Molecular and Cellular Biology Program, Oregon State University, Corvallis, OR 97333.
Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR 97333.
Mol Biol Cell. 2021 Jul 1;32(14):1293-1305. doi: 10.1091/mbc.E20-10-0657. Epub 2021 May 12.
Release of neurotransmitter from sensory hair cells is regulated by otoferlin. Despite the importance of otoferlin in the auditory and vestibular pathways, the functional contributions of the domains of the protein have not been fully characterized. Using a zebrafish model, we investigated a mutant otoferlin with a stop codon at the start of the transmembrane domain. We found that both the phenotype severity and the expression level of mutant otoferlin changed with the age of the zebrafish. At the early developmental time point of 72 h post fertilization, low expression of the otoferlin mutant coincided with synaptic ribbon deficiencies, reduced endocytosis, and abnormal transcription of several hair cell genes. As development proceeded, expression of the mutant otoferlin increased, and both synaptic ribbons and hair cell transcript levels resembled wild type. However, hair cell endocytosis deficits and abnormalities in the expression of GABA receptors persisted even after up-regulation of mutant otoferlin. Analysis of membrane-reconstituted otoferlin measurements suggests a function for the transmembrane domain in liposome docking. We conclude that deletion of the transmembrane domain reduces membrane docking, attenuates endocytosis, and results in developmental delay of the hair cell.
感觉毛细胞中神经递质的释放受 otoferlin 调节。尽管 otoferlin 在听觉和前庭途径中很重要，但该蛋白结构域的功能贡献尚未完全确定。我们使用斑马鱼模型研究了一种起始跨膜结构域处带有终止密码子的 otoferlin 突变体。我们发现，突变 otoferlin 的表型严重程度和表达水平都随斑马鱼的年龄而变化。在受精后 72 小时的早期发育时间点，突变 otoferlin 的低表达与突触带缺陷、内吞作用减少以及几个毛细胞基因的转录异常有关。随着发育的进行，突变 otoferlin 的表达增加，突触带和毛细胞转录本水平均类似于野生型。然而，即使突变 otoferlin 的表达上调后，毛细胞内吞作用缺陷和 GABA 受体表达异常仍然存在。对膜重建 otoferlin 测量的分析表明，跨膜结构域在脂质体对接中具有功能。我们得出结论，跨膜结构域的缺失会降低膜对接，减弱内吞作用，并导致毛细胞发育延迟。
