Neuronal primary cilium: a centralized signaling node for dendrite morphogenesis

神经元树突的适当大小和形状是神经元如何接收、整合和编码电路信息的关键决定因素。我们最近有了一个意想不到的发现，神经元初级纤毛是一个非常短(只有几米长)的信号天线，从神经元胞体伸出，是一个集中的信号节点，强烈地影响树突的形态。NSERC DG研究计划的目标是确定神经元初级纤毛在神经元形态发生和连接中的作用。 神经元的初级纤毛具有不同的信号成分，作为信号中枢，并将环境信号传递到胞体，以实现主要的细胞功能。然而，由于缺乏精确控制纤毛特异性信号事件的方法，纤毛信号在整个大脑发育和功能中的确切作用仍然是个谜。我的工作克服了这一技术限制，并建立了一个在发育中的神经元中询问纤毛信号的实验框架，我们将在这个新的NSERC项目中充分利用这一点。 在前期工作中，我们使用了Arl13b的小鼠模型突变体，Arl13b是一种纤毛特异的小GTP酶，对纤毛信号转导至关重要。我们发现，锥体神经元中纤毛信号的中断显著损害了大脑皮层中的树突树枝。这一建议解决了一个关键问题：初级纤毛信号如何对树突形态发生产生如此深远的影响？在三个目标中，我们将： 目的1.研究Arl13b突变型皮质神经元树突状突起的生长和形态变化。高分辨率共聚焦显微镜将用于树枝状形态的详细表征。采用延时实时成像技术分析树枝生长动态。 目的2.阐明纤毛驱动的细胞内信号转导机制在树突状细胞形态发生中的作用。我们将检验这一假设，即纤毛-PI3K(磷脂酰肌醇3-激酶)信号在调节树突发育中发挥核心作用。我们将产生新的光遗传学工具来剖析纤毛-PI3K信号级联如何控制树突生长动力学和形态。 目的3.确定调控树突状细胞形态发生的纤毛信号受体。为此，我们将重点研究已知的影响树突形态的纤毛受体，以及Arl13b突变纤毛中的错误定位。光遗传诱导的纤毛受体激活将被用来直接检测这些受体对树突形态发育的影响。 每个目标都是专门为HQP提供神经发育、分子遗传学和新的实验工具开发和实施方面的不同培训。完成这些目标后，我们将获得变革性的机制洞察力，了解神经元纤毛如何在发育中的大脑中以前所未有的水平发挥作用。

The appropriate size and shape of neuronal dendritic arbors is a key determinant of how neurons receive, integrate, and encode circuit information. We recently made an unexpected finding that the neuronal primary cilium, a remarkably short (just a few m long), singular signaling antenna that protrudes from the neuronal soma, is a centralized signaling node that robustly influences dendrite morphology. The goal of this NSERC DG Research Program is to define the role of the neuronal primary cilium in neuronal morphogenesis and connectivity. The neuronal primary cilium possesses a diverse cast of signaling components to act as a signaling hub and relay of environmental signals to the soma for major cell functions. However, due to a lack of methods to precisely control cilia-specific signaling events, the exact role of ciliary signaling in overall brain development and function remains enigmatic. My work has overcome this technical limitation and established an experimental framework to interrogate ciliary signaling in developing neurons, which we will fully exploit in this new NSERC project. In preliminary work, we used a mouse model mutant for Arl13b, a ciliary-specific small GTPase critical for ciliary signal transduction. We found that disruption of ciliary signaling in pyramidal neurons significantly impairs dendritic arborization in the cerebral cortex. This proposal addresses a key question: How does primary ciliary signaling exert such profound effects on dendrite morphogenesis? In three aims, we will: Aim 1. To characterize the altered dendritic arbor growth and patterning in Arl13b mutant cortical neurons. High-resolution confocal microscopy will be performed for a detailed characterization of the dendritic morphology. Time-lapse live imaging will be performed to analyze the dendritic arbor growth dynamics. Aim 2. To delineate the ciliary-driven intracellular signaling mechanisms in dendrite morphogenesis. We will test the hypothesis that the ciliary-PI3K (Phosphoinositide 3-kinase) signaling plays a central role in modulating dendrite development. We will generate novel optogenetic tools to dissect how ciliary-PI3K signaling cascades govern dendritic growth dynamics and morphology. Aim 3. To define the ciliary signaling receptors that modulate dendrite morphogenesis. We will focus on the ciliary receptors known to influence dendrite morphology and mislocalized in Arl13b mutant cilia for this aim. Optogenetically induced ciliary receptor activation will be used to directly examine these receptors' influence on dendrite morphology development. Each aim is specifically designed to provide HQP with diverse training in neurodevelopment, molecular genetics, and novel experimental tool development and implementation. Upon completion of these aims, we will have gained transformative mechanistic insights into how neuronal cilia function in the developing brain at a level never before achieved.