title: Conserved prefusion protein assembly in regulated exocytosis.
abstract: The regulated release of hormones and neurotransmitters is a fundamental process throughout the animal kingdom. The short time scale for the calcium triggering of vesicle fusion in regulated secretion suggests that the calcium sensor synaptotagmin and the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) membrane fusion machinery are well ordered before the calcium signal. To gain insight into the organization of the prefusion protein assembly in regulated exocytosis, we undertook a structural/functional study of the vesicular synaptotagmin1 and the plasma membrane SNARE proteins, which copurify from the brain in the absence of calcium. Based on an evolutionary analysis, mutagenesis screens, and a computational protein docking approach, we now provide the first testable description of the supramolecular prefusion assembly. Perturbing the determined synaptotagmin/SNARE-interacting interface in several models of regulated exocytosis altered the secretion of hormones and neurotransmitters. These mutations also disrupted the constitutive synaptotagmin/SNARE link in full agreement with our model. We conclude that the interaction of synaptotagmin with preassembled plasma membrane SNARE proteins, before the action of calcium, can provide a precisely organized "tethering" scaffold that underlies regulated secretion throughout evolution.
authors.affiliation: MRC Laboratory of Molecular Biology, Cambridge CB2 2QH, United Kingdom.
authors.forename: Colin
authors.initials: C
authors.lastname: Rickman
authors.forename: José L
authors.initials: JL
authors.lastname: Jiménez
authors.forename: Margaret E
authors.initials: ME
authors.lastname: Graham
authors.forename: Deborah A
authors.initials: DA
authors.lastname: Archer
authors.forename: Mikhail
authors.initials: M
authors.lastname: Soloviev
authors.forename: Robert D
authors.initials: RD
authors.lastname: Burgoyne
authors.forename: Bazbek
authors.initials: B
authors.lastname: Davletov
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002942:Circular Dichroism; D019295:Computational Biology; D017124:Conserved Sequence; D005089:Exocytosis / Q000502:physiology*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D050600:SNARE Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D050857:Synaptotagmins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D050600:SNARE Proteins; D050857:Synaptotagmins