Neurotrophic factor trafficking and signaling in development and disease

神经细胞生物学中的一个基本问题是膜蛋白是如何 在细胞体中生物合成后，长距离运输到轴突。轴突靶向 膜蛋白对神经元连接的形成和维持至关重要 对于功能正常的神经系统来说。然而，大多数膜蛋白是如何输送到 轴突仍未定义。 神经生物学中长期持有的观点是，信号受体是结构性的 通过分泌性交易传递给轴突。相反，我们发现TrkA神经营养因子 受体是神经元存活、轴突生长和炎症的重要调节因素 疼痛通过跨细胞作用主动招募到轴突，这是一种基于细胞内吞的机制 其中嵌入胞体表面的受体是内在化和顺序化的 被输送到轴突。引人注目的是，顺行TrkA转胞作用是由配体触发的， 神经生长因子(NGF)作用于轴突终末，提示正反馈 一种在时间段内动态放大轴突中受体可用性的机制 需要的时候。此外，我们还发现TrkA转胞作用是由以下活性启动的 PTP1B是细胞体中的一种内质网驻留蛋白酪氨酸磷酸酶。的总目标是 这一应用是为了阐明潜在的信号和交易机制 配基触发的受体靶向轴突的模式特征不佳。在目标1中， 我们将定义NGF介导的在细胞体中启动跨细胞作用的机制， 阐明受体转运的转运路线和转运动力学，以及 研究TrkA在体内的细胞转运。在目标2中，我们将检验ER- 锚定PTP1B磷酸酶通过调控TrkA促进TrkA生物学功能的获得 受体的长距离跨细胞作用。我们将采用实时成像、生化和 分室培养结合体内培养的神经元功能分析 对转基因小鼠的分析以实现这些目标。这些研究将 解决一个基本的，但研究很少的细胞生物学问题，即信号如何 受体被定向到轴突，并将提供对特定机制的洞察 增强神经元对空间作用的外部线索的反应性。

A fundamental question in neuronal cell biology is how membrane proteins are transported long-distance to axons after biosynthesis in cell bodies. Axon targeting of membrane proteins is critical for the formation and maintenance of neuronal connections and for a functional nervous system. Yet, how most membrane proteins are delivered to axons remains undefined. A long-held view in neurobiology is that signaling receptors are constitutively delivered to axons via secretory trafficking. In contrast, we found that TrkA neurotrophin receptors that are essential regulators of neuron survival, axon growth, and inflammatory pain are actively recruited to axons via transcytosis, an endocytosis-based mechanism where receptors embedded in soma surfaces are internalized and anterogradely transported to axons. Strikingly, anterograde TrkA transcytosis is triggered by the ligand, Nerve Growth Factor (NGF), acting on axon terminals, suggesting a positive feedback mechanism that serves to dynamically scale up receptor availability in axons during times of need. Furthermore, we identified that TrkA transcytosis is primed by the activity of PTP1B, an ER-resident protein tyrosine phosphatase, in cell bodies. The overall goal of this application is to elucidate the signaling and trafficking mechanisms underlying a poorly characterized mode of ligand-triggered targeting of receptors to axons. In Aim 1, we will define NGF-mediated mechanisms that initiate transcytosis in cell bodies, elucidate the trafficking itinerary and transport kinetics of receptor transcytosis, and investigate TrkA transcytosis in vivo. In Aim 2, we will test the hypothesis that ER- anchored PTP1B phosphatase promotes a gain of TrkA biological function by controlling the long-distance transcytosis of receptors. We will employ live imaging, biochemical, and functional analyses in compartmentalized neuron cultures in combination with in vivo analyses of genetically modified mice to accomplish these goals. These studies will address a fundamental, yet poorly studied, cell biological question of how signaling receptors are directed to axons, and will provide insight into specialized mechanisms that enhance neuronal responsiveness to spatially acting extrinsic cues.