组蛋白去甲基化酶PHF8在斑马鱼内耳发育中的作用及其机制

Sensorineural deafness is often caused by abnormalities in the development of the inner ear, but the inner ear teratogenic deafness mechanism is unclear. Our previous studies have found that histone modification is closely related to the zebrafish inner ear development, especially in the preliminary experiments of this project found that PHD zinc finger protein 8 (PHD finger protein 8, PHF8) expression in the feeling and semicircular canal of inner ear in zebrafish development. Furthermore, knockdown of PHF8 by the technology of morphine Lin modified antisense oligonucleotide, we found that zebrafish inner ear development obvious flaws, hair cells significantly reduced, and accompanied by semicircular canal dysplasia. These preliminary results indicate that PHF8 played an important role in the inner ear development. Therefore, this project would establish the PHF8 mutants and overexpression of zebrafish animal models to further confirm the role of PHF8 in inner ear phenotype, hair cells proliferation and differentiation, as well as the regeneration after damage. Then use a variety of biological methods, such as chromatin immunization coprecipitation sequencing and transcriptome sequencing, to study the molecular mechanism of PHF8 regulating the inner ear development of zebrafish. Our project would finally clarify the role of PHF8 in zebrafish inner ear development and its molecular mechanism, providing data of scientific experiment for clinical treatment of sensorineural deafness.

感音神经性耳聋常为内耳发育异常所导致，但内耳致畸致聋机理尚不清楚。我们前期研究发现组蛋白修饰与斑马鱼内耳发育密切相关，特别是在本项目预实验中发现，PHD锌指蛋白8（PHD finger protein8, PHF8）表达于斑马鱼发育中内耳的感觉斑和半规管，而且运用吗啡啉修饰反义寡核苷酸技术下调PHF8表达后，斑马鱼内耳发育明显缺陷，毛细胞数量显著减少，并伴有半规管发育异常。这些预实验结果提示PHF8是一个重要的内耳发育调节因子。因此，本项目拟运用PHF8突变体和过表达斑马鱼动物模型进一步确认PHF8在斑马鱼胚胎的内耳表型、毛细胞增殖与分化以及损伤再生中的作用；然后运用染色质免疫共沉淀测序及转录组测序等多种生物学手段深入研究PHF8调控斑马鱼内耳发育的分子机制，最终阐明PHF8在斑马鱼内耳发育中的作用及其分子机制，为临床治疗感音神经性耳聋提供科学实验依据。

感音神经性耳聋是一种常见的感觉障碍性疾病，受到遗传和环境等多重因素影响，致病机理复杂。临床上，很多感音神经性耳聋常为内耳发育异常所导致，然而，内耳致畸致聋病程复杂，致病机制尚不清楚，导致临床治疗方法非常有限。组蛋白去甲基酶PHF8在多种发育过程中起着关键作用，但是，对其在听觉器官发育中的作用的认识仍然极其有限。我们采用原位杂交技术检测了斑马鱼侧线系统和耳泡中PHF8的表达。并发现PHF8的敲低显著破坏了后侧线系统的发育，影响了细胞的迁移，减少了侧线神经丘的数量。在内耳发育过程中，PHF8的敲低也导致了半规管和耳石在大小、数量和位置上的严重畸形。PHF8的敲低也导致了侧线神经丘和内耳感觉毛细胞的分化缺陷，与细胞迁移和沉积有关的多种信号通路相关的的许多基因的表达也发生了改变，包括Wnt和FGF通路。总之，目前的研究结果证实PHF8是一种影响听觉器官发育的影响因子，使其成为治疗听力损失的潜在候选基因。