Light-controlled membrane remodeling in gel-fluid phase-separated giant vesicles using photoswitchable lipids

Photoswitchable lipids enable optical control of membrane area, mechanics and phase behavior, offering a platform to study stimuli-responsive biomimetic systems. However, it remains unclear how the spatial organization of coexisting membrane phases governs photoinduced mechanical responses. Here, we incorporate the photoswitch azobenzene-phosphatidylcholine (azoPC) and dipalmitoyphosphatidylcholine (DPPC) into gel-fluid phase-separated giant unilamellar vesicles (GUVs) to probe how domain architecture controls light-driven deformation. We combine differential scanning calorimetry, temperature-controlled confocal microscopy, and a calibrated heating stage to quantify phase transitions and visualize domain dynamics. Dispersed domains produce global GUV crumpling upon UV-light-induced trans -to- cis isomerization of azoPC, whereas coarsened fluid domains locally confine deformation to budding regions of the GUVs; both responses are reversed by blue light. Temperature-controlled imaging reveals that the gel-fluid transition in GUVs is considerably broader than the calorimetric profile suggests, with coexisting phases detectable well above the calorimetry peak transition temperature. Well above the transition temperature, i.e. in the fully melted membrane, UV irradiation unexpectedly induces reversible nucleation of gel-like flower domains, consistent with an increased transition temperature in the cis azoPC state due to lipid packing incompatibility with DPPC. These results demonstrate that membrane domain architecture dictates the spatial distribution of photoinduced remodeling and that photoswitchable lipids can tune both membrane morphology and phase equilibria, enabling new strategies for stimuli-responsive synthetic cells and soft actuators.