Mechanisms of Node of Ranvier Assembly

有髓轴突的兰维尔结节是跳跃性传导中动作电位传播的关键，也是神经元微妙的区域组织特征的显著例子。结点由多聚体复合体和细胞骨架复合体组成，多聚体复合体包括电压门控钠通道和钾通道、辅助β亚基、细胞黏附分子(CAM)，细胞骨架复合体包括与通道和CAM结合的骨架蛋白G。我们最近的研究表明，节点组装来自不同的来源。CAM，特别是NF186，通过与雪旺细胞相互作用的扩散捕获，从轴突表面的现有池重新分布，在形成节点时积累。相反，通道和细胞骨架蛋白主要通过轴突运输传递到结节。令人惊讶的是，似乎还有一个钠通道池，它独立于运输和Ankyrin G在结节上运输和积累。最后，我们的研究表明，有一个积极的计划来从结外位置清除结节蛋白，进一步加强它们在结节处的选择性浓缩。为了扩展这些发现并进一步阐明结节组装的机制，我们将调查结节组件的运输和组装，以及它们如何在髓鞘共培养中通过活体成像策略从结外部位清除。特别是，我们将：i)确定将组件运输到节点的小泡是否分离为专用于CaM的小泡和其他专用于离子通道的小泡，并显示任何区域特异性；ii)表征依赖运输和非运输的钠通道组件到节点的运输，包括ankyrin G和NF186的分别作用，并确定钠通道复合体是否在局部组装；以及i)研究如何将节点成分从结外部位清除，重点是NF186以进一步检查其细胞质片段以及内吞和蛋白降解在清除NF186的节点间表面池中的作用。相关性：这些研究将阐明调节兰维尔结节组装的机制，这对神经纤维适当地传导电脉冲的能力至关重要。因此，这项研究的发现可能对我们理解有髓纤维疾病的发病机制具有重要意义，包括导致神经传导异常的神经疾病，从而可能导致这些神经疾病的新治疗策略。

The nodes of Ranvier of myelinated axons are critical for action potential propagation by saltatory conduction and are a striking example of the exquisite domain organization characteristic of neurons. Nodes are comprised of a multimeric complex that includes voltage gated sodium and potassium channels, accessory beta subunits, cell adhesion molecules (CAMs), and a cytoskeletal complex that includes ankyrin G to which channels and CAMs bind. Our recent studies indicate that the node assembles from distinct sources. CAMs, notably NF186, accumulate at forming nodes by redistribution from existing pools on the axon surface via diffusion trapping from interactions with the Schwann cell. In contrast, channels and cytoskeletal proteins are delivered to the node primarily via axonal transport. Surprisingly, there also appears to be a pool of sodium channels that traffics and accumulates at the node independent of transport and of ankyrin G. Finally, our studies suggest there is an active program to clear nodal proteins from extranodal sites that further reinforces their selective enrichment at the node. To extend these findings and further elucidate the mechanisms of node assembly, we will investigate trafficking and assembly of components of the node, and how they are cleared from extranodal sites, by live imaging strategies in myelinating cocultures. In particular, we will: i) determine whether vesicles that transport components to the nodes segregate into those specific for CAMs and others for ion channels, and exhibit any domain specificity, ii) characterize transport-dependent and -independent trafficking of sodium channel components to the node, including the roles of ankyrin G and NF186, respectively, and determine whether the sodium channel complex assembles locally, and i) investigate how nodal components are cleared from extranodal sites, focusing on NF186 to examine further the role of its cytoplasmic segment and that of endocytosis and proteolysis in clearing the internodal, surface pool of NF186. Relevance: These studies will elucidate the mechanisms that regulate the assembly of the node of Ranvier, which is critical to the ability of nerve fibers to conduct electrical impulses appropriately. Findings in this study may therefore have important implications for our understanding of the pathogenesis of disorders of myelinated fibers, including neuropathies that result in aberrant nerve conduction, and may thereby lead to new therapeutic strategies for these neurological disorders.