Tubulin Mutations in Neuronal Migration Disorders

神经元迁移障碍中的微管蛋白突变

• 批准号: 8293824
• 负责人: NICHOLAS COWAN
• 金额: $ 48.29万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-01 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8293824
• 关键词: Alleles; Animal Model; Behavior; Binding; Biochemical; Biological; Biological Assay; Biological Models; Brain; Cell-Free System; Cells; Cognitive; Complex; Computing Methodologies; Cultured Cells; Data; Defect; Development; Disease; Embryo; Environment; Escherichia coli; Event; Failure; Genes; Histocytochemistry; Homologous Gene; Human; In Vitro; Induced Mutation; Knock-in Mouse; Lead; Mammals; Maps; Measures; Methods; Microtubule-Associated Proteins; Microtubules; Molecular; Molecular Chaperones; Monitor; Multigene Family; Mus; Mutation; Mutation Spectra; Neuronal Migration Disorder; Neurons; Pathway interactions; Pattern; Play; Polymerase; Population; Process; Property; Protein Binding; Proteins; Rattus; Recombinants; Resolution; Role; Structure; Testing; Time; Tissues; Transgenic Animals; Transgenic Mice; Tubulin; Work; base; chaperonin CCT; cytosolic chaperonin; disability; disease-causing mutation; in vivo; migration; mouse model; mutant; polypeptide; research study

DESCRIPTION (provided by applicant): Microtubules are assembled from ¿/¿ tubulin heterodimers. In mammals, a small multigene family encodes ¿- and ¿-tubulins, with each gene product (termed an isotype) having a distinct developmentally regulated and tissue-specific pattern of expression. Recently, mutations have been identified in genes (e.g. TUBA1A and TUBB2B) encoding these isotypes that cause certain human neuronal migration disorders, all of which are associated with severe cognitive disabilities. These discoveries reinforce the notion that microtubule based events play a central role in neuronal migration during brain development, and that disruption of critical microtubule processes results in cortical dysgeneses. Analysis of the properties of disease causing mutations in TUBA1A suggests two broad classes of mechanism: 1. Defects in the complex tubulin heterodimer assembly pathway This pathway involves sequential interactions of newly synthesized ¿- and ¿-tubulin polypeptides with multiple chaperone proteins (including the cytosolic chaperonin CCT and five tubulin specific chaperones, TBCA-TBCE). CCT facilitates productive folding by providing a sequestered environment in which folding can occur in the absence of off-pathway interactions that might otherwise lead to aggregation, while the tubulin specific chaperones function downstream of CCT as an ¿/¿ tubulin heterodimer assembly machine. 2. Defective microtubule dynamics and/or interactions with Microtubule Associated Proteins (MAPs) Disease causing mutations might compromise microtubule dynamics or interfere with interaction(s) between microtubules and MAPs that are critical for directing proper neuronal migration. The experiments proposed here constitute a multifaceted approach towards understanding the mechanism of these diseases. 1) We will exploit the mutation-induced defective interactions between CCT-generated ¿-tubulin folding intermediates and TBCB to define this interaction. These experiments will establish the mechanism of action of TBCB as a critical player in proper corticogenesis. 2) We will generate populations of isotypically homogeneous wild type and mutant tubulin heterodimers. These will be used to identify MAPs (such as the microtubule polymerase TOGp) that bind differently to microtubules polymerized from wild type and disease-causing mutant heterodimers. 3) For those disease-causing tubulin mutations that do not interfere with the heterodimer assembly machinery, we will examine their effect on microtubule dynamics in single cell experiments (including cultured neurons) in vivo. 4) The mechanism of disease will be explored via the construction and analysis of transgenic mice in which one copy of wild type tuba1a (the mouse homolog of TUBA1A) is replaced with the same allele harboring a disease-causing mutation. The brains of these mice will be examined in terms of their development and the behavior of their microtubules in vitro and in vivo. PUBLIC HEALTH RELEVANCE: We will establish the molecular basis of naturally occurring neuronal migration diseases caused by mutations in the ¿-tubulin gene TUBA1A. We will use structural, biochemical, cell biological and animal model approaches in order to understand the significance of these mutations in terms of the development of the mammalian brain.

描述（由应用程序提供）：微管由微管蛋白异二聚体组装。在哺乳动物中，一个小的多基因家族编码„ - 和 - tubulins，每个基因产物（称为同种型）具有独特的发育调节和组织特异性表达模式。最近，已经在编码这些同种型的基因（例如Tuba1a和Tubb2b）中鉴定出突变，这些同种型引起某些人类神经元迁移障碍，所有这些都与严重的认知障碍有关。这些发现加强了以下观点：基于微管的事件在脑发育过程中在神经元迁移中起着核心作用，而关键微管过程的破坏会导致皮质失调。对TUBA1A中引起突变的疾病特性的分析提出了两种广泛的机制：1。复杂的小节蛋白异二聚体组装中的缺陷，该途径涉及新合成的�-和 - tubul蛋白多肽与多链酮蛋白（包括细胞蛋白蛋白蛋白）的顺序相互作用。 CCT通过提供一个测序的环境来促进产品折叠，在该环境中可能会在没有校园相互作用的情况下进行折叠，否则可能会导致聚集，而小管蛋白特异性链酮在CCT下游起作用，将CCT作为« /»tubelin heterodimer组装机。 2。有缺陷的微管动力学和/或与微管相关蛋白（MAP）疾病引起突变的疾病可能会损害微管动力学或在微管和地图之间对指导正确神经元迁移至关重要的相互作用的干扰。这里提出的实验构成了一种多方面的方法，用于理解这些疾病的机制。 1）我们将利用CCT生成的–微型蛋白折叠中间体和TBCB之间的突变引起的缺陷相互作用来定义这种相互作用。这些实验将确定TBCB作为适当皮质生成的关键参与者的作用机理。 2）我们将产生同种均质野生型和突变型管蛋白异二聚体的种群。这些将用于识别与与野生型和引起疾病的突变异二聚体聚合的微管不同结合的图（例如微管聚合酶TOGP）。 3）对于那些不干扰异二聚体组装机械的那些引起疾病的小动蛋白突变，我们将检查它们对体内单细胞实验（包括培养的神经元）中微管动力学的影响。 4）将通过对转基因小鼠的构建和分析来探索疾病的机制，其中一份野生型管1a副本（Tuba1a的小鼠同源物）被带有引起疾病突变的同类等位基因代替。这些小鼠的大脑将根据其发育以及其在体外和体内的微管的行为来检查。 公共卫生相关性：我们将建立由 - 小氨基蛋白基因tuba1a突变引起的天然神经元迁移疾病的分子基础。我们将使用结构，生化，细胞生物学和动物模型方法，以了解这些突变在哺乳动物大脑的发展方面的重要性。