Identification of the Hair Cell Soma-1 Antigen, HCS-1, as Otoferlin
Identification of the Hair Cell Soma-1 Antigen, HCS-1, as Otoferlin
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DOI：
10.1007/s10162-010-0231-6
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发表时间：
2010-12-01
影响因子：
2.4
通讯作者：
Richardson, Guy P.
中科院分区：
文献类型：
--
作者：
Goodyear, Richard J.;Legan, P. Kevin;Richardson, Guy P.
毛细胞是内耳的机械感受器细胞，对我们的听觉和平衡至关重要。内耳中这些受体细胞的数量很少，阻碍了对毛细胞功能重要的蛋白质的识别和表征。单抗的结合特异性为鉴定毛细胞特异性蛋白并将其分离以供进一步研究提供了一种手段。我们已经产生了一种名为毛细胞胞体-1(HCS-1)的单抗，它能特异性地标记至少五类脊椎动物的毛细胞，包括鲨鱼和射线、硬骨鱼、两栖动物、鸟类和哺乳动物。我们用HCS-1对同源抗原进行免疫沉淀，鉴定其为Ferlin蛋白家族成员Otoferlin。DFNB9是一种隐性的、非综合征形式的语前耳聋，其特征是一种听神经病。用免疫细胞化学的方法，我们发现Otoferlin与毛细胞的整个基底膜和分布在大多数毛细胞胞浆中的泡状结构有关。生化分析表明，奥托费林与膜密切相关，因为它不会因钙或盐浓度的变化而增溶。Hcs-1免疫标记不与突触带的组成成分Ribeye共定位，这表明Otoferlin除了在突触小泡释放中发挥预期的功能外，可能在毛细胞中发挥额外的作用。
Hair cells, the mechanosensitive receptor cells of the inner ear, are critical for our senses of hearing and balance. The small number of these receptor cells in the inner ear has impeded the identification and characterization of proteins important for hair cell function. The binding specificity of monoclonal antibodies provides a means for identifying hair cell-specific proteins and isolating them for further study. We have generated a monoclonal antibody, termed hair cell soma-1 (HCS-1), which specifically immunolabels hair cells in at least five vertebrate classes, including sharks and rays, bony fish, amphibians, birds, and mammals. We used HCS-1 to immunoprecipitate the cognate antigen and identified it as otoferlin, a member of the ferlin protein family. Mutations in otoferlin underlie DFNB9, a recessive, nonsyndromic form of prelingual deafness characterized as an auditory neuropathy. Using immunocytochemistry, we find that otoferlin is associated with the entire basolateral membrane of the hair cells and with vesicular structures distributed throughout most of the hair cell cytoplasm. Biochemical assays indicate that otoferlin is tightly associated with membranes, as it is not solubilized by alterations in calcium or salt concentrations. HCS-1 immunolabeling does not co-localize with ribeye, a constituent of synaptic ribbons, suggesting that otoferlin may, in addition to its proposed function in synaptic vesicle release, play additional roles in hair cells.