Reconstitution of Rab- and SNARE-dependent membrane fusion by synthetic endosomes

Nature. 2009 Jun 25;459(7250):1091-7. doi: 10.1038/nature08107. Epub 2009 May 20.


Rab GTPases and SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) are evolutionarily conserved essential components of the eukaryotic intracellular transport system. Although pairing of cognate SNAREs is sufficient to fuse membranes in vitro, a complete reconstitution of the Rab-SNARE machinery has never been achieved. Here we report the reconstitution of the early endosomal canine Rab5 GTPase, its key regulators and effectors together with SNAREs into proteoliposomes using a set of 17 recombinant human proteins. These vesicles behave like minimal 'synthetic' endosomes, fusing with purified early endosomes or with each other in vitro. Membrane fusion measured by content-mixing and morphological assays requires the cooperativity between Rab5 effectors and cognate SNAREs which, together, form a more efficient 'core machinery' than SNAREs alone. In reconstituting a fusion mechanism dependent on both a Rab GTPase and SNAREs, our work shows that the two machineries act coordinately to increase the specificity and efficiency of the membrane tethering and fusion process.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Cricetinae
  • Cytosol / metabolism
  • Dogs
  • Endosomes / metabolism
  • Endosomes / physiology*
  • Humans
  • Membrane Fusion / physiology*
  • Microscopy, Electron
  • Proteolipids / metabolism
  • Proteolipids / ultrastructure
  • Recombinant Proteins / metabolism
  • SNARE Proteins / metabolism*
  • Vesicular Transport Proteins / metabolism
  • rab GTP-Binding Proteins / metabolism*


  • Proteolipids
  • Recombinant Proteins
  • SNARE Proteins
  • Vesicular Transport Proteins
  • proteoliposomes
  • rab GTP-Binding Proteins