Fatty acids (FAs) are essential for cell survival, yet their overaccumulation causes lipotoxicity. To prevent lipotoxicity, cells store excess FAs as triglycerides (TGs). In cultured cells TG synthesis is activated by excess unsaturated but not saturated FAs. Here, we identify Ubxd8 as a sensor for unsaturated FAs and regulator of TG synthesis. In cultured cells depleted of FAs, Ubxd8 inhibits TG synthesis by blocking conversion of diacylglycerols (DAGs) to TGs. Excess unsaturated but not saturated FAs relieve this inhibition. As a result, unsaturated FAs are incorporated into TGs, whereas saturated FAs are incorporated into DAGs. In vitro, unsaturated but not saturated FAs alter the structure of purified recombinant Ubxd8 as monitored by changes in its thermal stability, trypsin cleavage pattern, and oligomerization. These results suggest that Ubxd8 acts as a brake that limits TG synthesis, and this brake is released when its structure is altered by exposure to unsaturated FAs.

Hepatocytes play a crucial role in regulating lipid metabolism by exporting cholesterol and triglyceride into plasma through secretion of very low density lipoproteins (VLDL). VLDL production is also required for release of hepatitis C virus (HCV) from infected hepatocytes. Here, we show that long chain acylCoA synthetase 3 (ACSL3) plays a crucial role in secretion of VLDL and HCV from hepatocytes. In cultured human hepatoma Huh7 cells, ACSL3 is specifically required for incorporation of fatty acids into phosphatidylcholine. In cells receiving small interfering RNA targeting ACSL3, secretion of apolipoprotein B, the major protein component of VLDL, was inhibited and the lipoprotein was rapidly degraded. This inhibition in secretion was completely eliminated when these cells were treated with phosphatidylcholine. Treatment of cells with small interfering RNA targeting ACSL3 also inhibited secretion of HCV from Huh7-derived cells. These results identify ACSL3 as a new enzymatic target to limit VLDL secretion and HCV infection.