Genetic and Genomic Approaches to Study Hair Cell Regeneration in Zebrafish

内耳中机械感受器(毛细胞)的损坏或丢失是人类 耳聋，因为两者在出生后都是不可逆转的。与哺乳动物不同，鱼可以代替受损的毛细胞 贯穿整个成年期。这种再生能力在内耳中观察到，但也在进化和 与生理有关的器官，即侧线.这种肤浅的感觉器官为研究提供了一个有价值的工具 在整个动物的时间和背景下，再生的动态事件。极少的基因 到目前为止，参与再生的基因已被确定，阻碍了修复毛细胞的进展 人类。为了解决这一点，本应用程序的长期目标是阐明 侧线毛细胞再生的分子事件。作为入口点，我们使用 斑马鱼突变系(凤凰)，我们已经确定了它的特征。突变的基因是一种新的突变幼虫 侧线再生毛细胞明显减少。菲尼克斯基因在 支持细胞，以前被认为是祖细胞。他们没有进入牢房 在没有凤凰的情况下骑车。第一个具体目标是破译凤凰的功能。为了做到这一点，我们 将鱼类细胞系的细胞学方法与以前的突变幼虫的表达筛选相结合 已知或新发现的分子参与者在再生过程中。第二个具体目标是找到 凤凰卫视的合伙人。我们在野生型和凤凰突变体中使用基因组和蛋白质组学方法。 背景。我们正在鉴定侧线每种细胞类型的转录组，并评估它们的 在再生过程中发生变化。蛋白质组学方法是基于鱼类细胞系中的免疫共沉淀。 使用一种针对凤凰的抗体。在第三个具体目标中，我们将寻求初步数据显示， Hutchinson-Gilford Progeria综合征患者的人成纤维细胞中菲尼克斯定位发生改变 正在过早地衰老。利用免疫沉淀和质谱学，我们将分离和 对凤凰城的人类同源物进行排序。这项工作将为人类再生的遗传学提供洞察力。 毛细胞，开辟了新的治疗途径。

Damage or loss of the mechanoreceptors (hair cells) in the inner ear, are the leading cause for human deafness, because both are irreversible after birth. Unlike mammals, fish replace damaged hair cells throughout adulthood. This regenerative power is observed in the inner ear, but also in an evolutionary and physiologically related organ the lateral line. This superficial sensory organ offers a valuable tool to study in vivo the dynamic events of regeneration in time and in the context of a whole animal. Very few genes involved in regeneration have been identified so far, hampering progress towards restoring hair cells in humans. To address this particular point, the long-term objectives of this application are to elucidate the molecular events underlying the regeneration of hair cells in the lateral line. As an entry point, we are using a zebrafish mutant line (phoenix), which we have characterized. The mutated gene is novel and mutant larvae have a clear reduction in regenerated hair cells in the lateral line. The phoenix gene is expressed in supporting cells, which have been previously implicated as progenitor cells. They are not entering the cell cycle in the absence of phoenix. The first specific aim is to decipher the function of phoenix. To do so, we combine a cytological approach in fish cell lines with an expression screen in mutant larvae of previously known or newly discovered molecular players in the regeneration process. The second specific aim is to find partners of phoenix. We use genomic and a proteomic approaches in the wild type and phoenix mutant context. We are characterizing the transcriptome of each cell type of the lateral line and assess their changes during regeneration. The proteomic approach is based on co-immunoprecipitation in fish cells lines using an antibody against phoenix. In the third specific aim, we will pursue preliminary data showing that, phoenix localization is altered in human fibroblasts of Hutchinson-Gilford Progeria Syndrome patients, who are aging prematurely. Using immunoprecipitation coupled with mass spectroscopy, we will isolate and sequence the human homolog of phoenix. This work will provide insight into the genetics of regeneration of hair cells, opening up new therapeutic avenues.