The R-SNARE endobrevin/VAMP-8 mediates homotypic fusion of early endosomes and late endosomes.
Endobrevin/VAMP-8 is an R-SNARE localized to endosomes, but it is unknown in which intracellular fusion step it operates. Using subcellular fractionation and quantitative immunogold electron microscopy, we found that endobrevin/VAMP-8 is present on all membranes known to communicate with early endosomes, including the plasma membrane, clathrin-coated pits, late endosomes, and membranes of the trans-Golgi network. Affinity-purified antibodies that block the ability of endobrevin/VAMP-8 to form SNARE core complexes potently inhibit homotypic fusion of both early and late endosomes in vitro. Fab fragments were as active as intact immunoglobulin Gs. Recombinant endobrevin/VAMP-8 inhibited both fusion reactions with similar potency. We conclude that endobrevin/VAMP-8 operates as an R-SNARE in the homotypic fusion of early and late endosomes.
Antonin
W
W
Department of Neurobiology, Max-Planck-Institute for Biophysical Chemistry, Göttingen, Germany.
Holroyd
C
C
Tikkanen
R
R
Höning
S
S
Jahn
R
R
D000818:Animals; D002458:Cell Fractionation; D002462:Cell Membrane / Q000502:physiology / Q000648:ultrastructure; D002474:Cell-Free System; D003034:Coated Pits, Cell-Membrane / Q000502:physiology / Q000648:ultrastructure; D011992:Endosomes / Q000502:physiology* / Q000648:ultrastructure; D008561:Membrane Fusion / Q000502:physiology*; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology*; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; D051381:Rats; D050600:SNARE Proteins; D016708:Synaptophysin / Q000502:physiology; D033921:Vesicular Transport Proteins; D021601:trans-Golgi Network / Q000502:physiology / Q000648:ultrastructure
D008565:Membrane Proteins; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; D050600:SNARE Proteins; D016708:Synaptophysin; C491830:Vamp8 protein, rat; D033921:Vesicular Transport Proteins