The integrity of cochlear hair cells is established and maintained through the localization of Dia1 at apical junctional complexes and stereocilia
The integrity of cochlear hair cells is established and maintained through the localization of Dia1 at apical junctional complexes and stereocilia
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DOI：
10.1038/s41419-020-02743-z
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发表时间：
2020-07-16
影响因子：
9
通讯作者：
Ueyama, Takehiko
中科院分区：
文献类型：
--
作者：
Ninoyu, Yuzuru;Sakaguchi, Hirofumi;Ueyama, Takehiko
Dia 1属于透明相关的肌动蛋白家族，通过肌动蛋白的直线延伸活性影响多种细胞过程。最近，新的DIA 1突变体，如p.R1213X（p.R1204X）和p.A265S，已被报道导致常染色体显性感音神经性听力损失（DFNA 1）。此外，活性DIA 1突变体以功能获得的方式诱导进行性听力损失。然而，DIA 1（R1213 X/R1204 X）的亚细胞定位和病理功能仍不清楚。在本研究中，我们证明了本地化的内源性Dia 1和组成型活性Dia 1突变体在耳蜗中，使用转基因小鼠表达FLAG标记的Dia 1（R1204 X）（Dia 1-TG）。内源性Dia 1和DIA 1突变体从生命早期开始在Corti器官和螺旋神经节区域表达;随着耳蜗成熟，它们定位于毛细胞（HC）和支持细胞（SC）之间的顶端连接复合体（AJCs）。为了研究DIA 1-TG小鼠的HC易感性，我们将4周龄小鼠暴露于中等噪声，这诱导了噪声暴露后4周的耳蜗突触病和静纤毛超微结构变化的暂时性阈值偏移。此外，我们建立了表达AcGFP标记的DIA 1（R1213 X）（DIA 1-KI）的敲入（KI）小鼠系，并证实了突变体定位于AJC和HC中的静纤毛尖端。在稳定表达AcGFP-DIA 1（R1213 X）的MDCK（AcGFP-DIA 1（R1213 X））细胞中，AcGFP-DIA 1（R1213 X）显示出在细胞顶端表面的微绒毛上的显著定位和在细胞-细胞连接处的定位减少。DIA 1-TG小鼠在5月龄时表现出AJC的模糊和皱褶的周向肌动蛋白带和异常静纤毛伴随HC丢失。总之，Dia 1在AJCs和静纤毛的发育和维持中起着关键作用，确保耳蜗和HC的完整性。HC的亚临床/潜在脆弱性可能是DFNA 1患者中进行性听力损失的原因，因此提出了预防与DFNA 1相关的HC变性和进行性听力损失的新治疗靶点。
Dia1, which belongs to the diaphanous-related formin family, influences a variety of cellular processes through straight actin elongation activity. Recently, novel DIA1 mutants such as p.R1213X (p.R1204X) and p.A265S, have been reported to cause an autosomal dominant sensorineural hearing loss (DFNA1). Additionally, active DIA1 mutants induce progressive hearing loss in a gain-of-function manner. However, the subcellular localization and pathological function of DIA1(R1213X/R1204X) remains unknown. In the present study, we demonstrated the localization of endogenous Dia1 and the constitutively active DIA1 mutant in the cochlea, using transgenic mice expressing FLAG-tagged DIA1(R1204X) (DIA1-TG). Endogenous Dia1 and the DIA1 mutant were regionally expressed at the organ of Corti and the spiral ganglion from early life; alongside cochlear maturation, they became localized at the apical junctional complexes (AJCs) between hair cells (HCs) and supporting cells (SCs). To investigate HC vulnerability in theDIA1-TG mice, we exposed 4-week-old mice to moderate noise, which induced temporary threshold shifts with cochlear synaptopathy and ultrastructural changes in stereocilia 4 weeks post noise exposure. Furthermore, we established a knock-in (KI) mouse line expressing AcGFP-tagged DIA1(R1213X) (DIA1-KI) and confirmed mutant localization at AJCs and the tips of stereocilia in HCs. In MDCK(AcGFP-DIA1(R1213X))cells with stable expression of AcGFP-DIA1(R1213X), AcGFP-DIA1(R1213X) revealed marked localization at microvilli on the apical surface of cells and decreased localization at cell-cell junctions. TheDIA1-TG mice demonstrated hazy and ruffled circumferential actin belts at AJCs and abnormal stereocilia accompanied with HC loss at 5 months of age. In conclusion, Dia1 plays a pivotal role in the development and maintenance of AJCs and stereocilia, ensuring cochlear and HC integrity. Subclinical/latent vulnerability of HCs may be the cause of progressive hearing loss in DFNA1 patients, thus suggesting new therapeutic targets for preventing HC degeneration and progressive hearing loss associated with DFNA1.