Vesicle budding caused by lysolipid-induced asymmetry stress.

Abstract

The lipid membrane undergoes continuous and dynamic changes in a multitude of cellular processes. Exo- and endovesiculation, asexual reproduction or the forming of organelles require a change in local curvature. Although many protein-centered mechanisms have been identified to induce these changes, the properties of the lipid membrane itself have been neglected in most cases. Changes in lipid composition and asymmetry can also contribute to membrane curvature generation. Lysolipids dissolved or dispersed in the outer medium insert into the accessible membrane leaflet and expand its area but show only very slow translocation to the trans leaflet. This resulting asymmetry stress is partially relaxed by vesicle budding even in the absence of proteins. Budding induced by lysolipids have been studied with giant unilamellar vesicles (GUVs), but they lack a quantitative perspective, since bulk measurements are difficult to implement using GUVs. Using Asymmetric Flow Field-Flow Fractionation (AF4), we were able to establish an assay quantifying the fraction of lipid budding off from large unilamellar vesicles (LUVs) depending on specific conditions. We discovered that the extent of budding depends, above a threshold, not on the amount of lysolipid added. Instead, it is limited by the excess surface area stored in the initial LUV. As the vesicle becomes ideally spherical, a hypotonic vesicle interior opposes further budding and further lysolipid is prevented from entering the membrane. Our studies provide a detailed picture how lysolipids and other membrane-impermeant amphiphiles contribute to membrane curvature generation. Furthermore, it provides insight about alternative mechanisms of a membrane to relax asymmetry stress, including transient membrane leakage as a mode of action of antibiotic lipopeptides.