Developmentally regulated sphingolipid synthesis in African trypanosomes.
Sphingolipids are essential components of eukaryotic membranes, and many unicellular eukaryotes, including kinetoplastid protozoa, are thought to synthesize exclusively inositol phosphorylceramide (IPC). Here we characterize sphingolipids from Trypanosoma brucei, and a trypanosome sphingolipid synthase gene family (TbSLS1-4) that is orthologous to Leishmania IPC synthase. Procyclic trypanosomes contain IPC, but also sphingomyelin, while surprisingly bloodstream-stage parasites contain sphingomyelin and ethanolamine phosphorylceramide (EPC), but no detectable IPC. In vivo fluorescent ceramide labelling confirmed stage-specific biosynthesis of both sphingomyelin and IPC. Expression of TbSLS4 in Leishmania resulted in production of sphingomyelin and EPC suggesting that the TbSLS gene family has bi-functional synthase activity. RNAi silencing of TbSLS1-4 in bloodstream trypanosomes led to rapid growth arrest and eventual cell death. Ceramide levels were increased more than threefold by silencing suggesting a toxic downstream effect mediated by this potent intracellular messenger. Topology predictions support a revised six-transmembrane domain model for the kinetoplastid sphingolipid synthases consistent with the proposed mammalian sphingomyelin synthase structure. This work reveals novel diversity and regulation in sphingolipid metabolism in this important group of human parasites.
Sutterwala
Shaheen S
SS
Department of Medical Microbiology and Immunology, University of Wisconsin School of Medicine and Public Health, 1550 Linden Drive, Madison, WI 53706, USA.
Hsu
Fong-Fu
FF
Sevova
Elitza S
ES
Schwartz
Kevin J
KJ
Zhang
Kai
K
Key
Phillip
P
Turk
John
J
Beverley
Stephen M
SM
Bangs
James D
JD
D000349:Africa; D000595:Amino Acid Sequence; D000818:Animals; D015870:Gene Expression; D020868:Gene Silencing; D006028:Glycosphingolipids / Q000302:isolation & purification; D006801:Humans; D007891:Leishmania / Q000235:genetics; D008025:Ligases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D008954:Models, Biological; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015800:Protozoan Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013107:Sphingolipids / Q000096:biosynthesis*; D013109:Sphingomyelins / Q000302:isolation & purification; D014346:Trypanosoma brucei brucei / Q000737:chemistry* / Q000302:isolation & purification / Q000502:physiology*
D006028:Glycosphingolipids; D015800:Protozoan Proteins; D013107:Sphingolipids; D013109:Sphingomyelins; C025524:inositolphosphoceramides; C060281:ceramide phosphoethanolamine; D008025:Ligases