Molecular Organization and Function of Paranodal Axo-Glial Junctions

有髓轴突的标志是它们组织成分子上不同的结构域，这是神经元分化的先决条件。 动作电位的快速传播。结旁区建立轴胶质分隔连接 通过涉及轴突接触蛋白相关蛋白（Caspr）、接触蛋白（Cont）和 神经胶质神经成束蛋白NF 155。结域由神经元NF 186、电压门控钠（Nav） 通道和锚蛋白G（AnkG），一种细胞骨架衔接蛋白。我们发现Caspr和NF 155的缺失 导致AGSJs的丢失，结旁蛋白的错误定位，结旁组织的紊乱， 轴突细胞骨架和有髓轴突变性，但相对保留了节点组织 不受影响最近，我们发现带4.1B，一个结旁/结旁细胞骨架蛋白，是必需的， 稳定AGSJs和AGPARANODAL组织。我们在这里展示了另一种细胞骨架蛋白Whirlin， 蛋白质是结旁致密化和细胞骨架稳定性所必需的。在进一步的研究中，我们证明了 节点NF 186的缺失废除了节点处的Nav通道和AnkG的聚集， 结旁AGSJ侵入结间隙。最重要的是，我们在这里表明，在体内损失的AnkG不 不影响结的形成，但可能影响结的稳定性。虽然已经取得了重大进展， 关于轴突结构域的组成和组织，仍然存在一些基本问题 关于这些结构域的跨膜成分如何与局部细胞骨架相互作用， 域组织，并确保轴突结构的长期稳定性和维护。基于我们 发表的和初步的研究，我们的中心假设是，跨膜组件和 局部轴突细胞骨架对于轴突结构域组织、其稳定性和功能至关重要。我们 将使用遗传，分子和细胞生物学方法来确定副阳极，节点和 其相关的细胞骨架蛋白在结构域稳定和维护，以及恢复的功效 通过在进行性衰弱的成年小鼠突变体中重新表达NF 155和NF 186，我们 我们将在以下具体目标中实现我们的目标：（1）细胞骨架丢失的后果是什么？ 支架蛋白和NF 155在成年期的损失对结旁AGSJS的维持和功能的影响？ (2)在成年期，淋巴结细胞骨架蛋白和NF 186的丢失对淋巴结的生长有什么影响？ 有髓轴突中节点的稳定性和功能？和（3）是具有延长的周期的成年有髓轴突 能够重组轴突区域以恢复神经传导？ 总的来说，我们的研究将提供直接影响轴突域 形成和维持，以及如何重组这些结构和恢复神经功能。在 未来，这些研究将促进我们对脱髓鞘疾病，如多发性硬化症， (MS)导致轴突结构域紊乱并指导治疗干预的发展。

The hallmark of myelinated axons is their organization into molecularly distinct domains, a pre-requisite for the rapid propagation of action potentials. The paranodal domains establish the axo-glial septate junctions (AGSJs) through interactions involving axonal Contactin-associated protein (Caspr), Contactin (Cont), and glial Neurofascin NF155. The nodal domain is organized by neuronal NF186, voltage gated sodium (Nav) channels, and Ankyrin G (AnkG), a cytoskeletal adaptor protein. We showed that loss of Caspr and NF155 results in loss of AGSJs, mislocalization of the juxtaparanodal proteins, disorganization of the paranodal axonal cytoskeleton, and degeneration of myelinated axons, but nodal organization remains relatively unaffected. Recently, we showed that Band 4.1B, a paranodal/ juxtaparanodal cytoskeletal protein, is essential for the stability of AGSJs and juxtaparanodal organization. We show here that Whirlin, another cytoskeletal protein, is required for paranodal compaction and cytoskeletal stability. In further studies, we demonstrated that loss of nodal NF186 abolished clustering of Nav channels and AnkG at the nodes, allowing the flanking paranodal AGSJs to invade the nodal space. Most importantly, we show here that in vivo loss of AnkG does not abolish node formation, but may affect nodal stability. While significant advancements have been made regarding the composition and organization of axonal domains, there still remain fundamental questions regarding how the transmembrane components at these domains interact with local cytoskeleton to initiate domain organization, and to ensure long-term stability and maintenance of axonal architecture. Based on our published and preliminary studies, our central hypothesis is that the transmembrane components and local axonal cytoskeleton are critical for axonal domain organization, their stability and function. We will use genetic, molecular and cell biological methods to determine the specific role of paranodes, nodes and their associated cytoskeletal proteins in domain stabilization and maintenance, and the efficacy of restoration of key axonal domains by re-expression of NF155 and NF186 in progressively weak adult mouse mutants. We will accomplish our goals in the following specific aims: (1) What are the consequences of loss of cytoskeletal scaffolding proteins and loss of NF155 during adult life on the maintenance and function of paranodal AGSJs? (2) What are the consequences of loss of nodal cytoskeletal proteins and loss of NF186 during adult life on the stability and function of nodes in myelinated axons? and (3) Are adult myelinated axons with extended periods of disorganized domain structure able to re-organize axonal domains to restore nerve conduction? Collectively, our studies will provide insights that bear directly on the mechanisms by which axonal domains are formed and maintained, and how these structures can be reorganized and nerve function restored. In the future, these studies will advance our understanding of how demyelinating diseases, such as multiple sclerosis (MS) lead to axonal domain disorganization and guide the development of therapeutic interventions.