title: Proteomic screening of glutamatergic mouse brain synaptosomes isolated by fluorescence activated sorting.
abstract: For decades, neuroscientists have used enriched preparations of synaptic particles called synaptosomes to study synapse function. However, the interpretation of corresponding data is problematic as synaptosome preparations contain multiple types of synapses and non-synaptic neuronal and glial contaminants. We established a novel Fluorescence Activated Synaptosome Sorting (FASS) method that substantially improves conventional synaptosome enrichment protocols and enables high-resolution biochemical analyses of specific synapse subpopulations. Employing knock-in mice with fluorescent glutamatergic synapses, we show that FASS isolates intact ultrapure synaptosomes composed of a resealed presynaptic terminal and a postsynaptic density as assessed by light and electron microscopy. FASS synaptosomes contain bona fide glutamatergic synapse proteins but are almost devoid of other synapse types and extrasynaptic or glial contaminants. We identified 163 enriched proteins in FASS samples, of which FXYD6 and Tpd52 were validated as new synaptic proteins. FASS purification thus enables high-resolution biochemical analyses of specific synapse subpopulations in health and disease.
authors.affiliation: Department of Molecular Neurobiology, Max Planck Institute of Experimental Medicine, Göttingen, Germany.
authors.forename: Christoph
authors.initials: C
authors.lastname: Biesemann
authors.forename: Mads
authors.initials: M
authors.lastname: Grønborg
authors.forename: Elisa
authors.initials: E
authors.lastname: Luquet
authors.forename: Sven P
authors.initials: SP
authors.lastname: Wichert
authors.forename: Véronique
authors.initials: V
authors.lastname: Bernard
authors.forename: Simon R
authors.initials: SR
authors.lastname: Bungers
authors.forename: Ben
authors.initials: B
authors.lastname: Cooper
authors.forename: Frédérique
authors.initials: F
authors.lastname: Varoqueaux
authors.forename: Liyi
authors.initials: L
authors.lastname: Li
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Byrne
authors.forename: Henning
authors.initials: H
authors.lastname: Urlaub
authors.forename: Olaf
authors.initials: O
authors.lastname: Jahn
authors.forename: Nils
authors.initials: N
authors.lastname: Brose
authors.forename: Etienne
authors.initials: E
authors.lastname: Herzog
mesh_terms: D000818:Animals; D001921:Brain / Q000166:cytology* / Q000378:metabolism; D002469:Cell Separation / Q000379:methods; D005434:Flow Cytometry / Q000379:methods*; D018698:Glutamic Acid / Q000378:metabolism*; D007473:Ion Channels / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D009474:Neurons / Q000166:cytology* / Q000378:metabolism; D040901:Proteomics; D013569:Synapses / Q000378:metabolism; D013574:Synaptosomes / Q000502:physiology*; D050598:Vesicular Glutamate Transport Protein 1 / Q000235:genetics / Q000378:metabolism
chemical_list: C518765:FXYD6 protein, mouse; D007473:Ion Channels; C493465:Slc17a7 protein, mouse; D050598:Vesicular Glutamate Transport Protein 1; D018698:Glutamic Acid