Perturbation of transmembrane 6 superfamily member 2 expression alters lipid metabolism in a human liver cell line

Abstract Background Nonalcoholic fatty liver disease (NAFLD) is caused by accumulation of excess lipids in hepatocytes. Genome wide association studies have identified strong association of NAFLD with non-synonymous E167K amino acid mutation in transmembrane 6 superfamily member 2 (TM6SF2) protein. The E167K mutation affects TM6SF2 stability and its carriers display increased hepatic lipids levels and lower serum triglycerides. While similar phenotype is evident in mice with TM6SF2 knockdown, effects of TM6SF2 on hepatic lipid metabolism is not completely understood. Methods Here, we overexpressed wild-type or E167K variant of TM6SF2 or knocked down TM6SF2 expression in lipid-treated Huh-7 cells and used biochemical assays, untargeted lipidomic analysis, RNAseq transcriptome analysis and high-throughput fluorescent imaging to determine changes in lipid metabolism. Results Both knockdown and E167K overexpression increased acylglyceride levels which was decreased by wild-type TM6SF2 overexpression. Further, mean intensity of individual lipid droplets was increased by E167K overexpression and knockdown while wild-type TM6SF2 had no effects. We also observed lipid chain remodeling for acylglycerides by TM6SF2 knockdown leading to a relative increase in species with shorter and more saturated side chains. RNA sequencing revealed differential expression of several lipid metabolizing genes, including genes belonging to AKR1 family and lipases, primarily in cells with TM6SF2 knockdown. Conclusion Taken together, our data shows that overexpression of TM6SF2 gene or its loss-of-function changes hepatic lipid species composition and expression of lipid metabolizing genes. Further, overexpression of E167K variant and TM6SF2 knockdown similarly increased hepatic lipid accumulation and lipid droplets profile further confirming a loss-of-function effect for variant.