Datensatz

Zusammenfassung

Membrane asymmetry is ubiquitous in cell membranes particularly with respect to lipids, whereby charged lipids are mainly restricted to the inner monolayer. We investigate the influence of anionic lipid asymmetry on the stability of giant unilamellar vesicles (GUVs), minimal plasma membrane models. To quantify asymmetry, we apply a fluorescence quenching assay, which is often difficult to reproduce and caution in handling the quencher is generally underestimated. Thus, we first optimize this assay and then apply it to GUVs prepared with the inverted emulsion transfer protocol using increasing fractions of anionic lipids restricted to one leaflet. This protocol is found to produce highly asymmetric bilayers, but with ~20% interleaflet mixing. To probe the stability of asymmetric vs symmetric membranes, we expose the GUVs to porating DC pulses and monitor the fraction of destabilized vesicles. The pulses open macropores, and the GUVs either completely recover their integrity or become destabilized exhibiting leakage or bursting/collapse. Destabilization is much more pronounced in asymmetrically charged membranes, which is corroborated by pore edge tension data showing considerable decrease with asymmetry. Rendering GUV membrane asymmetric from exposure to different transmembrane pH, we confirm that poration-triggered destabilization does not depend on the approach used to generate membrane asymmetry.Competing Interest StatementThe authors have declared no competing interest.