Role of Cdc42 and Par6 Polarity Complex in CNS Neuronal Migration

Cdc42 和 Par6 极性复合物在 CNS 神经元迁移中的作用

• 批准号: 8627650
• 负责人: Mary Elizabeth Hatten
• 金额: $ 36.6万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-02 至 2016-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8627650
• 关键词: Adhesions; Affect; Architecture; Autistic Disorder; Biological Models; Brain; Brain Diseases; Brain region; Cell Adhesion Molecules; Cell Nucleus; Cell Polarity; Cells; Centrosome; Chimeric Proteins; Code; Complex; Computer software; Cortical Malformation; Cre-LoxP; Cytoplasmic Granules; Development; Diagnosis; Disease; Epilepsy; Exhibits; Extracellular Domain; Fiber; Fluorescence Resonance Energy Transfer; Genetic; Goals; Grant; Health; Human; Image; Immigration; Integrins; Life; LoxP-flanked allele; Maintenance; Mental Retardation; Molecular; Monitor; Motor; Movement; Mus; Myosin ATPase; Neuroglia; Neuronal Migration Disorder; Neurons; PHluorin; Pathway interactions; Pattern; Phenotype; Positioning Attribute; Process; Protein Binding; Protein Isoforms; Proteins; Relative (related person); Reporter; Research; Resolution; Role; Signal Pathway; Surface; Synapses; System; Takeda brand of pioglitazone hydrochloride; Testing; Time; Tubulin; adhesion receptor; axon guidance; base; cdc42 GTP-Binding Protein; cell motility; cell type; granule cell; insight; interstitial; loss of function; migration; mutant; neuronal cell body; progenitor; receptor; reconstruction; research study; spatiotemporal; trafficking

DESCRIPTION (provided by applicant): The development of the mammalian brain depends on the migrations of neuronal precursors from germinal zones, where they are generated, and their assembly into neuronal laminae, where synaptic connections form. Since CNS migration disorders are associated with a number of cortical malformations, and are a major cause of disease in the developing human brain, including mental retardation and epilepsy, a clearer understanding of the molecular control of CNS neuronal migration could be relevant to the diagnosis and treatment of human developmental brain disorders. Neuronal migration critically depends on the polarization of the neuron in the direction of moment, and with support from this grant, we previously established that the conserved mPar61 polarity complex localizes to the centrosome and coordinates the forward movement of the centrosome and soma, by a mechanism that includes activation of acto-myosin contractile motors in the proximal region of the leading process, in migrating cerebellar granule neurons. The overall goal of the proposed research is to define the relative contributions of the master polarity regulator Cdc42 and the three Par6 isoforms to critical steps in CNS migration, including the formation and maintenance of a highly polarized leading process, and to define their role(s) in receptor trafficking of neuronal adhesion proteins. Although a clear role for Cdc42 has been established in the migration of many non-neuronal cells, and in dendritic arborization and axon guidance, the roles of Cdc42 and the relative role of the Par6 isoforms have not been analyzed in high-resolution time-lapse imaging of live, migrating CNS neurons. Given the importance of glial-guided migration to the formation of neuronal layers in cortical regions of brain, we will focus on this highly specialized migration system, using cerebellar granule neurons migrating on glia as our model system. In Aim 1, we will study a conditional loss of function of Cdc42; a similar plan will be implemented in Aim 3 for each of the three Par6 isoforms expressed in granule cell progenitors (GCPs). As a complementary approach, in Aims 2 & 3, we will use siRNAs and shRNAs to knockdown Cdc42 and Par6 isoform levels and compare knockdown phenotypes with conditional loss of function phenotypes. If Cdc42 is in the same genetic pathway with any or all of the Par6 isoforms, we would expect to see similar phenotypes for Cdc42 and Par6 isoform loss of function. We will also use Raichu probes for Cdc42, kindly provided by Dr. Miki Matsuda, which are FRET-based probes that monitor Cdc42 activation in localized regions of a cell. These probes will enable us to evaluate the spatiotemporal localization of Cdc42 activation relative to the Par6 isoforms in migrating GCPs. The discovery of changing patterns of Par6 expression during cerebellar development is an exciting opportunity to understand their relative contributions to neuronal migration and whether they act within a Cdc42 signaling pathway.

描述（由申请人提供）：哺乳动物大脑的发育取决于神经元前体从产生它们的生发区的迁移，以及它们组装成突触连接形成的神经元层。由于中枢神经系统迁移障碍与许多皮质畸形有关，并且是人类大脑发育中疾病的主要原因，包括智力低下和癫痫，因此更清楚地了解中枢神经系统神经元迁移的分子控制可能有助于诊断和治疗。治疗人类大脑发育障碍。神经元迁移关键取决于神经元在矩方向上的极化，在这项资助的支持下，我们之前确定保守的 mPar61 极性复合物定位于中心体，并通过一种机制协调中心体和体细胞的向前运动这包括在迁移的小脑颗粒神经元中引导过程的近端区域激活肌球蛋白收缩运动。拟议研究的总体目标是确定主极性调节剂 Cdc42 和三种 Par6 同工型对 CNS 迁移关键步骤的相对贡献，包括高度极化主导过程的形成和维持，并确定它们的作用）在神经元粘附蛋白的受体运输中。尽管 Cdc42 在许多非神经元细胞的迁移以及树突分枝和轴突引导中的明确作用已被确定，但 Cdc42 的作用和 Par6 亚型的相对作用尚未在高分辨率延时中进行分析活体迁移中枢神经系统神经元的成像。鉴于神经胶质引导的迁移对大脑皮质区域神经元层形成的重要性，我们将重点关注这种高度专业化的迁移系统，使用在神经胶质上迁移的小脑颗粒神经元作为我们的模型系统。在目标 1 中，我们将研究 Cdc42 的功能有条件丧失；目标 3 中将针对颗粒细胞祖细胞 (GCP) 中表达的三种 Par6 同工型中的每一种实施类似的计划。作为补充方法，在目标 2 和 3 中，我们将使用 siRNA 和 shRNA 来敲低 Cdc42 和 Par6 同工型水平，并将敲低表型与条件性功能丧失表型进行比较。如果 Cdc42 与任何或所有 Par6 同工型处于同一遗传途径，我们预计会看到 Cdc42 和 Par6 同工型功能丧失的相似表型。我们还将使用由 Miki Matsuda 博士友情提供的 Cdc42 Raichu 探针，它们是基于 FRET 的探针，可监测细胞局部区域的 Cdc42 激活。这些探针将使我们能够评估迁移 GCP 中 Cdc42 激活相对于 Par6 同工型的时空定位。小脑发育过程中 Par6 表达模式变化的发现是一个令人兴奋的机会，可以了解它们对神经元迁移的相对贡献以及它们是否在 Cdc42 信号通路内发挥作用。