一个新的基因家族Fam49在胚胎发育中的分子细胞机理研究

From our screen for factors regulating neurogenesis, we isolated a novel gene,Fam49a. Fam49a is one member of the small Fam49 gene family,which is characterized by a conserved DUF1394 domain with unknown functions.Up to date, there is less report about the information of this gene family.To better characterize Fam49, RT-PCR and whole mount in situ hybridization were performed to detect their expressions in Xenopus tropicalis embryos.Our preliminary data reveal that Fam49 are mainly abundant expressed in the neural plate and neural crest during early embryogenesis.Further,antisense MO knockdown assays were implemented to investigate the function of Fam49a.Results both from ATG-MO and splicing MO knockdown strongly indicate that Fam49a may play potential roles during central nervous system and neural crest development through inhibiting apoptosis.. Based on the high level conservation on amino acid sequences and expression pattern,we propose that Fam49 members may play similar critical roles in central nervous system and neural crest development.To test our hypothesis, here in this study,we will perform molecular, evolutionary,cellular,developmental biolgoy and genetic studies in Xenopus tropicalis system by virtues of its advantages on classic develomnetal biology and functional genomics researchs.Our goal is to charaterize this novel gene family and elucidate the mechanisms that how Fam49 members exert their funtions to regulate cell fates and even the embryo development.And, at the same time, we will also investigate the expressions and functions of FAM49 genes in different human cell lines to explore the underlying clues for related diseases.

在调控神经形成因子的筛选中，我们发现一个参与神经形成的新因子Fam49a。Fam49a是一个全新的基因家族Fam49的一员，这个基因家族（Fam49a,b,c）具有一个保守的未知功能结构域DUF1394，有关Fam49基因的背景和研究还未有相关文献报道。生物信息学和比较基因组学的分析表明Fam49基因是一类在进化上高度保守的基因。我们的初步研究表明早期胚胎发育时Fam49基因的表达则主要富集于神经板以及神经嵴部位，并且对Fam49a的功能敲落表明其与神经系统发育密切相关。为了阐明Fam49基因家族的本质特性和功能，本项目以模式动物Xenopus tropicalis为研究对象，结合其经典发育生物学和基因功能研究的优势，从分子、进化、细胞、发育等多角度对这个基因家族在早期胚胎发育过程中的表达、功能和相关的分子、细胞、发育机理进行全面深入的研究。

我们前期的研究发现一个在蝾螈的附肢再生过程中特异性表达以及在脑部发育中差异表达的Fam49a基因的EST，Fam49 是一个全新的基因家族，目前还没有Fam49相关的任何基因功能和机制研究的报道。仅有的报道表明Fam49 基因可能人类与几种神经系统疾病的相关。本研究以热带爪蟾为模式动物, 通过分子克隆、表达检测和分子进化分析，发现Fam49是一类在进化上高度保守的蛋白，它们在神经系统和神经嵴中有高丰度的表达；用基因过表达、基因敲落和挽救等基因功能研究表明Fam49 基因是一类重要的调控中枢神经系统和神经嵴发育的因子；Crispr/Cas9基因编辑研究表明Fam49能够调控293T细胞、U251细胞以及早期胚胎细胞的细胞增殖水平；基因编辑研究还表明Fam49b敲除还会抑制U251细胞的分化和迁移；Fam49成员均包含一个未知的功能结构域--DUF1394，分子互作蛋白筛选以及基因编辑后细胞的表达谱分析表明它可能通过与Rac1的互作来影响细胞的增殖、分化和的转移。我们的研究结果暗示Fam49基因不仅对胚胎早期的发育有重要影响，而且可能在肿瘤的发生和转移中具重要作用，深入的后续研究预期可揭示新的肿瘤治疗靶点。