组蛋白去乙酰化酶(HDACs)调控斑马鱼侧线器发育及其机制研究

Histone deacetylases (HDACs) activity has been shown to be important for modulating many biological processes including cellular development and regeneration in vertebrates. We have previously reported important roles for HDACs in hair cell development and regeneration in zebrafish, and found a significant downregulation of HDAC1 and HDAC3 protein levels in zebrafish treated with the HDACs inhibitors. These findings highlight the importance of HDAC1 and HDAC3 for hair cell differentiation and regeneration in zebrafish lateral line system. Here, using gene interference methods, we try to examine the role of HDAC1 and HDAC3 in the development of zebrafish lateral line system, cell proliferation and survival, as well as the differentiation of hair cells. We performed whole-mount in situ hybridization analysis in combination with drug treatment and transgenic zebrafish to understand the effect of HDAC1 and HDAC3 on signaling pathways. We next explored the mechanisms of gene regulation by HDAC1 and HDAC3 in lateral line primordium using chromatin immunoprecipitation coupled with massive parallel sequencing (ChIP-seq) to identify genomic regions of HDAC1 and HDAC3 enrichment. RNA sequencing (RNA-seq) were performed to indicate the differential expression of genes in lateral line primordium. The project will contribute to a better understanding of the processes of auditory organ development and facilitate prevention and treatment of sensorineural hearing loss.

组蛋白去乙酰化酶（HDACs）在细胞增殖、分化、存活及早期胚胎发育和特定器官形成中发挥重要作用。我们前期研究发现采用HDACs抑制剂调控HDACs能够显著抑制HDAC1、HDAC3活性，并影响斑马鱼侧线神经丘毛细胞的早期分化和再生，提示HDAC1、HDAC3参与毛细胞分化和再生过程。本课题我们拟采用斑马鱼模型，从基因水平调控HDAC1、HDAC3的表达，通过形态结构分析、原位杂交、免疫组化等方法研究HDAC1、HDAC3在侧线系统形成，细胞增殖和存活及毛细胞分化中的作用。采用原位杂交技术结合信号通路调控等方法探讨其作用的机制。最后进一步应用染色质免疫共沉淀测序（ChIP-Seq）结合转录组测序技术（RNA-Seq）深入研究HDAC1、HDAC3调控侧线发育的分子机制。本项目的研究将有利于深入理解听觉器官的发育调控机制，为感音神经性耳聋的防治提供理论依据。

组蛋白去乙酰化酶（HDACs）在细胞增殖、分化、存活及早期胚胎发育和特定器官形成中发挥重要作用。我们前期研究发现采用HDACs抑制剂调控HDACs能够显著抑制HDAC1、HDAC3活性，并影响斑马鱼侧线神经丘毛细胞的早期分化和再生，提示HDAC1、HDAC3参与毛细胞分化和再生过程。本课题我们采用斑马鱼模型，从基因水平调控HDAC1、HDAC3的表达，通过形态结构分析、原位杂交、免疫组化等方法研究HDAC1、HDAC3在侧线系统形成，内耳发育及毛细胞分化中的作用。研究发现抑制HDAC3后导致侧线系统形成障碍，包括神经丘数目减少、位置排列紊乱、原基细胞增殖减少、神经丘中的毛细胞数目减少等。采用整胚原位杂交方法检测HDAC3敲降后侧线发育相关信号途径分子的表达变化，发现Fgf、cxcl12a/cxcr7b等信号途径分子的表达发生改变，提示HDAC3可能通过这些信号途径调控斑马鱼侧线发育。我们采用基因阻抑的方法构建HDAC1表达下调的斑马鱼模型，发现HDAC1低表达后导致斑马鱼胚胎内耳发育障碍，包括听泡变小、耳石数目位置异常、半规管发育异常、内耳毛细胞数目减少等。研究还发现HDAC1敲降后，内耳Fgf信号通路基因的表达明显降低，提示HDAC1可能通过该信号通路调控斑马鱼内耳发育。本项目的研究将有利于深入理解听觉器官的发育调控机制，为感音神经性耳聋的防治提供理论依据。