Dishevelled-Mediated Control of Wnt/PCP Pathways

描述（由申请人提供）：高度保守的经典和非经典Wnt通路对哺乳动物发育的所有方面都很重要，包括中枢神经系统的发育。一个悬而未决的问题仍然是：如何在体内整合的各种Wnt途径，调节发展？Dvl是解决这个问题的杰出候选者，因为这些保守蛋白在所有真核生物中对于经典和非经典Wnt途径都是必需的。我们已经发现了三个Dvl基因中部分独特但主要是冗余的功能。单突变体在社会行为和圆锥干心脏发育方面显示出一些独特的缺陷，而双Dvl突变体显示出严重的神经管缺陷（颅脊柱裂）和严重的耳蜗缺陷。在冗余的进一步支持中，Dvl 1/2/3三重突变体不能经历原肠胚形成并且不形成中胚层。我们计划剖析Dvl调节正常发育并在Dvl突变体中被破坏以产生这些表型的体内途径。我们在小鼠中产生每个Dvl基因的体内条件等位基因以及可以区分典型Wnt或非典型Wnt/PCP途径功能的体内等位基因。我们使用这些等位基因提供明确的证据表明，颅脊柱裂表型显示Dvl 1; Dvl 2双突变体的收敛延伸运动通过Wnt/PCP途径的破坏。我们将使用这些工具，提供一个全面的分析的作用，典型的Wnt和非典型的Wnt/PCP途径在神经元发育过程中，从第一次发展的神经褶皱在原肠胚形成和神经形成整个神经发生和神经元迁移。根据我们发表的和初步的数据，我们预测Dvls和它们调节的通路在大脑发育的所有阶段都是至关重要的。我们将使用以下具体目标：1）确定体内原肠胚形成过程中经典Wnt和非经典Wnt/PCP通路的作用; 2）表征神经胚形成过程中负责神经管闭合的Dvl依赖性通路; 3）使用双Dvl突变体确定Dvl和经典Wnt和非经典Wnt/PCP通路在前脑/中脑-后脑发育中的作用;和4）使用三重Dvl突变体确定Dvl和经典Wnt和非经典Wnt/PCP途径在前脑/中脑-后脑发育期间的作用。 公共卫生相关性：了解细胞机制和途径，介导的Wnt途径使用Dvl突变小鼠神经元发育可能会提供重要的见解人类神经管缺陷和中枢神经系统的发展。使用复杂的小鼠突变体，这些突变体使每个Dvl突变或表达条件或突变等位基因，这些等位基因表达荧光标记的蛋白质，将允许以在人类中不可能的方式详细研究这些机制和途径。

DESCRIPTION (provided by applicant): The well-conserved canonical and non-canonical Wnt pathways are important for all aspects of mammalian development, including the development of the central nervous system. An outstanding question remains: how are the various Wnt pathways that regulate development integrated in vivo? Dvls are outstanding candidates to address this question, since these conserved proteins are required in all eukaryotes for both canonical and non-canonical Wnt pathways. We have uncovered partially unique but predominantly redundant functions among the three Dvl genes. Single mutants display some unique defects in social behavior and conotruncal heart development, while double Dvl mutants display severe neural tube defects (craniorachischisis) and severe cochlear defects. In further support of redundancy, Dvl1/2/3 triple mutants are unable to undergo gastrulation and do not form mesoderm. We plan to dissect the in vivo pathways that Dvls regulate normal development and are disrupted in the Dvl mutants to produce these phenotypes. We produced in vivo conditional alleles in mice for each of the Dvl genes as well as in vivo alleles that can distinguish either canonical Wnt of non-canonical Wnt/PCP pathway function. We used these alleles to provide definitive evidence that the craniorachischisis phenotype displayed by Dvl1;Dvl2 double mutants resulted from disruption of convergent extension movements via the Wnt/PCP pathway. We will use these tools to provide a comprehensive analysis of the role of the canonical Wnt and non-canonical Wnt/PCP pathways during neuronal development from the first development of neural folds during gastrulation and neurulation throughout neurogenesis and neuronal migration. Based on our published and preliminary data, we predict that Dvls and the pathways they regulate are critical at all stages of brain development. We will use the following specific aims: 1) Determine the role of canonical Wnt and non-canonical Wnt/PCP pathways during gastrulation in vivo; 2) Characterize the Dvl dependent pathways responsible for neural tube closure during neurulation; 3) Determine the role of Dvls and the canonical Wnt and non-canonical Wnt/PCP pathways during forebrain/midbrain-hindbrain development using double Dvl mutants; and 4) Determine the role of Dvls and the canonical Wnt and non-canonical Wnt/PCP pathways during forebrain/midbrain-hindbrain development using triple Dvl mutants. PUBLIC HEALTH RELEVANCE: Understanding cellular mechanisms and pathways that mediate neuronal development by the Wnt pathways using Dvl mutant mice will likely provide important insights into human neural tube defects and the development of the central nervous system. The use of sophisticated mouse mutants that inactivate each of the Dvls or express conditional or mutant alleles that express fluorescently tagged proteins will allow for the detailed study of these mechanisms and pathways in ways that are impossible in the human.