Phase Specific Membrane Fusion with SNARE Mimetics

Abstract

Biological membrane fusion is involved in a number of essential processes such as neurotransmission, exocytosis, and viral infection. It is crucial for cells to spatially confine this process to specific organelles or sites in the plasma membrane. In nature, the SNARE complex induces fusion in eukaryotic cells. Its different constituents integrated into opposing membranes approximate the two bilayers by interacting in a zipper-like fashion. Different studies examined membrane fusion in artificial systems using SNARE mimetics. Fusion between pairs of small unilamellar vesicles have been shown using lipidated DNA [Chan et al., Biointerphases, 3:FA17, 2008] and peptide [Pahler et al., Biophysical Journal, 103:2295, 2012] ligand-receptor pairs. Here, we show strong evidence of the fusion of large unilamellar vesicles (LUVs) to a spatially confined region within liquid-liquid phase-separated giant unilamellar vesicles using the abovementioned (i) hybridised DNA and (ii) coiled-coil peptide pairs as SNARE mimetics. We quantified the (hemi-)fusion process using two different FRET-based approaches which yielded similar results. Moreover, our system potentially offers to determine LUV docking and fusion efficiencies. Complementary experiments are currently performed using reconstituted SNARE proteins (SNARE proteins were kindly provided by Claudia Steinem, Department of Biomolecular Chemistry at Gottingen University). This work is part of the MaxSynBio consortium which is jointly funded by the Federal Ministry of Education and Research of Germany and the Max Planck Society.