Self-interaction of a SNARE transmembrane domain promotes the hemifusion-to-fusion transition.
SNARE proteins mediate intracellular fusion of eukaryotic membranes. Some SNAREs have previously been shown to dimerise via interaction of their transmembrane domains. However, the functional significance of these interactions had remained unclear. Here, we show that mutating alternate faces of the transmembrane helix of the yeast vacuolar Q-SNARE Vam3p reduces the ability of the full-length protein to induce contents mixing in yeast vacuole fusion to different extents. Examination of liposome fusion induced by synthetic transmembrane domains revealed that inner leaflet mixing is delayed relative to outer leaflet mixing, suggesting that fusion transits through a hemifusion intermediate. Interestingly, one of the mutations impaired inner leaflet mixing in the liposome system. This suggests that the defect seen in vacuolar contents mixing is due to partial arrest of the reaction at hemifusion. Since covalent dimerisation of this mutant recovered wild-type behaviour, homodimerisation of a SNARE transmembrane domain appears to control the transition of a hemifusion intermediate to complete lipid mixing.
Hofmann
Mathias W
MW
Lehrstuhl Chemie der Biopolymere, Technische Universität München, Weihenstephaner Berg 3, 85354 Freising, Germany.
Peplowska
Karolina
K
Rohde
Jan
J
Poschner
Bernhard C
BC
Ungermann
Christian
C
Langosch
Dieter
D
D002942:Circular Dichroism; D003432:Cross-Linking Reagents; D007425:Intracellular Membranes / Q000378:metabolism*; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008561:Membrane Fusion; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D050765:Qa-SNARE Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization; D014617:Vacuoles / Q000737:chemistry / Q000378:metabolism
D003432:Cross-Linking Reagents; D008051:Lipid Bilayers; D008081:Liposomes; D010446:Peptide Fragments; D050765:Qa-SNARE Proteins; D029701:Saccharomyces cerevisiae Proteins