Department of Otolaryngology - Head and Neck Surgery, Chungnam National University College of Medicine, Daejeon, South Korea.
Department of Otorhinolaryngology - Head and Neck Surgery, Seoul National University Bundang Hospital, Seoul National University College of Medicine, Seongnam, South Korea.
Hum Mutat. 2019 May;40(5):525-531. doi: 10.1002/humu.23719. Epub 2019 Feb 28.
Otoancorin (OTOA), encoded by OTOA, is required for the development of the tectorial membrane in the inner ear. Mutations in this gene cause nonsyndromic hearing loss (DFNB22). The molecular mechanisms underlying most DFNB22 remain poorly understood. Disruption of glycosylphosphatidylinositol (GPI) anchorage has been assumed to be the pathophysiology mandating experimental validation. From a Korean deaf family, we identified two trans OTOA variants (c.1320 + 5 G > C and p.Gln589ArgfsX55 [NM_144672.3]) . The pathogenic potential of c.1320 + 5 G > C was confirmed by a minigene splicing assay. To experimentally determine the GPI anchorage, wild-type (WT) and mutant OTOA harboring p.Gln589ArgfsX55 were expressed in HEK293T cells. The mutant OTOA with p.Gln589ArgfsX55 resulted in an uncontrolled release of OTOA into the medium in contrast with phosphatidylinositol-specific phospholipase C-induced controlled release of WT OTOA from the cell surface. Together, the results of this reverse translational study confirmed GPI-anchorage of OTOA and showed that downstream sequences from the 589th amino acid are critical for GPI-anchorage.
耳钙黏蛋白（Otoancorin，OTOA）由 OTOA 编码，是内耳盖膜发育所必需的。该基因的突变会导致非综合征性听力损失（DFNB22）。大多数 DFNB22 的分子机制仍知之甚少。破坏糖基磷脂酰肌醇（GPI）锚定被认为是需要实验验证的病理生理学。我们从一个韩国聋人家族中鉴定出两个跨 OTOA 变体（c.1320 + 5 G > C 和 p.Gln589ArgfsX55 [NM_144672.3]）。通过小基因剪接分析证实了 c.1320 + 5 G > C 的致病性。为了实验确定 GPI 锚定，我们在 HEK293T 细胞中表达了携带 p.Gln589ArgfsX55 的野生型（WT）和突变型 OTOA。与磷脂酰肌醇特异性磷脂酶 C 诱导的 WT OTOA 从细胞表面受控释放相比，携带 p.Gln589ArgfsX55 的突变型 OTOA 导致 OTOA 不受控制地释放到培养基中。综上所述，这项反向转化研究的结果证实了 OTOA 的 GPI 锚定，并表明 589 号氨基酸下游序列对于 GPI 锚定至关重要。
