This article is available online at http://www.jlr.org leafl et of the plasma membrane (PM) where its high packing density and affi nity for cholesterol contributes to the barrier function of the organelle ( 1 ). SM and cholesterol form concentration gradients along the exocytic pathway that are thought to affect protein sorting through hydrophobic matching of membrane spans ( 2, 3 ). Moreover, SM can be hydrolyzed by neutral SMases at the PM to form phosphocholine and the proapoptotic signaling molecule ceramide. This enzymatic reaction is part of an antiproliferative, sphingolipid-mediated signal transduction pathway that controls cell cycle arrest, differentiation, and apoptosis in response to growth factor deprivation, cytokines, ionizing radiation, heat, and chemotherapy ( 4 ). SM synthesis is mediated by a phosphatidylcholine (PC):ceramide cholinephosphotransferase or SM synthase (EC 2.7.8.27). This enzyme catalyzes the transfer of phosphocholine from PC onto ceramide, yielding SM and diacylglycerol ( 5 ). Mammalian cells also produce small amounts of the SM analog ceramide phosphoethanolamine (CPE). However, very little is known about the biological role of this analog or about the enzyme(s) responsible for its production. Two CPE synthase activities have been described in mammalian cells, one enriched in a microsomal fraction [presumably endoplasmic reticulum (ER)] and the other one associated with the PM ( 6-9 ). As PE serves as the headgroup donor for both activities, the enzyme(s) involved can be classifi ed as PE:ceramide Abstract Sphingolipids are vital components of eukaryotic membranes involved in the regulation of cell growth, death, intracellular traffi cking, and the barrier function of the plasma membrane (PM). While sphingomyelin (SM) is the major sphingolipid in mammals, previous studies indicate that mammalian cells also produce the SM analog ceramide phosphoethanolamine (CPE). Little is known about the biological role of CPE or the enzyme(s) responsible for CPE biosynthesis. SM production is mediated by the SM synthases SMS1 in the Golgi and SMS2 at the PM, while a closely related enzyme, SMSr, has an unknown biochemical function. We now demonstrate that SMS family members display striking differences in substrate specifi city, with SMS1 and SMSr being monofunctional enzymes with SM and CPE synthase activity, respectively, and SMS2 acting as a bifunctional enzyme with both SM and CPE synthase activity. In agreement with the PM residency of SMS2, we show that both SM and CPE synthase activities are enhanced at the surface of SMS2-overexpressing HeLa cells. Our fi ndings reveal an unexpected diversity in substrate specifi city among SMS family members that should enable the design of specifi c inhibitors to target the biological role of each enzyme individually.-Ternes, P., J. EMBO Long-Term Fellowship (to P.T.) (to J.C.M.H.).
and by grants from the Dutch Organization of Sciences (NWO-CW) and the Utrecht University High Potential Program