A genome-wide in vivo RNAi screen in Drosophila identifies regulators of cholesterol-dependent steroid production

Speaker: Kim Furbo Rewitz, Cell Biology and Neurobiology
Host: Peter Brodersen, Computational and RNA Biology

Abstract
Steroid hormones are signal molecules synthesized from cholesterol that regulate a variety of processes during embryogenesis, postembryonic development and reproduction. Although steroid synthesis in endocrine cells requires uptake of cholesterol, the mechanisms regulating cholesterol uptake, storage and availability for steroid production are poorly understood. We have performed a genome-wide in vivo RNAi screen to uncover the genes required for steroid production in the endocrine steroid-producing cells of Drosophila. Using this approach, we reduced the expression of 12,600 genes in the steroid producing prothoracic gland (PG) cells to investigate their potential role in steroid biosynthesis.

Several genes previously shown to be required for delivery of cholesterol for steroidogenesis were identified including genes involved in the Niemann pick type C disease, scavenger receptors and lipid droplet proteins. Cholesterol is a low density lipoprotein (LDL)-derived lipid, taken up by cells through endocytosis and stored as cholesteryl ester in lipid droplets. To identify novel genes involved in cholesterol uptake and transportation, we performed a secondary screen to analyze which of genes identified in our primary screen for steroidogenesis that are specifically involved in cholesterol trafficking. In this screen we identified 20 novel genes that seem to be involved in regulating cholesterol uptake and trafficking for steroid production.

Loss of genes identified in our screen caused defects in formation of cholesterol-rich lipid droplets indicating that the genes are involved in uptake, trafficking and storage of cholesterol. Among the pathways identified, we show that genes involved in fatty acid elongation and sphingolipid biosynthesis are required for endosomal trafficking of cholesterol, which is coordinated by TOR and depends on autophagosomal trafficking. Many of these novel genes have conserved human homologs that have been associated with diseases that involve dysregulation of cholesterol homeostasis and steroid signaling.