MOLECULAR REGULATION OF NEOCORTICAL NEUROGENESIS

描述(由申请人提供)：对啮齿动物和人类的自发突变的分析已被证明在识别大脑发育所必需的分子机制方面具有非常宝贵的价值。最值得注意的是，通过遗传分析和位置克隆已经确定了对大脑皮层神经元模式迁移至关重要的蛋白质。相比之下，很少有自发突变导致识别参与控制细胞质分裂和相关细胞动力学的通路，这些通路发生在脑室表面的神经发生过程中。胞质分裂的调节在新皮质神经发生中起着核心作用，并可能决定新生子细胞的命运。在这笔赠款的第一阶段，我们发现了一种新的突变体--平头，它现在揭示了一种基本的分子机制，它可以调节神经元前体细胞的胞质分裂和神经发生。该扁头突变体的CNS特异性表型特征是严重的小脑，不同类型神经细胞的相对数量发生变化，以及异常的神经元肥大。用定位克隆的方法将该扁头突变鉴定为位于大鼠和人12号染色体长臂上的CitronK基因突变。该扁头突变是外显子1的单碱基缺失，导致增殖区无意义密码子和CitronK蛋白表达。CitronK蛋白主要集中在VZ表面的贴壁和胞质皱纹上。我们假设，CitK包含多个蛋白质相互作用结构域，将决定命运的信号与VZ表面的细胞骨架重组联系在一起。我们建议结合成像实验、体内转基因和生化实验来进一步阐明神经发生所必需的细胞和分子途径。

DESCRIPTION (provided by applicant): Analysis of spontaneous mutations in rodents and humans has proven invaluable in identifying molecular mechanisms essential to brain development. Most notably, proteins essential to patterned migration of cortical neurons have been identified through genetic analyses and positional cloning. In contrast, there have been few spontaneous mutations that have led to the identification of pathways involved in the control of cytokinesis and associated cellular dynamics that occur throughout neurogenesis at the surface of the ventricles. The regulation of cytokinesis plays a central role in neocortical neurogenesis, and may determine the fates of newly generated daughter cells. In the first period of this grant we discovered a novel mutant, flathead, which has now revealed an essential molecular mechanism that operates to regulate cytokinesis and neurogenesis in neuronal progenitors. The CNS-specific phenotype of the flathead mutant is characterized by severe micrencephaly with alterations in the relative numbers of different neuronal cell types, and in abnormal neuronal hypertrophy. A positional cloning strategy was used to identify the flathead mutation as a mutation in the Citron-kinase gene (CitronK or CitK) located on the long arm of rat and human chromosome 12. The flathead mutation is a single base deletion in exon 1 that results in a nonsense codon and absence of CitronK protein expression in proliferative zones. CitronK protein is concentrated at the VZ surface at adherens unctions and cytokinesis furrows. We hypothesize that CitK, which contains multiple protein interaction domains, links fate determining signals with cytoskeletal restructuring at the VZ surface. We propose to use a combination of imaging experiments, in vivo transfection, and biochemical experiments to further elucidate a cellular and molecular pathway necessary for neurogenesis.