title: The Munc18-1 domain 3a hinge-loop controls syntaxin-1A nanodomain assembly and engagement with the SNARE complex during secretory vesicle priming.
abstract: Munc18-1 and syntaxin-1A control SNARE-dependent neuroexocytosis and are organized in nanodomains on the plasma membrane of neurons and neurosecretory cells. Deciphering the intra- and intermolecular steps via which they prepare secretory vesicles (SVs) for fusion is key to understanding neuronal and hormonal communication. Here, we demonstrate that expression of a priming-deficient mutant lacking 17 residues of the domain 3a hinge-loop (Munc18-1(Δ317-333)) in PC12 cells engineered to knockdown Munc18-1/2 markedly prolonged SV docking. Single-molecule analysis revealed nonhomogeneous diffusion of Munc18-1 and syntaxin-1A in and out of partially overlapping nanodomains. Whereas Munc18-1(WT) mobility increased in response to stimulation, syntaxin-1A became less mobile. These Munc18-1 and syntaxin-1A diffusional switches were blocked by the expression of Munc18-1(Δ317-333), suggesting that a conformational change in the Munc18-1 hinge-loop controls syntaxin-1A and subsequent SNARE complex assembly. Accordingly, syntaxin-1A confinement was prevented by expression of botulinum neurotoxin type E. The Munc18-1 domain 3a hinge-loop therefore controls syntaxin-1A engagement into SNARE complex formation during priming.
authors.affiliation: Queensland Brain Institute, Clem Jones Centre for Ageing Dementia Research, The University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Ravikiran
authors.initials: R
authors.lastname: Kasula
authors.affiliation: Queensland Brain Institute, Clem Jones Centre for Ageing Dementia Research, The University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Ye Jin
authors.identifier: 0000-0002-3022-7690
authors.initials: YJ
authors.lastname: Chai
authors.affiliation: Queensland Brain Institute, Clem Jones Centre for Ageing Dementia Research, The University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Adekunle T
authors.initials: AT
authors.lastname: Bademosi
authors.affiliation: Queensland Brain Institute, Clem Jones Centre for Ageing Dementia Research, The University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Callista B
authors.initials: CB
authors.lastname: Harper
authors.affiliation: Queensland Brain Institute, Clem Jones Centre for Ageing Dementia Research, The University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Rachel S
authors.identifier: 0000-0003-1604-0253
authors.initials: RS
authors.lastname: Gormal
authors.affiliation: Queensland Brain Institute, Clem Jones Centre for Ageing Dementia Research, The University of Queensland, Brisbane, Queensland 4072, Australia Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Isabel C
authors.initials: IC
authors.lastname: Morrow
authors.affiliation: Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5297, 33077 Bordeaux, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Hosy
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Brett M
authors.initials: BM
authors.lastname: Collins
authors.affiliation: Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5297, 33077 Bordeaux, France Bordeaux Imaging Center, Unité Mixte de Service 3420, Centre National de la Recherche Scientifique, US4 Institut National de la Santé et de la Recherche Médicale, University of Bordeaux, 33077 Bordeaux, France.
authors.forename: Daniel
authors.initials: D
authors.lastname: Choquet
authors.affiliation: Queensland Brain Institute, Clem Jones Centre for Ageing Dementia Research, The University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Andreas
authors.identifier: 0000-0002-3376-1979
authors.initials: A
authors.lastname: Papadopulos
authors.affiliation: Queensland Brain Institute, Clem Jones Centre for Ageing Dementia Research, The University of Queensland, Brisbane, Queensland 4072, Australia f.meunier@uq.edu.au.
authors.forename: Frédéric A
authors.identifier: 0000-0001-6400-1107
authors.initials: FA
authors.lastname: Meunier
mesh_terms: D000818:Animals; D019540:Area Under Curve; D001905:Botulinum Toxins / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D051938:Munc18 Proteins / Q000737:chemistry* / Q000378:metabolism*; D053758:Nanoparticles / Q000737:chemistry*; D016716:PC12 Cells; D000072417:Protein Domains; D017433:Protein Structure, Secondary; D051381:Rats; D050600:SNARE Proteins / Q000378:metabolism*; D022142:Secretory Vesicles / Q000378:metabolism*; D050827:Syntaxin 1 / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D051938:Munc18 Proteins; D050600:SNARE Proteins; D050827:Syntaxin 1; D001905:Botulinum Toxins; C096326:botulinum toxin type E