title: Sphingolipid synthesis and scavenging in the intracellular apicomplexan parasite, Toxoplasma gondii.
abstract: Sphingolipids are essential components of eukaryotic cell membranes, particularly the plasma membrane, and are involved in a diverse array of signal transduction pathways. Mammals produce sphingomyelin (SM) as the primary complex sphingolipid via the well characterised SM synthase. In contrast yeast, plants and some protozoa utilise an evolutionarily related inositol phosphorylceramide (IPC) synthase to synthesise IPC. This activity has no mammalian equivalent and IPC synthase has been proposed as a target for anti-fungals and anti-protozoals. However, detailed knowledge of the sphingolipid biosynthetic pathway of the apicomplexan protozoan parasites was lacking. In this study bioinformatic analyses indicated a single copy orthologue of the putative SM synthase from the apicomplexan Plasmodium falciparum (the causative agent of malaria) was a bona fide sphingolipid synthase in the related model parasite, Toxoplasma gondii (TgSLS). Subsequently, TgSLS was indicated, by complementation of a mutant cell line, to be a functional orthologue of the yeast IPC synthase (AUR1p), demonstrating resistance to the well characterised AUR1p inhibitor aureobasidin A. In vitro, recombinant TgSLS exhibited IPC synthase activity and, for the first time, the presence of IPC was demonstrated in T. gondii lipid extracts by mass spectrometry. Furthermore, host sphingolipid biosynthesis was indicated to influence, but be non-essential for, T. gondii proliferation, suggesting that whilst scavenging does take place de novo sphingolipid synthesis may be important for parasitism.
authors.affiliation: Biophysical Sciences Institute, Department of Chemistry and School of Biological Sciences, University Science Laboratories, Durham DH1 3LE, UK.
authors.forename: Steven
authors.initials: S
authors.lastname: Pratt
authors.forename: Nilu K
authors.initials: NK
authors.lastname: Wansadhipathi-Kannangara
authors.forename: Catherine R
authors.initials: CR
authors.lastname: Bruce
authors.forename: John G
authors.initials: JG
authors.lastname: Mina
authors.forename: Hosam
authors.initials: H
authors.lastname: Shams-Eldin
authors.forename: Josefina
authors.initials: J
authors.lastname: Casas
authors.forename: Kentaro
authors.initials: K
authors.lastname: Hanada
authors.forename: Ralph T
authors.initials: RT
authors.lastname: Schwarz
authors.forename: Sabrina
authors.initials: S
authors.lastname: Sonda
authors.forename: Paul W
authors.initials: PW
authors.lastname: Denny
mesh_terms: D019295:Computational Biology; D017353:Gene Deletion; D005816:Genetic Complementation Test; D006602:Hexosyltransferases / Q000378:metabolism; D013107:Sphingolipids / Q000378:metabolism*; D014122:Toxoplasma / Q000378:metabolism*
chemical_list: D013107:Sphingolipids; D006602:Hexosyltransferases; C088714:phosphatidylinositol-ceramide phosphoinositol transferase