Atoh1启动子近端缺失小鼠中不同的小脑和内耳表型

文献信息

中文摘要

目的： Atoh1对小脑和内耳的发育至关重要，但其启动子近端区域在体内的作用仍不完全清楚。我们构建了靶向包含C位点的Atoh1启动子近端区域的缺失品系，并检查了其在小脑和内耳中的表型。
方法： 使用CRISPR-Cas9基因组编辑方法，构建了包含C位点的Atoh1启动子近端区域缺失的小鼠，并建立了四个独立的缺失品系。对2月龄小鼠的小脑和内耳进行了大体形态学和组织学检查。
结果： 获得了四个独立的缺失等位基因，按缺失大小递增顺序命名为A系至D系。在小脑中，大体检查和解剖矢状切片显示，从A系到D系，发育不全逐渐加重，伴有小叶减少和皮质结构紊乱。颗粒细胞层逐渐减少，而浦肯野细胞相对保留。行为异常仅在受影响最严重的品系中检测到。在内耳中，所有品系的耳石器官的位觉斑相对保留。壶腹嵴在A-C系中相对保留，但在D系中显示毛细胞丢失。耳蜗表型更为严重：A系保留了相对完整的内毛细胞，但外毛细胞减少，而B-D系在分析区域显示毛细胞几乎完全丢失，且Corti器明显破坏。对可用标本的描述性定量分析支持进行性小脑发育不全、位觉斑相对保留、D系壶腹嵴受累以及B-D系严重的耳蜗毛细胞丢失。
结论： Atoh1启动子近端区域的缺失在体内产生了不同的组织特异性表型。这些发现表明，Atoh1启动子近端区域在小脑和内耳中的需求存在差异，并且其贡献在前庭和耳蜗感觉器官之间也有所不同。

英文摘要

OBJECTIVE: Atoh1 is essential for the development of the cerebellum and inner ear, but the in vivo role of its promoter-proximal region remains incompletely understood. We generated deletion lines targeting the Atoh1 promoter-proximal region containing C sites and examined their phenotypes in the cerebellum and inner ear.
METHODS: Using the CRISPR-Cas9 genome editing method, mice with deletions in the Atoh1 promoter-proximal region containing C sites were generated, and four independent deletion lines were established. Gross morphology and histology of the cerebellum and inner ear were examined in 2-month-old mice.
RESULTS: Four independent deletion alleles were obtained and designated line A to line D in ascending order of deletion size. In the cerebellum, gross examination and sagittal sections showed progressively more severe hypoplasia from line A to line D, accompanied by reduced foliation and disorganization of cortical architecture. The granule cell layer was progressively reduced, whereas Purkinje cells were relatively preserved. Behavioral abnormalities were detected only in the most severely affected line. In the inner ear, the maculae of the otolith organs were relatively preserved across all lines. The ampullary cristae were relatively preserved in lines A-C but showed hair-cell loss in line D. Cochlear phenotypes were more severe: line A retained relatively preserved inner hair cells with reduced outer hair cells, whereas lines B-D showed near-complete loss of hair cells and marked disruption of the organ of Corti in the analyzed regions. Descriptive quantitative analyses of available specimens supported progressive cerebellar hypoplasia, relative macular preservation, line D crista involvement, and severe cochlear hair-cell loss in lines B-D.
CONCLUSION: Deletion of the Atoh1 promoter-proximal region produced distinct tissue-specific phenotypes in vivo. These findings suggest that the Atoh1 promoter-proximal region is differentially required in the cerebellum and inner ear and that its contribution varies among vestibular and cochlear sensory organs.