Colocalization on the same synaptic vesicles of syntaxin and SNAP-25 with synaptic vesicle proteins: a re-evaluation of functional models required?
Synaptic vesicle docking and calcium dependent exocytosis are thought to require the specific interaction of proteins of the synaptic vesicle membrane (such as VAMP/synaptobrevin and synaptotagmin) and their plasma membrane-located counterparts (such as syntaxin and SNAP-25). When isolating synaptic vesicles by glycerol velocity gradient centrifugation we found cosedimentation of the presumptive presynaptic plasma membrane proteins syntaxin and SNAP-25 with synaptic vesicle membrane proteins. In order to further identify the antibody binding organelles we performed an immunoelectron microscopical analysis of synaptosomal profiles. Syntaxin and SNAP-25 were not only associated with the plasma membrane but to a large extent also with synaptic vesicle profiles. In order to answer the question whether the syntaxin and SNAP-25 containing vesicular compartment would also carry classical synaptic vesicle membrane markers we performed double labeling experiments using poly- and monoclonal antibodies. We found colocalization on the same vesicle not only of SNAP-25 and syntaxin but also of SNAP-25 with the synaptic vesicle membrane proteins SV2 and synaptotagmin and of syntaxin with the vesicular membrane protein synaptophysin. Our results demonstrate that syntaxin and SNAP-25 are colocalized with classical vesicle membrane proteins on the same vesicle and suggest that the functional models for the interaction of presynaptic proteins need to be re-evaluated.
Kretzschmar
S
S
AK Neurochemie, Zoologisches Institut der J.W. Goethe-Universität, Biozentrum der J.W. Goethe-Universität, Frankfurt am Main, Germany.
Volknandt
W
W
Zimmermann
H
H
D000818:Animals; D001923:Brain Chemistry; D015151:Immunoblotting; D008565:Membrane Proteins / Q000032:analysis*; D016253:Microscopy, Immunoelectron; D009419:Nerve Tissue Proteins / Q000032:analysis*; D050765:Qa-SNARE Proteins; D051381:Rats; D013572:Synaptic Vesicles / Q000737:chemistry*; D016708:Synaptophysin / Q000032:analysis; D050825:Synaptosomal-Associated Protein 25
D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D050765:Qa-SNARE Proteins; C494210:Snap25 protein, rat; D016708:Synaptophysin; D050825:Synaptosomal-Associated Protein 25