Group IVA phospholipase A₂ is necessary for the biogenesis of lipid droplets

Lipid droplets (LD) are organelles present in all cell types, consisting of a hydrophobic core of triacylglycerols and cholesteryl esters, surrounded by a monolayer of phospholipids and cholesterol. This work shows that LD biogenesis induced by serum, by long-chain fatty acids, or the combination of both in CHO-K1 cells was prevented by phospholipase A₂ inhibitors with a pharmacological profile consistent with the implication of group IVA cytosolic phospholipase A₂ (cPLA₂α). Knocking down cPLA ₂α expression with short interfering RNA was similar to pharmacological inhibition in terms of enzyme activity and LD biogenesis. A Chinese hamster ovary cell clone stably expressing an enhanced green fluorescent protein-cPLA₂α fusion protein (EGFP-cPLA₂) displayed higher LD occurrence under basal conditions and upon LD induction. Induction of LD took place with concurrent phosphorylation of cPLA ₂α at Ser⁵⁰⁵. Transfection of a S505A mutant cPLA₂α showed that phosphorylation at Ser⁵⁰⁵ is key for enzyme activity and LD formation. cPLA₂α contribution to LD biogenesis was not because of the generation of arachidonic acid, nor was it related to neutral lipid synthesis. cPLA₂α inhibition in cells induced to form LD resulted in the appearance of tubulo-vesicular profiles of the smooth endoplasmic reticulum, compatible with a role of cPLA ₂α in the formation of nascent LD from the endoplasmic reticulum.