Role of proteoglycan-mediated trans-axonal signaling in pre-target topographic sorting

项目摘要 轴突投射到地形图中的精确组织对大脑功能至关重要，特别是 尤其是在感觉系统中。地形图形成的一个重要发展机制 是预先靶向轴突分类，即轴突在到达其靶点的途中根据其身份预先排序 和/或位置原点。例如，在视觉系统中，轴突分类沿着视束进行，在视束处， 视网膜的腹侧和腹侧轴突分别分离成束的腹侧和背侧分支， 到达他们的大脑目标虽然前靶轴突分类在地形图绘制中具有指导作用，但它如何 在发展过程中建立的机制仍然知之甚少。使用独特的透明度和可访问性 我们的研究表明，在斑马鱼胚胎中，沿着视束的沿着的地形秩序并不确定 而是通过选择性变性错配的背侧轴突来实现的 错误地沿着背分支走错了路硫酸乙酰肝素（HS），一种携带 通过硫酸乙酰肝素蛋白聚糖（HSPG），这种选择性变性需要非细胞自主。 然而，有几个问题仍然没有解决。HS如何指导错误分类的背侧轴突退化， 保留那些沿着神经束腹分支正确延伸的神经束？是否涉及特定的HSPG？ HS调节哪个分子通路？错误排序和正确定位之间的关键区别 背侧轴突与腹侧先驱轴突的距离。当错误地沿着背分支导航时， 错配的背侧轴突与已经伸长的腹侧轴突紧密接触。而且我们 初步数据显示，HSPG磷脂酰肌醇蛋白聚糖-3（Gpc 3）选择性地表达于成熟视网膜神经节细胞的腹侧RGCs， 视网膜。因此，我们假设Gpc 3介导的腹侧先锋和腹侧前叶之间的跨轴突信号传导， 错误分类的背侧跟随视网膜轴突触发了错误分类的背侧轴突的变性， 在视觉系统中的lish预目标地形排序。我们将通过描述 Gpc 3在腹侧视网膜神经节细胞和相应轴突中的表达和定位（Aim 1）， 以细胞特异性方式测试Gpc 3在视束分选中的功能（目的2）。gpc 3在pio的表面- neer轴突可以作为直接作用于错误分类的背侧轴突以触发它们的变性的引导线索， 或作为通过调节跨轴突信号传导途径间接控制分选的调节因子。获得 深入了解哪些信号传导因子可能受到Gpc 3的调节，我们将测试脑信号蛋白-神经纤毛蛋白- 已知丛蛋白途径调节其他系统中的轴突排序，也有助于视束排序， ing（目标3）。总之，这些研究将是第一个确定跨轴突信号之间的先驱 跟随轴突在视觉系统中建立了前目标地形分类，从而解决了一个主要的 我们对控制地形图形成的分子机制的理解存在差距。

Project summary The precise organization of axonal projections into topographic maps is crucial for brain function, espe- cially in sensory systems. An important developmental mechanism contributing to topographic map formation is pre-target axon sorting, whereby axons are pre-ordered en route to their target according to their identity and/or positional origin. In the visual system, for instance, axon sorting occurs along the optic tract, where dor- sal and ventral retinal axons segregate respectively into the ventral and dorsal branches of the tract before reaching their brain target. While pre-target axon sorting has an instructive role in topographic mapping, how it is established during development remains poorly understood. Using the unique transparency and accessibility of the zebrafish embryo, our studies have shown that topographic order along the optic tract is not established during initial axon guidance but instead achieved through the selective degeneration of missorted dorsal axons that have erroneously misrouted along the dorsal branch. Heparan Sulfate (HS), a glycosaminoglycan carried by Heparan Sulfate Proteoglycans (HSPGs), is required non-cell autonomously for this selective degeneration. Yet, several questions remain unsolved. How does HS instruct missorted dorsal axons to degenerate while preserving those correctly elongating along the ventral branch of the tract? Is there a specific HSPG involved? Which molecular pathway(s) does HS regulate? A key difference between missorted and correctly targeted dorsal axons is their proximity to ventral pioneer axons. While erroneously navigating along the dorsal branch, missorted dorsal axons appear in close contact with ventral axons that have already elongated. Moreover, our preliminary data show that the HSPG glypican-3 (Gpc3) is selectively expressed in ventral RGCs in the mature retina. Thus, we hypothesize that Gpc3-mediated trans-axonal signaling between ventral pioneer and missorted dorsal follower retinal axons triggers the degeneration of missorted dorsal axons to estab- lish pre-target topographic sorting in the visual system. We will test that hypothesis by characterizing the expression and localization of Gpc3 in ventral retinal ganglion cells and corresponding axons (Aim 1), and by testing the function of Gpc3 in optic tract sorting in a cell specific manner (Aim 2). Gpc3 at the surface of pio- neer axons may act as a guidance cue acting directly on missorted dorsal axons to trigger their degeneration, or as a modulating factor controlling sorting indirectly by regulating a trans-axonal signaling pathway. To gain insight into which signaling factors might be regulated by Gpc3, we will test whether the semaphorin-neuropilin- plexin pathway, which is known to regulate axon ordering in other systems, also contributes to optic tract sort- ing (Aim 3). Altogether, these studies will be the first to determine how trans-axonal signaling between pioneer and follower axons establishes pre-target topographic sorting in the visual system, thus addressing a major gap in our understanding of the molecular mechanisms controlling topographic map formation.