title: A new probe for super-resolution imaging of membranes elucidates trafficking pathways.
abstract: The molecular composition of the organelles involved in membrane recycling is difficult to establish as a result of the absence of suitable labeling tools. We introduce in this paper a novel probe, named membrane-binding fluorophore-cysteine-lysine-palmitoyl group (mCLING), which labels the plasma membrane and is taken up during endocytosis. It remains attached to membranes after fixation and permeabilization and can therefore be used in combination with immunostaining and super-resolution microscopy. We applied mCLING to mammalian-cultured cells, yeast, bacteria, primary cultured neurons, Drosophila melanogaster larval neuromuscular junctions, and mammalian tissue. mCLING enabled us to study the molecular composition of different trafficking organelles. We used it to address several questions related to synaptic vesicle recycling in the auditory inner hair cells from the organ of Corti and to investigate molecular differences between synaptic vesicles that recycle actively or spontaneously in cultured neurons. We conclude that mCLING enables the investigation of trafficking membranes in a broad range of preparations.
authors.affiliation: Department of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, GermanyDepartment of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, Germany International Max Planck Research School for Neurosciences, 37077 Göttingen, Germany Collaborative Research Center 889 and Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, University of Göttingen, 37099 Göttingen, Germany.
authors.forename: Natalia H
authors.initials: NH
authors.lastname: Revelo
authors.affiliation: Department of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, GermanyDepartment of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, Germany srizzol@gwdg.de dirk.kamin@mpibpc.mpg.de.
authors.forename: Dirk
authors.initials: D
authors.lastname: Kamin
authors.affiliation: Department of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, GermanyDepartment of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, Germany International Max Planck Research School for Molecular Biology, 37077 Göttingen, Germany.
authors.forename: Sven
authors.initials: S
authors.lastname: Truckenbrodt
authors.affiliation: Department of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, Germany International Max Planck Research School for Neurosciences, 37077 Göttingen, Germany Collaborative Research Center 889 and Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, University of Göttingen, 37099 Göttingen, Germany.
authors.forename: Aaron B
authors.initials: AB
authors.lastname: Wong
authors.affiliation: Department of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, Germany Collaborative Research Center 889 and Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, University of Göttingen, 37099 Göttingen, Germany.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Reuter-Jessen
authors.affiliation: Department of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, Germany Collaborative Research Center 889 and Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, University of Göttingen, 37099 Göttingen, Germany.
authors.forename: Ellen
authors.initials: E
authors.lastname: Reisinger
authors.affiliation: Department of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, Germany Collaborative Research Center 889 and Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, University of Göttingen, 37099 Göttingen, Germany Collaborative Research Center 889 and Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, University of Göttingen, 37099 Göttingen, Germany.
authors.forename: Tobias
authors.initials: T
authors.lastname: Moser
authors.affiliation: Department of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, GermanyDepartment of Neuro- and Sensory Physiology; European Neuroscience Institute; and InnerEarLab and Molecular Biology of Cochlear Neurotransmission Group, Department of Otolaryngology; University Medical Center Göttingen, 37099 Göttingen, Germany Collaborative Research Center 889 and Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, University of Göttingen, 37099 Göttingen, Germany Collaborative Research Center 889 and Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, University of Göttingen, 37099 Göttingen, Germany srizzol@gwdg.de dirk.kamin@mpibpc.mpg.de.
authors.forename: Silvio O
authors.initials: SO
authors.lastname: Rizzoli
mesh_terms: D000818:Animals; D001419:Bacteria; D019556:COS Cells; D002462:Cell Membrane / Q000502:physiology*; D002522:Chlorocebus aethiops; D004331:Drosophila melanogaster; D004705:Endocytosis / Q000502:physiology; D005089:Exocytosis / Q000502:physiology; D005456:Fluorescent Dyes; D006199:Hair Cells, Auditory, Inner / Q000502:physiology; D006624:Hippocampus / Q000166:cytology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence / Q000379:methods*; D009474:Neurons / Q000502:physiology; D009925:Organ of Corti / Q000502:physiology; D015388:Organelles / Q000502:physiology*; D061251:Primary Cell Culture; D021381:Protein Transport / Q000502:physiology*; D051381:Rats; D012441:Saccharomyces cerevisiae; D013572:Synaptic Vesicles / Q000502:physiology
chemical_list: D005456:Fluorescent Dyes