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Roles of a Wnt activator, Ccd1, in neural development

Wnt 激活剂 Ccd1 在神经发育中的作用

基本信息

批准号：
17300098
负责人：
MASU Masayuki
金额：
$ 10.16万
依托单位：
University of Tsukuba
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2007
项目状态：
已结题

项目摘要

Wnt signaling plays important roles in axis formation, differentiation, growth, axon guidance, and synaptogenesis. Wnt signal is transduced via three pathways, beta-catenin pathway, PCP pathway, and Calcium pathway. Although many molecules involved in the Wnt pathways have been identified, many questions regarding the regulation of Wnt signaling remain unresolved. Recently we identified a novel protein Ccd1 (Coiled-coil-DIX1) as third DIX-domain-containing protein from zebrafish. We showed that Ccd1 activates Wnt/beta-catenin pathway both in vitro and in vivo, and that it regulates neural patterning in zebrafish embryos. We also found that many Ccd1 isoforms are generated via differential promoter usage and alternative splicing in mouse. Among them, a novel isoform, named Ccd1A, contains one calponin homology (CH) domain in the N-terminus in addition to the coiled-coil and DIX domains found in the original isoform (named Ccd1B). From zebrafish, we isolated second gene, Ccd2, which encode both A and B subtypes. When heterogouly expressed in cultured cell lines, Ccd1B protein is present as small vesicles throughout cells, Ccd2A protein is co-localized with actin fibers. In addition, the cytoplasmic localization of Dishevelled (Dvl) protein is altered by forced expression of Ccd1A and Ccd1B, suggesting that Ccd1 protein interacts with Dvl in the cell. Although we generated knockout mice, they did not show any obvious phenotypes. When we overexpressed Ccd2 in zebrafish embryos, they showed curly tail-down phenotype, suggesting that Ccd1 plays a role in midline signaling.

Wnt信号在轴的形成、分化、生长、轴突引导和突触发生中起重要作用。Wnt信号通过三种途径转导，β -连环蛋白途径、PCP途径和钙途径。虽然许多参与Wnt信号通路的分子已经被确定，但关于Wnt信号调控的许多问题仍未解决。最近，我们从斑马鱼中发现了一个新的蛋白Ccd1 （Coiled-coil-DIX1）作为第三个含dix结构域的蛋白。我们发现Ccd1在体外和体内激活Wnt/ β -catenin通路，并调节斑马鱼胚胎的神经模式。我们还发现许多Ccd1异构体是通过小鼠的不同启动子使用和选择性剪接产生的。其中，一种名为Ccd1A的新型异构体，除了在原始异构体（名为Ccd1B）中发现的coil -coil和DIX结构域外，在n端还含有一个钙钙蛋白同源结构域（CH）。从斑马鱼中，我们分离到了编码A和B亚型的第二基因Ccd2。当在培养细胞系中异种表达时，Ccd1B蛋白以小泡的形式存在于细胞中，Ccd2A蛋白与肌动蛋白纤维共定位。此外，disheveled （Dvl）蛋白的细胞质定位被Ccd1A和Ccd1B的强制表达所改变，这表明Ccd1蛋白在细胞中与Dvl相互作用。虽然我们产生了敲除小鼠，但它们没有表现出任何明显的表型。当我们在斑马鱼胚胎中过表达Ccd2时，它们表现出卷曲的尾部向下表型，这表明Ccd1在中线信号传导中起作用。