The synaptic vesicle cycle: a single vesicle budding step involving clathrin and dynamin.
Strong evidence implicates clathrin-coated vesicles and endosome-like vacuoles in the reformation of synaptic vesicles after exocytosis, and it is generally assumed that these vacuoles represent a traffic station downstream from clathrin-coated vesicles. To gain insight into the mechanisms of synaptic vesicle budding from endosome-like intermediates, lysed nerve terminals and nerve terminal membrane subfractions were examined by EM after incubations with GTP gamma S. Numerous clathrin-coated budding intermediates that were positive for AP2 and AP180 immunoreactivity and often collared by a dynamin ring were seen. These were present not only on the plasma membrane (Takei, K., P.S. McPherson, S.L.Schmid, and P. De Camilli. 1995. Nature (Lond.). 374:186-190), but also on internal vacuoles. The lumen of these vacuoles retained extracellular tracers and was therefore functionally segregated from the extracellular medium, although narrow connections between their membranes and the plasmalemma were sometimes visible by serial sectioning. Similar observations were made in intact cultured hippocampal neurons exposed to high K+ stimulation. Coated vesicle buds were generally in the same size range of synaptic vesicles and positive for the synaptic vesicle protein synaptotagmin. Based on these results, we suggest that endosome-like intermediates of nerve terminals originate by bulk uptake of the plasma membrane and that clathrin- and dynamin-mediated budding takes place in parallel from the plasmalemma and from these internal membranes. We propose a synaptic vesicle recycling model that involves a single vesicle budding step mediated by clathrin and dynamin.
Takei
K
K
Department of Cell Biology, Yale University School of Medicine, New Haven, Connecticut 06510, USA.
Mundigl
O
O
Daniell
L
L
De Camilli
P
P
D033962:Adaptor Protein Complex 2; D033965:Adaptor Protein Complex alpha Subunits; D033942:Adaptor Proteins, Vesicular Transport; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002135:Calcium-Binding Proteins; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D002478:Cells, Cultured; D002966:Clathrin / Q000032:analysis*; D003600:Cytosol; D034281:Dynamins; D011992:Endosomes / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D020558:GTP Phosphohydrolases / Q000032:analysis*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006624:Hippocampus / Q000166:cytology; D006735:Horseradish Peroxidase; D008562:Membrane Glycoproteins / Q000032:analysis; D008565:Membrane Proteins / Q000032:analysis; D034041:Monomeric Clathrin Assembly Proteins; D009419:Nerve Tissue Proteins / Q000032:analysis; D009474:Neurons / Q000166:cytology; D051381:Rats; D013572:Synaptic Vesicles / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D013574:Synaptosomes / Q000737:chemistry / Q000648:ultrastructure; D050857:Synaptotagmins
D033962:Adaptor Protein Complex 2; D033965:Adaptor Protein Complex alpha Subunits; D033942:Adaptor Proteins, Vesicular Transport; D002135:Calcium-Binding Proteins; D002966:Clathrin; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D034041:Monomeric Clathrin Assembly Proteins; D009419:Nerve Tissue Proteins; C439023:clathrin assembly protein AP180; D050857:Synaptotagmins; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D006735:Horseradish Peroxidase; D020558:GTP Phosphohydrolases; D034281:Dynamins