LYPLAL1 rare loss-of-function variants in humans and deletion in human hepatoma cells protect against MASLD

Genetic variants near LYPLAL1 are associated with Metabolic dysfunction-Associated Steatotic Liver Disease (MASLD) in humans, but their impact on LYPLAL1 function is unknown. We identified LYPLAL1 loss-of-function variants from UK BioBank (UKBB) whole-exome sequencing data that had AlphaMissense or GPN-MSA scores in the top 20% of LYPLAL1 variants for being disruptive. We aggregated these variants and carried out burden analysis for effects on MRI Proton Density-Fat Fraction (MRI-PDFF) and ICD-based MASLD in UKBB. Rare loss-of-function LYPLAL1 variants were associated with reduced MRI-PDFF and ICD diagnosed MASLD across sexes. We used CRISPR to knockout and overexpress LYPLAL1 in human hepatoma cells (HuH-7), measuring lipid content, lipid uptake/export, and changes in de novo lipogenesis and mitochondrial β-oxidation. LYPLAL1 subcellular localization was determined by overexpressing LYPLAL1-HA tagged protein. We purified GST tagged human LYPLAL1 protein and conducted in vitro tests for esterase and depalmitoylase activity. Knocking out LYPLAL1 reduced triglycerides biochemically as well as lipid intensity after oleic (18:1, n-9) acid treatment. LYPLAL1 KO cells had increased expression of PPARα and MLXIPL, increased mitochondrial β-oxidation, and reduced capacity to both import fatty acids (FAs) and export lipoproteins. Overexpression of LYPLAL1 increased lipid droplet accumulation and decreased PPARα and MLXIPL. LYPLAL1-HA is partly localized to mitochondria when treated with oleic acid. Biochemical analyses showed that LYPLAL1 has strong esterase activity but lacks depalmitoylase activity. Reduction of LYPLAL1 esterase function likely increases β-oxidation of FAs in mitochondria through PPARα and MLXIPL and decreases FA import to protect against lipid accumulation in human liver cancer cells. Together, our results indicate that LYPLAL1 loss-of-function protects against MASLD in Europeans and in vitro, LYPLAL1 is an esterase for short-chain substrates which is involved in the regulation of mitochondrial β-oxidation and uptake of fatty acids, influencing lipid accumulation in the liver.