title: Characterization of Ribeye subunits in zebrafish hair cells reveals that exogenous Ribeye B-domain and CtBP1 localize to the basal ends of synaptic ribbons.
abstract: Synaptic ribbons are presynaptic structures formed by the self-association of RIBEYE-the main structural component of ribbon synapses. RIBEYE consists of two domains: a unique N-terminal A-domain and a C-terminal B-domain that is identical to the transcription co-repressor C-terminal binding protein 2 (CtBP2). Previous studies in cell lines have shown that RIBEYE A-domain alone is sufficient to form ribbon-like aggregates and that both A- and B- domains form homo-and heterotypic interactions. As these interactions are likely the basis for synaptic-ribbon assembly and structural plasticity, we wanted to examine how zebrafish Ribeye A- and B- domains interact with synaptic ribbons in vivo. To that end, we characterized the localization of exogenously expressed Ribeye A- and B- domains and the closely related protein, CtBP1, in the hair cells of transgenic zebrafish larvae. Unexpectedly, exogenously expressed Ribeye A-domain showed variable patterns of localization in hair cells; one zebrafish paralog of A-domain failed to self-associate or localize to synaptic ribbons, while the other self-assembled but sometimes failed to localize to synaptic ribbons. By contrast, Ribeye B-domain/CtBP2 was robustly localized to synaptic ribbons. Moreover, both exogenously expressed B-domain/CtBP2 and CtBP1 were preferentially localized to the basal end of ribbons adjacent to the postsynaptic density. Overexpression of B-domain/CtBP2 also appeared to affect synaptic-ribbon composition; endogenous levels of ribbon-localized Ribeye were significantly reduced as hair cells matured in B-domain/CtBP2 transgenic larvae compared to wild-type. These results reveal how exogenously expressed Ribeye domains interact with synaptic ribbons, and suggest a potential organization of elements within the ribbon body.
authors.affiliation: Department of Otology and Laryngology, Harvard Medical School, Boston, Massachusetts, United States of America; Eaton-Peabody Laboratory, Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, United States of America.
authors.forename: Lavinia
authors.initials: L
authors.lastname: Sheets
authors.affiliation: Oregon Hearing Research Center and Vollum Institute, Oregon Health & Science University, Portland, Oregon, United States of America.
authors.forename: Matthew W
authors.initials: MW
authors.lastname: Hagen
authors.affiliation: Oregon Hearing Research Center and Vollum Institute, Oregon Health & Science University, Portland, Oregon, United States of America.
authors.forename: Teresa
authors.initials: T
authors.lastname: Nicolson
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D005136:Eye Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D006198:Hair Cells, Auditory / Q000378:metabolism* / Q000648:ultrastructure; D007814:Larva / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D013569:Synapses / Q000378:metabolism* / Q000648:ultrastructure; D009435:Synaptic Transmission; D014162:Transfection; D015027:Zebrafish / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029961:Zebrafish Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D005136:Eye Proteins; D021122:Protein Subunits; D012097:Repressor Proteins; D029961:Zebrafish Proteins; C577847:ctbp2a protein, zebrafish