title: Molecular characterization of the ankle-link complex in cochlear hair cells and its role in the hair bundle functioning.
abstract: Several lines of evidence indicate that very large G-protein-coupled receptor 1 (Vlgr1) makes up the ankle links that connect the stereocilia of hair cells at their base. Here, we show that the transmembrane protein usherin, the putative transmembrane protein vezatin, and the PDZ (postsynaptic density-95/Discs large/zona occludens-1) domain-containing submembrane protein whirlin are colocalized with Vlgr1 at the stereocilia base in developing cochlear hair cells and are absent in Vlgr1-/- mice that lack the ankle links. Direct in vitro interactions between these four proteins further support their involvement in a molecular complex associated with the ankle links and scaffolded by whirlin. In addition, the delocalization of these proteins in myosin VIIa defective mutant mice as well as the myosin VIIa tail direct interactions with vezatin, whirlin, and, we show, Vlgr1 and usherin, suggest that myosin VIIa conveys proteins of the ankle-link complex to the stereocilia. Adenylyl cyclase 6, which was found at the base of stereocilia, was both overexpressed and mislocated in Vlgr1-/- mice. In postnatal day 7 Vlgr1-/- mice, mechanoelectrical transduction currents evoked by displacements of the hair bundle toward the tallest stereocilia (i.e., in the excitatory direction) were reduced in outer but not inner hair cells. In both cell types, stimulation of the hair bundle in the opposite direction paradoxically resulted in significant transduction currents. The absence of ankle-link-mediated cohesive forces within hair bundles lacking Vlgr1 may account for the electrophysiological results. However, because some long cadherin-23 isoforms could no longer be detected in Vlgr1-/- mice shortly after birth, the loss of some apical links could be involved too. The premature disappearance of these cadherin isoforms in the Vlgr1-/- mutant argues in favor of a signaling function of the ankle links in hair bundle differentiation.
authors.affiliation: Institut National de la Santé et de la Recherche Médicale Unité Mixte de Recherche en Santé 587, Collège de France, Institut Pasteur, 75724 Paris cedex 15, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Michalski
authors.forename: Vincent
authors.initials: V
authors.lastname: Michel
authors.forename: Amel
authors.initials: A
authors.lastname: Bahloul
authors.forename: Gaëlle
authors.initials: G
authors.lastname: Lefèvre
authors.forename: Jérémie
authors.initials: J
authors.lastname: Barral
authors.forename: Hideshi
authors.initials: H
authors.lastname: Yagi
authors.forename: Sébastien
authors.initials: S
authors.lastname: Chardenoux
authors.forename: Dominique
authors.initials: D
authors.lastname: Weil
authors.forename: Pascal
authors.initials: P
authors.lastname: Martin
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Hardelin
authors.forename: Makoto
authors.initials: M
authors.lastname: Sato
authors.forename: Christine
authors.initials: C
authors.lastname: Petit
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D002352:Carrier Proteins / Q000378:metabolism; D002614:Chelating Agents / Q000494:pharmacology; D002923:Cilia / Q000502:physiology; D003051:Cochlea / Q000166:cytology*; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004622:Embryo, Mammalian; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D006198:Hair Cells, Auditory / Q000378:metabolism* / Q000648:ultrastructure; D040542:Mechanotransduction, Cellular / Q000235:genetics / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D008855:Microscopy, Electron, Scanning / Q000379:methods; D009924:Organ Culture Techniques; D018408:Patch-Clamp Techniques; D043562:Receptors, G-Protein-Coupled / Q000172:deficiency; D020860:Subtilisin / Q000494:pharmacology
chemical_list: D000262:Adenylyl Cyclases; D002352:Carrier Proteins; D002614:Chelating Agents; D004533:Egtazic Acid; D016326:Extracellular Matrix Proteins; D008565:Membrane Proteins; D043562:Receptors, G-Protein-Coupled; D020860:Subtilisin; C485949:Adgrv1 protein, mouse; C491810:Ush2a protein, mouse; C477218:Whrn protein, mouse; C506603:vezatin protein, mouse; C121100:adenylyl cyclase 6; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid