Lipid cosorting mediated by shiga toxin induced tubulation.

To maintain cell membrane homeostasis, lipids must bedynamically redistributed during the formation of trans-port intermediates, but the mechanisms driving lipidsorting are not yet fully understood. Lowering sphin-golipid concentration can reduce the bending energy of amembrane, and this effect could account for sphingolipiddepletion along the retrograde pathway. However, sph-ingolipids and cholesterol are enriched along the antero-grade pathway, implying that other lipid sorting mecha-nisms, such as protein-mediated sorting, can dominate.To characterize the influence of protein binding on thelipid composition of highly curved membranes, we stud-ied the interactions of the B-subunit of Shiga toxin (STxB)with giant unilamellar vesicles containing its glycosph-ingolipid receptor [globotriaosylceramide (Gb3)]. STxBbinding induced the formation of tubular membraneinvaginations, and fluorescence microscopy images ofthese highly curved membranes were consistent withco-enrichment of Gb3 and sphingolipids. In agreementwith theory, sorting was stronger for membrane compo-sitions close to demixing. These results strongly supportthe hypothesis that proteins can indirectly mediate thesorting of lipids into highly curved transport intermedi-ates via interactions between lipids and the membranereceptor of the protein.Key words: curvature, Gb3, GM1, Laurdan, lipid sorting,model membrane, Shiga toxin, sphingolipidReceived 6 April 2010, revised and accepted forpublication 30 August 2010, uncorrected manuscriptpublished online 7 September 2010, published online1 October 2010