Biochem. J. (2008) 415
(87–96) (Printed in Great Britain)
Lipidomic analysis of Toxoplasma gondii tachyzoites rhoptries: further insights into the role of cholesterol
Sébastien BESTEIRO*†1, Justine BERTRAND-MICHEL‡, Maryse LEBRUN*†, Henri VIAL*† and Jean-François DUBREMETZ*
*Laboratoire de Dynamique des Interactions Membranaires Normales et Pathologiques (DIMNP), Universités de Montpellier II et I, Centre National de la Recherche Scientifique (CNRS), UMR 5235, Montpellier, F-34095, France, †INSERM, DIMNP, Montpellier, France, and ‡INSERM, Institut Claude de Préval, IFR30, Plateau technique de Lipidomique, Toulouse, F-31300, France
Rhoptries are secretory organelles involved in the virulence of the human pathogen Toxoplasma gondii. In the present study we have used HPLC and capillary GLC to isolate and quantify lipids from whole Toxoplasma cells and their purified rhoptries. This comparative lipidomic analysis revealed an enrichment of cholesterol, sphingomyelin and, most of all, saturated fatty acids in the rhoptries. These lipids are known, when present in membranes, to contribute to their rigidity and, interestingly, fluorescence anisotropy measurements confirmed that rhoptry-derived membranes have a lower fluidity than membranes from whole T. gondii cells. Moreover, although rhoptries were initially thought to be highly enriched in cholesterol, we demonstrated that cholesterol is present in lower proportions, and we have provided additional evidence towards a lack of involvement of rhoptry cholesterol in the process of host-cell invasion by the parasite. Indeed, depleting the cholesterol content of the parasites did not prevent the secretion of protein-containing rhoptry-derived vesicles and the parasites could still establish a structure called the moving junction, which is necessary for invasion. Instead, the crucial role of host cholesterol for invasion, which has already been demonstrated [Coppens and Joiner (2003) Mol. Biol. Cell 14, 3804–3820], might be explained by the need of a cholesterol-rich region of the host cell we could visualize at the point of contact with the attached parasite, in conditions where parasite motility was blocked.
Key words: cholesterol, invasion, lipidomic analysis, raft, rhoptry, Toxoplasma gondii.
Abbreviations used: CE, cholesterol ester; CytD, cytochalasin D; DAG, diacylglycerol; DMEM, Dulbecco's modified Eagle's medium; DPH, 1,6-diphenyl-1,3,5-hexatriene; FAME, fatty acid methyl ester; FBS, fetal bovine serum; GPI, glycerophosphatidylinositol; HFF, human foreskin fibroblast; i.d., internal diameter, LPL, lysophospholipid; MJ, moving junction; MUFA, monounsaturated fatty acid; NL, neutral lipid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PI, phosphatidylinositol; PL, phospholipid; PS, phosphatidylserine; PUFA, polyunsaturated fatty acid; PV, parasitophorous vacuole; PVM, PV membrane; SAFA, saturated fatty acid; SM, sphingomyelin; TAG, triacylglycerol; UFA, unsaturated fatty acid.
1To whom correspondence should be addressed (email sebastien.besteiro@univ-montp2.fr).
Received 17 April 2008/9 June 2008; accepted 16 June 2008
Published as BJ Immediate Publication 16 June 2008, doi:10.1042/BJ20080795
© The Authors Journal compilation © 2008 Biochemical Society
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