Lipid Cosorting Mediated by Shiga Toxin Induced Tubulation

Abstract

† These authors contributed equally to this work. ‡ These authors contributed equally to this work.To maintain cell membrane homeostasis, lipids must be dynamically redistributed during the formation of transport intermediates, but the mechanisms driving lipid sorting are not yet fully understood. Lowering sphingolipid concentration can reduce the bending energy of a membrane, and this effect could account for sphingolipid depletion along the retrograde pathway. However, sphingolipids and cholesterol are enriched along the anterograde pathway, implying that other lipid sorting mechanisms, such as protein-mediated sorting, can dominate. To characterize the influence of protein binding on the lipid composition of highly curved membranes, we studied the interactions of the B-subunit of Shiga toxin (STxB) with giant unilamellar vesicles containing its glycosphingolipid receptor [globotriaosylceramide (Gb3)]. STxB binding induced the formation of tubular membrane invaginations, and fluorescence microscopy images of these highly curved membranes were consistent with co-enrichment of Gb3 and sphingolipids. In agreement with theory, sorting was stronger for membrane compositions close to demixing. These results strongly support the hypothesis that proteins can indirectly mediate the sorting of lipids into highly curved transport intermediates via interactions between lipids and the membrane receptor of the protein.

Polarized sorting and trafficking in epithelial cells

“…The growing curvature of a membrane close to the demixing point (phase separation) further induces lipid sorting based solely on their underlying connectivity, which is greatly amplified by their clustering [111]. Since curvature of a membrane can also drive protein sorting, a growing bud can generate a feedback system whereby curvature-preferring proteins would be recruited to a growing lipid raft platform, further increasing the propensity to generate curvature [112][113][114][115]. Once a curved membrane is generated, phase separation in membrane tubes can trigger membrane fission arising from the difference in elastic constants between the domains [115,116].…”

Curvature-Driven Lipid Sorting in Biomembranes

“…Confocal microscopy of membrane nanotubes has also revealed the sorting of the curvaturesensitive GTP-ase ArfGap1 (Ambroggio et al 2010). In addition, cholera (Sorre et al 2009;) and Shiga toxins (Safouane et al 2010) bound to DOPC membranes with small amounts of the toxin lipid receptor have been observed to be strongly excluded from tubes. Both types of toxin prefer negative curvature, whereas tubes have positive curvature, and hence on energetic grounds should be excluded from tubes when bound to the outer leaflet.…”

“…Generally, clustering a certain type of lipid by a protein or even actin filaments can induce phase separation in a homogeneous membrane even if this component is present in a very small quantity (Hammond et al 2005;Liu and Fletcher 2006;Römer et al 2010) and (Safouane et al 2010). The effects of protein on the phase behavior of lipid membrane are an interesting phenomenon and can be understood as follows.…”

“…We discuss briefly how protein may play a key role in concentrating sphingolipids into highly curved transport intermediates. In Safouane et al (2010), tubular invaginations were induced on GUVs by the binding of the B-subunit of Shiga toxin to its lipid receptor Gb3. Using different fluorophores, it was found that the concentration of sphingolipids in the tubes increased as the membrane composition approached that of phase separation.…”

Bending "On the Rocks"--A Cocktail of Biophysical Modules to Build Endocytic Pathways

“…This is explained by lipid -lipid interactions that become dominant over mixing entropy as membranes approach lipiddemixing conditions. Of interest, GSLs can be cosorted under such conditions (Safouane et al 2010), which may contribute to the generation of specific compositional environments that are programmed to carry out downstream functions. A striking example is the above-mentioned actin-driven scission process on Shiga toxin-induced membrane invaginations (Rö-mer et al 2010).…”