Molecular characterization of the ankle-link complex in cochlear hair cells and its role in the hair bundle functioning
Molecular characterization of the ankle-link complex in cochlear hair cells and its role in the hair bundle functioning
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DOI：
10.1523/jneurosci.0342-07.2007
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发表时间：
2007-06-13
影响因子：
5.3
通讯作者：
Petit, Christine
中科院分区：
文献类型：
--
作者：
Michalski, Nicolas;Michel, Vincent;Petit, Christine
一些证据表明，非常大的g蛋白偶联受体1 （Vlgr1）构成了连接毛细胞基部立体纤毛的踝链。本研究表明，在发育中的耳蜗毛细胞中，跨膜蛋白引导素、推测的跨膜蛋白vezatin和含有PDZ（突触后密度-95/Discslarge/带状occluden -1）结构域的亚膜蛋白whirlin与Vlgr1在立体纤毛基础上共定位，而在缺乏踝环的Vlgr1（-/-）小鼠中则不存在。这四种蛋白之间的直接体外相互作用进一步支持它们参与与踝关节相关的分子复合物，并由旋转蛋白支撑。此外，这些蛋白在myosin VIIa缺陷突变小鼠中的脱位，以及myosin VIIa尾巴与vezatin， whirlin，以及我们发现的Vlgr1和usher的直接相互作用，表明myosin VIIa将脚环复合物的蛋白传递到立体纤毛。腺苷酸环化酶6位于静纤毛基部，在Vlgr1（-/-）小鼠中存在过表达和错位。在出生后第7天的Vlgr1（-/-）小鼠中，毛束向最高的立体纤毛位移（即在兴奋方向）引起的机电转导电流在外部毛细胞中减少，而在内部毛细胞中没有。在这两种细胞类型中，相反方向的毛束刺激反而导致了显著的转导电流。在缺乏Vlgr1的毛束中，踝关节连接介导的凝聚力的缺失可能解释了电生理结果。然而，由于Vlgr1（-/-）小鼠出生后不久就不能再检测到一些长钙粘蛋白-23同工型，因此也可能涉及一些根尖连接的丢失。这些钙粘蛋白异构体在Vlgr1（-/-）突变体中过早消失，支持踝链在毛束分化中的信号功能。
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/Discslarge/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.