Perilipin A increases triacylglycerol storage by decreasing the rate of triacylglycerol hydrolysis

The perilipins are the most abundant proteins at the surfaces of lipid drop lets in adipocytes and are also found in steroidogenic cells. To investigat e perilipin function, perilipin A, the predominant isoform, was ectopically expressed in fibroblastic 3T3-L1 pre-adipocytes that normally lack the per ilipins. In control cells, fluorescent staining of neutral lipids with Bodi py 493/503 showed a few minute and widely dispersed lipid droplets, while i n cells stably expressing perilipin A, the lipid droplets were more numerou s and tightly clustered in one or two regions of the cytoplasm. Immunofluor escence microscopy revealed that the ectopic perilipin A localized to the s urfaces of the tiny clustered lipid droplets; subcellular fractionation of the cells using sucrose gradients confirmed that the perilipin A localized exclusively to lipid droplets. Cells expressing perilipin A stored 6-30-fol d more triacylglycerol than control cells due to reduced lipolysis of triac ylglycerol stores. The lipolysis of stored triacylglycerol was 5 times slow er in lipid-loaded cells expressing perilipin A than in lipid-loaded contro l cells, when triacylglycerol synthesis was blocked with 6 muM triacsin C. This stabilization of triacylglycerol was not due to the suppression of tri acylglycerol lipase activity by the expression of perilipin A. We conclude that perilipin A increases the triacylglycerol content of cells by forming a barrier that reduces the access of soluble lipases to stored lipids, thus inhibiting triacylglycerol hydrolysis. These studies suggest that perilipi n A plays a major role in the regulation of triacylglycerol storage and lip olysis in adipocytes.