Drosophila glucosylceramide synthase: a negative regulator of cell death mediated by proapoptotic factors

Glucosylceramide synthase (GlcT-1) catalyzes the formation of glucosylceramide (GlcCer), the core structure of major glycosphingolipids (GSLs), from ceramide and UDP-glucose. Ceramide and its metabolites, such as sphingosine-1-phosphate, are now known to be important mediators of apoptosis and cell survival. Recently, we have shown that GlcT-1 functions to regulate intracellular ceramide levels via glycosylation of ceramide. In this study, we employ the fruit fly Drosophila melanogaster as a model system for understanding the in vivo roles of GlcT-1. We isolated and characterized a GlcT-1 homologue (DGlcT-1) from Drosophila. When DGlcT-1 was expressed in GM-95 cells deficient in GSLs (because of the absence of GlcT-1 activity), these cells regained the ability to synthesize GSLs. Northern blot and in situ hybridization analyses revealed that the expression of DGlcT-1 mRNA was ubiquitous throughout development, suggesting that DGlcT-1 is important for development and differentiation. Indeed, RNA interference experiments demonstrated that the loss of GlcT-1 function enhances apoptotic cell death. Conversely, targeted expression of GlcT-1 partially rescued cell death caused by the proapoptotic factors Reaper and Grim, suggesting that ceramide generation might be one signal pathway that executes the cell death program. We also found that GlcT-1 localized not only in the Golgi apparatus but also in the perinuclear endoplasmic reticulum, providing the first visual evidence of GlcT-1 in membranes. These results indicate that GlcT-1 might down-regulate ceramide generated in these membranes.