Oligomeric complexes link Rab5 effectors with NSF and drive membrane fusion via interactions between EEA1 and syntaxin 13.
SNAREs and Rab GTPases cooperate in vesicle transport through a mechanism yet poorly understood. We now demonstrate that the Rab5 effectors EEA1 and Rabaptin-5/Rabex-5 exist on the membrane in high molecular weight oligomers, which also contain NSF. Oligomeric assembly is modulated by the ATPase activity of NSF. Syntaxin 13, the t-SNARE required for endosome fusion, is transiently incorporated into the large oligomers via direct interactions with EEA1. This interaction is required to drive fusion, since both dominant-negative EEA1 and synthetic peptides encoding the FYVE Zn2+ finger hinder the interaction and block fusion. We propose a novel mechanism whereby oligomeric EEA1 and NSF mediate the local activation of syntaxin 13 upon membrane tethering and, by analogy with viral fusion proteins, coordinate the assembly of a fusion pore.
McBride
H M
HM
European Molecular Biology Laboratory, Heidelberg, Germany.
Rybin
V
V
Murphy
C
C
Giner
A
A
Teasdale
R
R
Zerial
M
M
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000595:Amino Acid Sequence; D001324:Autoantigens / Q000378:metabolism; D015374:Biosensing Techniques; D002352:Carrier Proteins / Q000378:metabolism*; D011992:Endosomes / Q000187:drug effects / Q000502:physiology; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D008561:Membrane Fusion / Q000187:drug effects / Q000502:physiology*; D008565:Membrane Proteins / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D052066:N-Ethylmaleimide-Sensitive Proteins; D009842:Oligopeptides / Q000737:chemistry / Q000494:pharmacology; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology; D050765:Qa-SNARE Proteins; D050600:SNARE Proteins; D033921:Vesicular Transport Proteins; D016335:Zinc Fingers; D020696:rab5 GTP-Binding Proteins
D001324:Autoantigens; D002352:Carrier Proteins; D008565:Membrane Proteins; D009842:Oligopeptides; D010446:Peptide Fragments; D050765:Qa-SNARE Proteins; D050600:SNARE Proteins; D033921:Vesicular Transport Proteins; C093846:early endosome antigen 1; D000251:Adenosine Triphosphatases; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D052066:N-Ethylmaleimide-Sensitive Proteins; D020696:rab5 GTP-Binding Proteins