A Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis

Benoît Hastoy, Pier A. Scotti, Alexandra Milochau, Zahia Fezoua-Boubegtiten, Jorge Rodas, Rémi Megret, Bernard Desbat, Michel Laguerre, Sabine Castano, David Perrais, Patrik Rorsman, Reiko Oda (+1 others)
2017 Scientific Reports  
Exocytosis depends on cytosolic domains of SNARE proteins but the function of the transmembrane domains (TMDs) in membrane fusion remains controversial. The TMD of the SNARE protein synaptobrevin2/VAMP2 contains two highly conserved small amino acids, G 100 and C 103 , in its central portion. Substituting G 100 and/or C 103 with the β-branched amino acid valine impairs the structural flexibility of the TMD in terms of α-helix/β-sheet transitions in model membranes (measured by infrared
more » ... n-absorption or evanescent wave spectroscopy) during increase in protein/lipid ratios, a parameter expected to be altered by recruitment of SNAREs at fusion sites. This structural change is accompanied by reduced membrane fluidity (measured by infrared ellipsometry). The G 100 V/C 103 V mutation nearly abolishes depolarization-evoked exocytosis (measured by membrane capacitance) and hormone secretion (measured biochemically). Single-vesicle optical (by TIRF microscopy) and biophysical measurements of ATP release indicate that G 100 V/C 103 V retards initial fusion-pore opening, hinders its expansion and leads to premature closure in most instances. We conclude that the TMD of VAMP2 plays a critical role in membrane fusion and that the structural mobility provided by the central small amino acids is crucial for exocytosis by influencing the molecular re-arrangements of the lipid membrane that are necessary for fusion pore opening and expansion. Published: xx xx xxxx OPEN www.nature.com/scientificreports/ 2 Scientific RepoRts | 7: 2835 |
doi:10.1038/s41598-017-03013-3 pmid:28588281 pmcid:PMC5460238 fatcat:c7gzc4anjzgz3pzagrexgkhsza