Genetic dissection of peripheral glia and glial sheath development

概述：我的研究计划的长期目标是研究包裹和隔离周围神经的神经胶质鞘发育过程中的蛋白质相互作用。周围神经对环境信号做出反应并控制肌肉收缩，周围环绕着神经胶质细胞层，神经胶质细胞是神经系统的主要绝缘体。轴突周围神经胶质鞘的形成是所有动物神经系统发育和功能的重要组成部分。人们对触发神经胶质细胞迁移并包裹其相关轴突的机制知之甚少。神经胶质细胞的迁移和围绕轴突的神经胶质包裹的形成都涉及细胞间的蛋白质黏附和通讯，导致细胞骨架的广泛重排。神经胶质细胞必须从细长的迁移细胞分化为具有精心设计的延伸的细胞，这些细胞包围着它们包裹的轴突。在果蝇中，三个不同的神经胶层构成成熟的外周神经。在后期阶段，神经胶质鞘必须保持和扩张，以适应动物的生长，以确保基本轴突的完全隔离。神经胶质鞘的丧失会导致神经元功能的中断和致命性的破坏。 我们之前由NSERC资助的工作表明，一类被称为整合素的黏附蛋白及其在细胞内的结合伙伴在建立和维持神经胶质鞘方面起着关键作用。我们发现，在周围神经系统(PNS)中不同的胶质细胞层之间的整合素通讯对于维持胶质细胞的功能和隔离神经胶质细胞膜至关重要。但整合素复合体如何控制神经胶质鞘的形成尚不清楚。我们有证据表明，在周围神经的核心，整合素介导的黏附不是通过细胞外基质成分，而是通过膜结合的配体。我们对潜在整合素配体的研究发现，basigin是一种跨膜Ig结构域蛋白，对胶质细胞与胶质细胞的黏附至关重要，以维持胶质细胞的包裹和胶质层。我们NSERC资助的研究还发现了缝隙连接蛋白Innexin 2在胶质鞘的发育或维持中的关键作用，这些数据也指出了胶质细胞-胶质细胞通讯的关键作用。但是，无论是作为缝隙连接还是作为黏附蛋白，Innexin 2如何促进鞘形成尚不清楚。因此，我们的研究计划正在巩固这一中心主题，即轴突周围胶质鞘的发育和维持不仅是神经胶质细胞到轴突通讯的功能，也是神经胶质细胞到神经胶质细胞通讯的功能。我们的短期目标将解决作为这种沟通基础的机制。 方法：我的实验室有丰富的经验，使用分子和细胞生物学结合高分辨率成像分析周围神经系统神经胶质细胞的分子和发育动力学。我们已经产生了一系列细胞和遗传标记，使我们能够操纵外围神经胶质细胞中表达的基因，然后在发育过程中评估结果。因此，我们可以使用果蝇模型系统来分离和表征所有动物中神经胶质细胞发育和鞘形成的全球基本机制。 目的：研究整合素介导的神经胶质鞘形成和维持的分子机制，以及包括basigin在内的Ig结构域蛋白在此过程中的作用。我们将使用功能丧失和挽救的方法来确定innexin 2如何在神经胶质鞘中发挥作用，作为缝隙连接，以及通过什么信号机制。

Overview: The long-term goal of my research program is to study the protein interactions that underlies the development of the glial sheath that ensheaths and insulates the peripheral nerve. The peripheral nerve responds to environment cues and controls muscle contraction and are surrounded by layers of glial cells, the major insulator of the nervous system. The formation of a glial sheath around axons is an essential component of the development and function of the nervous system in all animals. Little is known about the mechanisms that trigger glial cells to migrate along and then wrap their associated axons. Both glial migration and the formation of the glial wrap around axons involves protein adhesion and communication between cells leading to extensive rearrangements of the cytoskeleton. Glial cells must differentiate from elongated migrating cells into cells with elaborate extensions that encompass the axons they wrap. In Drosophila three distinct glial layers contribute to the mature peripheral nerve. In later stages the glial sheath must be maintained and expand to match animal growth to ensure complete insulation of the underlying axons. Loss of the glial sheath leads to a disruption of neuronal function and lethality. Our previous work funded by NSERC has shown that a class of adhesion proteins called the integrins and their binding partners within the cell play a critical role in establishing and maintaining the glial sheath. We found that integrin communication between the distinct glia layers in the peripheral nervous system (PNS) is critical to maintain glia function and insulating glia wrap. But how the integrin complex functions to control glia sheath formation is not known. We have evidence within the core of the peripheral nerve, integrin mediated adhesion is not through extracellular matrix components but rather through membrane bound ligands. Our investigation of potential integrin ligands identified Basigin, a transmembrane Ig domain protein, as critical for glia-glial adhesion to maintain the glial wrap and the glial layers. Our NSERC funded research has also uncovered a critical role of the gap junction protein, Innexin 2, in the development or maintenance of the glial sheath and these data also point to an essential role for glia-glia communication. But how Innexin 2 contributes to sheath formation either as a gap junction or an adhesion protein is not known. Therefore our research program is consolidating on the central theme that the development and maintenance of the glia sheath around axons is a function of not just of glia to axon communication but also of glia to glia communication. Our short terms goals will address the mechanisms that underlie this communication. Approach: My laboratory has extensive experience analyzing the molecules and developmental dynamics of the glia of the peripheral nervous system using a combination of molecular and cell biology paired with high resolution imaging. We have generated a collection of cellular and genetic markers that allow us to manipulate the genes expressed in the peripheral glia and then assess the results during the development. Therefore we can use the Drosophila model system to isolate and characterize the global fundamental mechanisms that underlie glial cell development and sheath formation in all animals. Objectives: We will investigate the molecular mechanisms that underlie integrin mediated glial sheath formation and maintenance, and the role of the Ig domain proteins including Basigin in this process. We will use a loss of function and rescue approach to determine how innexin 2 functions in the glial sheath, as a gap junction and through what signaling mechanism.