Functions of Heparan Sulfate Proteoglycans in Axon Guidance and Degeneration.

项目总结/摘要 神经元连接的精确组织对于处理信息至关重要。的一大挑战 神经科学的目的是了解这些连接是如何正确建立的，以及这种连接中的错误是如何发生的。 这一过程可能导致精神疾病，如自闭症或精神分裂症。在发育过程中， 轴突被吸引和排斥的线索引导沿着确定的路径到达它们的大脑目标。在 除了这个指导过程，涉及修剪或变性的机制纠正了轴突， 偏离正确的路径，从而确保电路形成的准确性。有几个因素调节轴突 它们提供的组合信息不足以塑造整个神经网络。 硫酸乙酰肝素蛋白聚糖（HSPGs）是一种与乙酰肝素结合的细胞表面和细胞外核心蛋白 硫酸化（HS）糖链，在轴突寻路中起着至关重要的作用，并且是触发神经元凋亡所必需的。 被误导的轴突的选择性退化。然而，HSPG如何控制这些不同的过程仍然是一个问题。 谜HS链经过许多修饰，特别是硫酸化，赋予它们独特的多样性 和结构复杂性。这些修饰被提出来产生一个复杂的“糖密码” 通过调节大脑布线所必需的大多数因素来协调轴突连接的形成。而 吸引人的是，这种“糖密码”假说仍然是理论上的，还没有在脊椎动物中得到验证。而且 携带这些特定HS模式的核心蛋白的贡献是未知的。利用斑马鱼 视网膜顶盖系统作为体内模型，我发现三种不同的HSPG（SDC 2，GPC 1a和 GPC 1b）调节视网膜轴突导向的不同步骤。在这个项目中，我建议剖析这些 核心蛋白和HS链修饰使用体内的独特组合调节轴突导航， 方法，遗传学和HS生物化学。本提案的第一个目的是确认 SDC 2，GPC 1a和GPC 1b，并在细胞和分子水平上研究其作用方式。它将 确定它们的缺失如何影响轴突行为并导致指导错误，它们在哪里起作用， HS链是其功能所必需的，它们与哪些因素相互作用。第二个目的是 该项目将确定特定HS结构基序对轴突寻路和变性的贡献。它 将测试6-O和3-O磺基转移酶的作用，这些酶在特定的条件下用硫酸化修饰HS链。 位置，并研究具有定义的硫酸化基序和大小的合成HS寡糖如何调节 视网膜轴突导航，无论是在体外培养系统和在体内发育胚胎。通过研究 核心蛋白和HS精细结构的作用，拟议的研究提供了一个独特的机会来测试 糖代码是否在发育过程中协调大脑线路。它也可能提供新的研究方向 为未来发展的治疗和再生战略的背景下，神经系统疾病。

Project Summary/Abstract Precise organization of neuronal connections is crucial for processing information. A major challenge in neuroscience is to understand how these connections are properly established, and how errors in this wiring process can lead to psychiatric disorders like autism or schizophrenia. During development, neurons extend axons that are guided along defined paths by attractive and repulsive cues to reach their brain target. In addition to this guidance process, mechanisms involving pruning or degeneration correct axons that have deviated from the right path, thereby ensuring accuracy of circuit formation. Several factors regulate axon guidance, but the combined information they provide is not sufficient to sculpt the entire neuronal network. Heparan sulfate proteoglycans (HSPGs) are cell-surface and extracellular core proteins with attached heparan sulfate (HS) sugar chains that play crucial roles in axon pathfinding, and are essential for triggering the selective degeneration of misguided axons. How HSPGs control these different processes remains however a mystery. HS chains undergo many modifications, especially sulfations, that confer on them a unique diversity and structural complexity. These modifications have been proposed to generate a complex “sugar code” orchestrating the formation of axonal connections by regulating most factors essential for brain wiring. While appealing, this “sugar code” hypothesis is still theoretical and has not been tested in vertebrates. Moreover, the contribution of the core proteins carrying these specific HS patterns is not known. Using the zebrafish retinotectal system as an in vivo model, I have discovered that three distinct HSPGs (SDC2, GPC1a and GPC1b) regulate different steps of retinal axon guidance. In this project, I propose to dissect out how these core proteins and HS chain modifications regulate axon navigation using a unique combination of in vivo approaches, genetics and HS biochemistry. The first aim of this proposal will confirm the novel functions of SDC2, GPC1a and GPC1b, and investigate their mode of action at a cellular and molecular level. It will determine how their absence affects axon behavior and leads to guidance errors, where they act, whether their HS chains are required for their function, and what are the factors they interact with. The second aim of this project will determine the contribution of specific HS structural motifs for axon pathfinding and degeneration. It will test the roles of the 6-O and 3-O sulfotransferases that modify HS chains with sulfations at specific positions, and examine how synthetic HS oligosaccharides with defined sulfation motifs and sizes regulate retinal axon navigation, both in vitro in culture systems and in vivo in the developing embryo. By studying the roles of both core proteins and HS fine structure, the proposed studies give a unique opportunity to test whether a sugar code orchestrates brain wiring during development. It might also provide new lines of research for future development of therapeutic and regenerative strategies in the context of neurological disorders.