title: Synaptic proteins promote calcium-triggered fast transition from point contact to full fusion.
abstract: The molecular underpinnings of synaptic vesicle fusion for fast neurotransmitter release are still unclear. Here, we used a single vesicle-vesicle system with reconstituted SNARE and synaptotagmin-1 proteoliposomes to decipher the temporal sequence of membrane states upon Ca(2+)-injection at 250-500 μM on a 100-ms timescale. Furthermore, detailed membrane morphologies were imaged with cryo-electron microscopy before and after Ca(2+)-injection. We discovered a heterogeneous network of immediate and delayed fusion pathways. Remarkably, all instances of Ca(2+)-triggered immediate fusion started from a membrane-membrane point-contact and proceeded to complete fusion without discernible hemifusion intermediates. In contrast, pathways that involved a stable hemifusion diaphragm only resulted in fusion after many seconds, if at all. When complexin was included, the Ca(2+)-triggered fusion network shifted towards the immediate pathway, effectively synchronizing fusion, especially at lower Ca(2+)-concentration. Synaptic proteins may have evolved to select this immediate pathway out of a heterogeneous network of possible membrane fusion pathways.DOI:http://dx.doi.org/10.7554/eLife.00109.001.
authors.affiliation: Departments of Molecular and Cellular Physiology, Neurology and Neurological Sciences, Structural Biology, Photon Science and Howard Hughes Medical Institute ,  Stanford University ,  Stanford ,  USA.
authors.forename: Jiajie
authors.initials: J
authors.lastname: Diao
authors.forename: Patricia
authors.initials: P
authors.lastname: Grob
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Cipriano
authors.forename: Minjoung
authors.initials: M
authors.lastname: Kyoung
authors.forename: Yunxiang
authors.initials: Y
authors.lastname: Zhang
authors.forename: Sachi
authors.initials: S
authors.lastname: Shah
authors.forename: Amie
authors.initials: A
authors.lastname: Nguyen
authors.forename: Mark
authors.initials: M
authors.lastname: Padolina
authors.forename: Ankita
authors.initials: A
authors.lastname: Srivastava
authors.forename: Marija
authors.initials: M
authors.lastname: Vrljic
authors.forename: Ankita
authors.initials: A
authors.lastname: Shah
authors.forename: Eva
authors.initials: E
authors.lastname: Nogales
authors.forename: Steven
authors.initials: S
authors.lastname: Chu
authors.forename: Axel T
authors.initials: AT
authors.lastname: Brunger
mesh_terms: D000200:Action Potentials; D033942:Adaptor Proteins, Vesicular Transport / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D015870:Gene Expression; D008561:Membrane Fusion; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D011510:Proteolipids / Q000378:metabolism* / Q000648:ultrastructure; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D009435:Synaptic Transmission; D013572:Synaptic Vesicles / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D050825:Synaptosomal-Associated Protein 25 / Q000235:genetics / Q000378:metabolism*; D050863:Synaptotagmin I / Q000235:genetics / Q000378:metabolism*; D050827:Syntaxin 1 / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D050988:Vesicle-Associated Membrane Protein 2 / Q000235:genetics / Q000378:metabolism
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; D009419:Nerve Tissue Proteins; D011510:Proteolipids; D011994:Recombinant Proteins; C494210:Snap25 protein, rat; C494213:Stx1a protein, rat; D050825:Synaptosomal-Associated Protein 25; D050863:Synaptotagmin I; D050827:Syntaxin 1; C494312:Syt1 protein, rat; C494706:Vamp2 protein, rat; D050988:Vesicle-Associated Membrane Protein 2; C095992:complexin I; C020485:proteoliposomes; D002118:Calcium