Disrupting the PNPLA3-ABHD5 Interaction Stimulates Lipolysis and Improves PNPLA3-I148M–Associated Hepatic Steatosis

People with an I148M mutation in the PNPLA3 protein are at high risk of developing hepatic steatosis and progressive liver disease. The PNPLA3-I148M mutant protein accumulates on the membrane of lipid droplets (LDs) where it can interfere with the function of adipose tissue triglyceride lipase (ATGL) by depleting their shared cofactor α/β-hydrolase domain-containing protein 5 (ABHD5). We hypothesized that restoring ABHD5 supply to ATGL by either ABHD5 overexpression or interruption of the protein-protein interaction (PPI) between ABHD5 and PNPLA3-I148M would recover lipolysis and ameliorate steatosis. In PNPLA3-I148M–expressing HuH7 cells and primary human hepatocytes homozygous for the PNPLA3-I148M mutation, overexpression of ABHD5 stimulated lipolysis and reduced LD accumulation. In PNPLA3-I148M knockin mice, adeno-associated virus–mediated overexpression of ABHD5 improved diet-induced hepatic steatosis. The amino acids (AAs) essential for the PPI between ABHD5 and PNPLA3 were identified using AlphaFold structural predictions, and AA substitutions (F17A/R120A) were verified to disrupt the PPI via competitive binding assay. Disruption of the PNPLA3-ABHD5 PPI reduced LD accumulation in HuH7 cells and improved hepatic steatosis in PNPLA3-I148M knockin mice. Together, our data support the hypothesis that PNPLA3-I148M causes hepatic steatosis via sequestration of ABHD5 from ATGL and that disruption of the PPI between ABHD5 and PNPLA3-I148M can resolve hepatic dysfunction. Article Highlights We evaluate the importance of the protein-protein interaction (PPI) between PNPLA3-I148M and its cofactor α/β-hydrolase domain-containing protein 5 (ABHD5) in PNPLA3-I148M–mediated hepatic steatosis. We demonstrate that disrupting the PNPLA3-ABHD5 PPI improves I148M-associated steatosis and markers of liver injury. This work supports the development of therapies targeting the PNPLA3-ABHD5 PPI for the treatment of PNPLA3-I148M–associated metabolic dysfunction–associated steatotic liver disease.