Resolving the function of distinct Munc18-1/SNARE protein interaction modes in a reconstituted membrane fusion assay

J Biol Chem. 2011 Sep 2;286(35):30582-30590. doi: 10.1074/jbc.M111.269886. Epub 2011 Jul 5.


Sec1p/Munc18 proteins and SNAP receptors (SNAREs) are key components of the intracellular membrane fusion machinery. Compartment-specific v-SNAREs on a transport vesicle pair with their cognate t-SNAREs on the target membrane and drive lipid bilayer fusion. In a reconstituted assay that dissects the sequential assembly of t-SNARE (syntaxin 1·SNAP-25) and v-/t-SNARE (VAMP2·syntaxin 1·SNAP-25) complexes, and finally measures lipid bilayer merger, we resolved the inhibitory and stimulatory functions of the Sec1p/Munc18 protein Munc18-1 at the molecular level. Inhibition of membrane fusion by Munc18-1 requires a closed conformation of syntaxin 1. Remarkably, the concurrent preincubation of Munc18-1-inhibited syntaxin 1 liposomes with both VAMP2 liposomes and SNAP-25 at low temperature releases the inhibition and effectively stimulates membrane fusion. VAMP8 liposomes can neither release the inhibition nor exert the stimulatory effect, demonstrating the need for a specific Munc18-1/VAMP2 interaction. In addition, Munc18-1 binds to the N-terminal peptide of syntaxin 1, which is obligatory for a robust stimulation of membrane fusion. In contrast, this interaction is neither required for the inhibitory function of Munc18-1 nor for the release of this block. These results indicate that Munc18-1 and the neuronal SNAREs already have the inherent capability to function as a basic stage-specific off/on switch to control membrane fusion.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • DNA / chemistry
  • Lipids / chemistry
  • Liposomes / chemistry
  • Liposomes / metabolism
  • Membrane Fusion / physiology*
  • Models, Biological
  • Munc18 Proteins / metabolism*
  • Peptides / chemistry
  • Protein Binding
  • Protein Conformation
  • Protein Interaction Mapping
  • Rats
  • SNARE Proteins / chemistry*
  • Temperature


  • Lipids
  • Liposomes
  • Munc18 Proteins
  • Peptides
  • SNARE Proteins
  • DNA