Zona Pellucida Domain-Containing Protein β-Tectorin is Crucial for Zebrafish Proper Inner Ear Development

BackgroundThe zona pellucida (ZP) domain is part of many extracellular proteins with diverse functions from structural components to receptors. The mammalian β-tectorin is a protein of 336 amino acid residues containing a single ZP domain and a putative signal peptide at the N-terminus of the protein. It is 1 component of a gel-like structure called the tectorial membrane which is involved in transforming sound waves into neuronal signals and is important for normal auditory function. β-Tectorin is specifically expressed in the mammalian and avian inner ear.Methodology/Principal FindingsWe identified and cloned the gene encoding zebrafish β-tectorin. Through whole-mount in situ hybridization, we demonstrated that β-tectorin messenger RNA was expressed in the otic placode and specialized sensory patch of the inner ear during zebrafish embryonic stages. Morpholino knockdown of zebrafish β-tectorin affected the position and number of otoliths in the ears of morphants. Finally, swimming behaviors of β-tectorin morphants were abnormal since the development of the inner ear was compromised.Conclusions/SignificanceOur results reveal that zebrafish β-tectorin is specifically expressed in the zebrafish inner ear, and is important for regulating the development of the zebrafish inner ear. Lack of zebrafish β-tectorin caused severe defects in inner ear formation of otoliths and function.背景：透明带（zona pellucida, ZP）结构域广泛存在于功能多样的各类细胞外蛋白质中，其功能涵盖结构组分与受体等多个范畴。哺乳动物β-盖膜蛋白（β-tectorin）是由336个氨基酸残基组成的蛋白质，其序列包含单个ZP结构域，且在蛋白质N端带有一段推定信号肽。该蛋白是一种名为盖膜（tectorial membrane）的凝胶状结构的组分之一，盖膜可介导声波向神经信号的转换，对维持正常听觉功能至关重要。β-盖膜蛋白特异性表达于哺乳动物与鸟类的内耳组织中。 方法与主要发现：我们成功鉴定并克隆了编码斑马鱼β-盖膜蛋白的基因。通过整体原位杂交（whole-mount in situ hybridization）实验，我们证实斑马鱼胚胎发育阶段，β-盖膜蛋白的信使RNA（messenger RNA, mRNA）可在听板以及内耳的特化感觉斑中表达。利用吗啉代寡核苷酸（Morpholino）对斑马鱼β-盖膜蛋白进行敲低后，吗啉代敲低胚胎内耳中耳石的位置与数量均出现异常。最终，由于内耳发育受损，β-盖膜蛋白敲低胚胎的游泳行为表现异常。 结论与意义：本研究结果表明，斑马鱼β-盖膜蛋白特异性表达于斑马鱼内耳，且对调控斑马鱼内耳发育具有关键作用。斑马鱼β-盖膜蛋白的缺失会导致内耳耳石形成及功能出现严重缺陷。