression without altering HTGC. Successful completion of these 3 aims will provide new insights into hepatic TG homeostasis and new strategies and targets for the treatment of chronic liver disease.

Public Health Relevance

Fatty liver disease (FLD) is a burgeoning health problem with few therapeutic options. We have used human genetics to identify genetic variants that confer increased susceptibility (PNPLA3 and TM6SF2) to FLD, and resistance to disease progression (HSD17B13). In this application we will build on these discoveries to glean new insights into pathogenesis of FLD with a goal of developing new strategies and targets for the treatment of chronic liver disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK090066-08
Application #
9522203
Study Section
Integrative Nutrition and Metabolic Processes Study Section (INMP)
Program Officer
Doo, Edward
Project Start
2011-08-01
Project End
2023-07-31
Budget Start
2018-08-01
Budget End
2019-07-31
Support Year
8
Fiscal Year
2018
Total Cost
Indirect Cost
Name
University of Texas Sw Medical Center Dallas
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
800771545
City
Dallas
State
TX
Country
United States
Zip Code
75390
Mitsche, Matthew A; Hobbs, Helen H; Cohen, Jonathan C (2018) Patatin-like phospholipase domain-containing protein 3 promotes transfer of essential fatty acids from triglycerides to phospholipids in hepatic lipid droplets. J Biol Chem 293:6958-6968
Hobbs, Helen H (2018) Science, serendipity, and the single degree. J Clin Invest 128:4218-4223
Stender, Stefan; Smagris, Eriks; Lauridsen, Bo K et al. (2018) Relationship between genetic variation at PPP1R3B and levels of liver glycogen and triglyceride. Hepatology 67:2182-2195
Stender, Stefan; Kozlitina, Julia; Nordestgaard, Børge G et al. (2017) Adiposity amplifies the genetic risk of fatty liver disease conferred by multiple loci. Nat Genet 49:842-847
BasuRay, Soumik; Smagris, Eriks; Cohen, Jonathan C et al. (2017) The PNPLA3 variant associated with fatty liver disease (I148M) accumulates on lipid droplets by evading ubiquitylation. Hepatology 66:1111-1124
Smagris, Eriks; Gilyard, Shenise; BasuRay, Soumik et al. (2016) Inactivation of Tm6sf2, a Gene Defective in Fatty Liver Disease, Impairs Lipidation but Not Secretion of Very Low Density Lipoproteins. J Biol Chem 291:10659-76
Smagris, Eriks; BasuRay, Soumik; Li, John et al. (2015) Pnpla3I148M knockin mice accumulate PNPLA3 on lipid droplets and develop hepatic steatosis. Hepatology 61:108-18
Stender, Stefan; Chakrabarti, Rima S; Xing, Chao et al. (2015) Adult-onset liver disease and hepatocellular carcinoma in S-adenosylhomocysteine hydrolase deficiency. Mol Genet Metab 116:269-74
Wang, Yan; Gusarova, Viktoria; Banfi, Serena et al. (2015) Inactivation of ANGPTL3 reduces hepatic VLDL-triglyceride secretion. J Lipid Res 56:1296-307
Kozlitina, Julia; Smagris, Eriks; Stender, Stefan et al. (2014) Exome-wide association study identifies a TM6SF2 variant that confers susceptibility to nonalcoholic fatty liver disease. Nat Genet 46:352-6

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