Probing Microtubule Function in Neuronal Development

神经元是多细胞生物体中形态最复杂的细胞类型，这种复杂性是 错综复杂地与神经系统信息处理有关。因此，神经元的损伤 形态发生总是会导致神经系统功能的改变，并经常削弱神经功能 疾病。对微管细胞骨架组织的精确控制是神经元大部分(如果不是全部)的核心 发育和功能的范围从新生神经元的迁移，轴突的延长周期， 向生长锥体引导和突触形成的回缩和分支。神经元的这种重要性 微管反映在广泛的神经发育和神经退行性疾病中，与 与微管细胞骨架相关的蛋白质中的遗传缺陷。人类遗传学与现代 测序技术发现了与频繁重症相关的大量基因突变 皮质畸形。这些疾病包括微管蛋白基因本身的两种突变--所谓的微管蛋白病 -以及神经元微管相关蛋白的突变，这些蛋白通常具有类似的范围 神经发育表型。虽然通常认为这些突变会扰乱发育 未成熟神经元通过发育中的皮质迁移，我们不了解支配规则 MT在神经形态发生中的作用在机制水平上。在此应用程序中，我们建议使用Emerging 结合最新基因组的人诱导多能干细胞(IPSC)技术 工程学和我们在定量显微镜方面的成熟专业知识来模拟和解剖神经细胞 肾小管病样疾病的生物学和动态MTS在神经形态发生中的作用。在目标1中，我们 关注导致皮质畸形的DCX和微管蛋白突变，我们将分析DCX是如何控制的 基于我们最近的数据，神经元微管动力学和力学，以及神经元的形态发生 DCX以一种独特的几何依赖方式结合微管。在目标2中，我们使用新的光遗传学来 控制蛋白质与生长中的微管和末端的相互作用，以秒和微米精度绘制地图 微管+末端复合体如何促进神经元发育动力学。我们相信 定量和严格地理解MT在神经元发育和发育中的作用原理 神经发育疾病的问题将对人类健康产生切实而重要的后果， 并可为未来的治疗方法奠定基础。

Neurons are the most morphologically complex cell type in multicellular organisms, and this complexity is intricately linked to the complexity of nervous system information processing. Thus, impairments in neuronal morphogenesis invariably result in alterations of nervous system function and often debilitating neurological disease. Precise control of microtubule cytoskeleton organization is central to most if not all aspects of neuron development and function ranging from the migration of newborn neurons, cycles of neurite elongation, retraction and branching to growth cone guidance and synapse formation. This importance of neuronal microtubules is reflected in the wide range of neurodevelopmental and neurodegenerative diseases linked to genetic defects in proteins associated with the microtubule cytoskeleton. Human genetics and modern sequencing techniques identified numerous mutations in related genes associated with frequently severe cortical malformations. These include both mutations of tubulin genes themselves – so-named tubulinopathies – as well as mutations in neuronal microtubule-associated proteins that often have a similar range of neurodevelopmental phenotypes. While it is generally assumed that these mutations disrupt the developmental migration of immature neurons through the developing cortex, we do not understand the rules governing MT function in neuromorphogenesis at a mechanistic level. In this application, we propose to use emerging human induced pluripotent stem cell (iPSC) technology in combination with state-of-the-art genome engineering and our established expertise in quantitative microscopy to model and dissect the neuronal cell biology of tubulinopathy-like diseases and the function of dynamic MTs in neuromorphogenesis. In Aim 1, we focus on DCX and tubulin mutations that cause cortical malformations, and we will analyze how DCX controls neuronal microtubule dynamics and mechanics, and neuronal morphogenesis based on our recent data that DCX binds microtubules in a unique geometry-dependent way. In Aim 2, we employ novel optogenetics to control protein interactions with growing microtubule plus ends with second and micrometer precision to map how microtubule plus end complexes contribute to neuronal development dynamics. We believe that quantitative and rigorous understanding of the principles that govern MT function in neuronal development and what goes wrong in neurodevelopmental disease will have tangible and important outcomes for human health, and can lay the foundation for future therapeutic approaches.