Complexin arrests a pool of docked vesicles for fast Ca2+-dependent release.
Regulated exocytosis requires that the assembly of the basic membrane fusion machinery is temporarily arrested. Synchronized membrane fusion is then caused by a specific trigger--a local rise of the Ca(2+) concentration. Using reconstituted giant unilamellar vesicles (GUVs), we have analysed the role of complexin and membrane-anchored synaptotagmin 1 in arresting and synchronizing fusion by lipid-mixing and cryo-electron microscopy. We find that they mediate the formation and consumption of docked small unilamellar vesicles (SUVs) via the following sequence of events: Synaptotagmin 1 mediates v-SNARE-SUV docking to t-SNARE-GUVs in a Ca(2+)-independent manner. Complexin blocks vesicle consumption, causing accumulation of docked vesicles. Together with synaptotagmin 1, complexin synchronizes and stimulates rapid fusion of accumulated docked vesicles in response to physiological Ca(2+) concentrations. Thus, the reconstituted assay resolves both the stimulatory and inhibitory function of complexin and mimics key aspects of synaptic vesicle fusion.
Heidelberg University Biochemistry Center, Heidelberg, Germany.
Jörg
J
Malsam
Daniel
D
Parisotto
Tanmay A M
TA
Bharat
Andrea
A
Scheutzow
Jean Michel
JM
Krause
John A G
JA
Briggs
Thomas H
TH
Söllner
D033942:Adaptor Proteins, Vesicular Transport / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002118:Calcium / Q000494:pharmacology*; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D008561:Membrane Fusion / Q000187:drug effects; D008954:Models, Biological; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D011485:Protein Binding / Q000187:drug effects; D009435:Synaptic Transmission / Q000187:drug effects; D013572:Synaptic Vesicles / Q000187:drug effects* / Q000378:metabolism*; D050863:Synaptotagmin I / Q000737:chemistry / Q000378:metabolism / Q000502:physiology; D013997:Time Factors; D053835:Unilamellar Liposomes / Q000378:metabolism
D033942:Adaptor Proteins, Vesicular Transport; D009419:Nerve Tissue Proteins; D050863:Synaptotagmin I; D053835:Unilamellar Liposomes; C095992:complexin I; D002118:Calcium