Abstract
Membrane fusion is the core process in membrane trafficking and is essential for cellular transport of proteins and other biomacromolecules. During protein-mediated membrane fusion, membrane proteins are often excluded from the membrane–membrane contact, indicating that local structural transformations in lipid domains play a major role. However, the rearrangements of lipid domains during fusion have not been thoroughly examined. Here using a newly developed Fluorescence Surface Forces Apparatus (FL-SFA), migration of liquid-disordered clusters and depletion of liquid-ordered domains at the membrane–membrane contact are imaged in real time during hemifusion of model lipid membranes, together with simultaneous force–distance and lipid membrane thickness measurements. The load and contact time-dependent hemifusion results show that the domain rearrangements decrease the energy barrier to fusion, illustrating the significance of dynamic domain transformations in membrane fusion processes. Importantly, the FL-SFA can unambiguously correlate interaction forces and in situ imaging in many dynamic interfacial systems.
Highlights
Membrane fusion is the core process in membrane trafficking and is essential for cellular transport of proteins and other biomacromolecules
Previous studies on combined lipid and protein systems show that lipid domains localize SNARE proteins[2,3,9], and the formation of lipid domain/SNARE complexes is essential for lowering the energy barrier to fusion[10]
Using a custom-built Fluorescence Surface Forces Apparatus (FL-SFA, see Methods section and Fig. 1 for a detailed description of the set-up), we measured the interaction forces between supported lipid membranes and simultaneously imaged lipid domains during pressure-induced and protein-free hemifusion, allowing for real-time correlations to be made between the interaction forces, membrane thickness and spatial and temporal domain rearrangements
Summary
Membrane fusion is the core process in membrane trafficking and is essential for cellular transport of proteins and other biomacromolecules. The involvement of lipid domains during biological fusion processes is well established, their dynamic rearrangements during fusion are yet to be elucidated Such domains are seen in reconstituted myelin lipid bilayers extracted from the brain, for example, where the domains are observed to be different in healthy versus pathological (for example, multiple sclerosis) membranes[16]. In this case, as in many other cellular structures, the membranes are planar and closely stacked in vivo and strongly interacting with each other across the water spaces. The results on the model membranes provide mechanistic details of domain rearrangements during membrane fusion, demonstrating that the FL-SFA should find wide utility in correlating fluorescent images with interaction forces during compression and separation of a broad range of materials between confined surfaces
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