title: Regulated vesicular trafficking of specific PCDH15 and VLGR1 variants in auditory hair cells.
abstract: Usher syndrome is a genetically heterogeneous disorder characterized by hearing and balance dysfunction and progressive retinitis pigmentosa. Mouse models carrying mutations for the nine Usher-associated genes have splayed stereocilia, and some show delayed maturation of ribbon synapses suggesting these proteins may play different roles in terminal differentiation of auditory hair cells. The presence of the Usher proteins at the basal and apical aspects of the neurosensory epithelia suggests the existence of regulated trafficking through specific transport proteins and routes. Immature mouse cochleae and UB/OC-1 cells were used in this work to address whether specific variants of PCDH15 and VLGR1 are being selectively transported to opposite poles of the hair cells. Confocal colocalization studies between apical and basal vesicular markers and the different PCDH15 and VLGR1 variants along with sucrose density gradients and the use of vesicle trafficking inhibitors show the existence of Usher protein complexes in at least two vesicular subpools. The apically trafficked pool colocalized with the early endosomal vesicle marker, rab5, while the basally trafficked pool associated with membrane microdomains and SNAP25. Moreover, coimmunoprecipitation experiments between SNAP25 and VLGR1 show a physical interaction of these two proteins in organ of Corti and brain. Collectively, these findings establish the existence of a differential vesicular trafficking mechanism for specific Usher protein variants in mouse cochlear hair cells, with the apical variants playing a potential role in endosomal recycling and stereocilia development/maintenance, and the basolateral variants involved in vesicle docking and/or fusion through SNAP25-mediated interactions.
authors.affiliation: Boys Town National Research Hospital, Omaha, Nebraska 68131, USA.
authors.forename: Marisa
authors.initials: M
authors.lastname: Zallocchi
authors.forename: Duane
authors.initials: D
authors.lastname: Delimont
authors.forename: Daniel T
authors.initials: DT
authors.lastname: Meehan
authors.forename: Dominic
authors.initials: D
authors.lastname: Cosgrove
mesh_terms: D020823:ADP-Ribosylation Factor 1 / Q000032:analysis; D000818:Animals; D001923:Brain Chemistry; D000070585:Cadherin Related Proteins; D015820:Cadherins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D002451:Cell Compartmentation; D002454:Cell Differentiation; D016764:Cell Polarity / Q000502:physiology*; D004195:Disease Models, Animal; D055785:Gene Knockdown Techniques; D006198:Hair Cells, Auditory / Q000378:metabolism / Q000648:ultrastructure*; D047468:Immunoprecipitation; D051379:Mice; D008818:Mice, Neurologic Mutants; D009154:Mutation; D009925:Organ of Corti / Q000737:chemistry / Q000648:ultrastructure; D025941:Protein Interaction Mapping; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D034622:RNA Interference; D043562:Receptors, G-Protein-Coupled / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship; D050825:Synaptosomal-Associated Protein 25 / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D022161:Transport Vesicles / Q000737:chemistry / Q000502:physiology*; D052245:Usher Syndromes / Q000378:metabolism; D020696:rab5 GTP-Binding Proteins / Q000032:analysis
chemical_list: D020823:ADP-Ribosylation Factor 1; D000070585:Cadherin Related Proteins; D015820:Cadherins; D011498:Protein Precursors; D043562:Receptors, G-Protein-Coupled; D050825:Synaptosomal-Associated Protein 25; D020696:rab5 GTP-Binding Proteins; C418976:CDHR15 protein, mouse; C485949:Adgrv1 protein, mouse; C494209:Snap25 protein, mouse