Abstract
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.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adenosine Triphosphatases / metabolism
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Amino Acid Sequence
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Autoantigens / metabolism
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Biosensing Techniques
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Carrier Proteins / metabolism*
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Endosomes / drug effects
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Endosomes / physiology
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GTP Phosphohydrolases / metabolism
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GTP-Binding Proteins / metabolism*
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HeLa Cells
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Humans
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Intracellular Membranes / drug effects
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Intracellular Membranes / physiology
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Intracellular Membranes / ultrastructure
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Membrane Fusion / drug effects
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Membrane Fusion / physiology*
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Membrane Proteins / metabolism*
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Models, Biological
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Molecular Sequence Data
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N-Ethylmaleimide-Sensitive Proteins
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Oligopeptides / chemistry
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Oligopeptides / pharmacology
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Peptide Fragments / chemistry
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Peptide Fragments / pharmacology
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Qa-SNARE Proteins
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SNARE Proteins
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Vesicular Transport Proteins*
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Zinc Fingers
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rab5 GTP-Binding Proteins
Substances
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Autoantigens
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Carrier Proteins
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Membrane Proteins
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Oligopeptides
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Peptide Fragments
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Qa-SNARE Proteins
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SNARE Proteins
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Vesicular Transport Proteins
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early endosome antigen 1
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Adenosine Triphosphatases
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GTP Phosphohydrolases
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GTP-Binding Proteins
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N-Ethylmaleimide-Sensitive Proteins
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rab5 GTP-Binding Proteins