title: Mapping the dynamics and nanoscale organization of synaptic adhesion proteins using monomeric streptavidin.
abstract: The advent of super-resolution imaging (SRI) has created a need for optimized labelling strategies. We present a new method relying on fluorophore-conjugated monomeric streptavidin (mSA) to label membrane proteins carrying a short, enzymatically biotinylated tag, compatible with SRI techniques including uPAINT, STED and dSTORM. We demonstrate efficient and specific labelling of target proteins in confined intercellular and organotypic tissues, with reduced steric hindrance and no crosslinking compared with multivalent probes. We use mSA to decipher the dynamics and nanoscale organization of the synaptic adhesion molecules neurexin-1β, neuroligin-1 (Nlg1) and leucine-rich-repeat transmembrane protein 2 (LRRTM2) in a dual-colour configuration with GFP nanobody, and show that these proteins are diffusionally trapped at synapses where they form apposed trans-synaptic adhesive structures. Furthermore, Nlg1 is dynamic, disperse and sensitive to synaptic stimulation, whereas LRRTM2 is organized in compact and stable nanodomains. Thus, mSA is a versatile tool to image membrane proteins at high resolution in complex live environments, providing novel information about the nano-organization of biological structures.
authors.affiliation: Interdisciplinary Institute for Neuroscience, UMR 5297, Centre National de la Recherche Scientifique, 33077 Bordeaux, France.|Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France.
authors.forename: Ingrid
authors.initials: I
authors.lastname: Chamma
authors.affiliation: Interdisciplinary Institute for Neuroscience, UMR 5297, Centre National de la Recherche Scientifique, 33077 Bordeaux, France.|Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France.
authors.forename: Mathieu
authors.initials: M
authors.lastname: Letellier
authors.affiliation: Interdisciplinary Institute for Neuroscience, UMR 5297, Centre National de la Recherche Scientifique, 33077 Bordeaux, France.|Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France.|Imagine Optic, 18 rue Charles de Gaulle, 91400 Orsay, France.
authors.forename: Corey
authors.initials: C
authors.lastname: Butler
authors.affiliation: Interdisciplinary Institute for Neuroscience, UMR 5297, Centre National de la Recherche Scientifique, 33077 Bordeaux, France.|Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France.
authors.forename: Béatrice
authors.initials: B
authors.lastname: Tessier
authors.affiliation: Department of Chemical and Biological Engineering, University at Buffalo, Buffalo, New York 14260, USA.
authors.forename: Kok-Hong
authors.initials: KH
authors.lastname: Lim
authors.affiliation: Interdisciplinary Institute for Neuroscience, UMR 5297, Centre National de la Recherche Scientifique, 33077 Bordeaux, France.|Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France.
authors.forename: Isabel
authors.initials: I
authors.lastname: Gauthereau
authors.affiliation: Interdisciplinary Institute for Neuroscience, UMR 5297, Centre National de la Recherche Scientifique, 33077 Bordeaux, France.|Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France.|Bordeaux Imaging Center, UMS 3420 Centre National de la Recherche Scientifique, University of Bordeaux, US 4 INSERM, 33077 Bordeaux, France.
authors.forename: Daniel
authors.initials: D
authors.lastname: Choquet
authors.affiliation: Interdisciplinary Institute for Neuroscience, UMR 5297, Centre National de la Recherche Scientifique, 33077 Bordeaux, France.|Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France.
authors.forename: Jean-Baptiste
authors.initials: JB
authors.lastname: Sibarita
authors.affiliation: Department of Chemical and Biological Engineering, University at Buffalo, Buffalo, New York 14260, USA.
authors.forename: Sheldon
authors.initials: S
authors.lastname: Park
authors.affiliation: Interdisciplinary Institute for Neuroscience, UMR 5297, Centre National de la Recherche Scientifique, 33077 Bordeaux, France.|Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France.
authors.forename: Matthieu
authors.identifier: http://orcid.org/0000-0001-5465-5641
authors.initials: M
authors.lastname: Sainlos
authors.affiliation: Interdisciplinary Institute for Neuroscience, UMR 5297, Centre National de la Recherche Scientifique, 33077 Bordeaux, France.|Interdisciplinary Institute for Neuroscience, University of Bordeaux, 33077 Bordeaux, France.
authors.forename: Olivier
authors.initials: O
authors.lastname: Thoumine
mesh_terms: D000818:Animals; D019921:Biotinylation; D015816:Cell Adhesion Molecules, Neuronal / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D057809:HEK293 Cells; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D007202:Indicators and Reagents; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D056004:Molecular Dynamics Simulation; D036103:Nanotechnology / Q000379:methods*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D019006:Neural Cell Adhesion Molecules / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D051381:Rats; D013194:Staining and Labeling / Q000379:methods*; D019809:Streptavidin; D013569:Synapses / Q000378:metabolism*
chemical_list: D015816:Cell Adhesion Molecules, Neuronal; D007202:Indicators and Reagents; C561320:LRRTM2 protein, human; C561491:LRRTM2 protein, mouse; C547192:LRRTM2 protein, rat; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D019006:Neural Cell Adhesion Molecules; C093438:neuroligin 1; D049452:Green Fluorescent Proteins; C086959:neurexin Ibeta; D019809:Streptavidin