Regulation of axon outgrowth by retrograde Ret signaling

在发育过程中或损伤后，轴突终末必须通过复杂的环境才能形成 功能连接。RET是一种神经营养素受体，存在于生长轴突的顶端。雷特 激活和内化诱导激活的受体逆行运输到细胞体，触发 转录反应，进而促进轴突生长。尽管它在紧张的时候扮演着重要的角色 系统开发，目前尚不清楚激活的Ret受体是如何在轴突中运输的，其性质 逆行Ret信号诱导的转录反应，以及这些转录靶点是如何 最终促进轴突生长。为了解决这些问题，我们正在利用斑马鱼的独特优势， 包括活体成像和遗传方法，以确定支配逆行的分子机制 神经营养素受体Ret的运输和随后的转录反应。在我们的预赛中 研究发现，Ret的逆行转运依赖于与支架蛋白JIP3的结合，JIP3 将货物连接到逆行马达上进行运输。Ret和JIP3突变体都显示了截短的感觉轴突， 活体成像显示生长锥体动力学异常，轴突终末细化减少。使用RNA 测序，我们确定了一些转录诱导的因子，以响应Ret-JIP3 在这种情况下，逆行信号和肌动蛋白为基础的突触行为的假定调节者。基于 在这些数据中，我们假设依赖JIP3的Ret逆行运输诱导了促进生长的因素 感觉轴突延伸所需的锥体动力学。我们将在三个具体目标中检验这一假设。在……里面 目的1，我们将确定Ret逆行转运的分子机制以及JIP3在该过程中的作用。 第二个目标中的实验将确定导致轴突失败的生长锥动力学缺陷。 在ret和JIP3突变体中延伸，并确定Ret逆行信号在这一过程中的作用。这个 最后一个目的是研究逆行Ret信号引起的转录反应及其影响因素 受此转录程序调控的基因可促进生长锥动态。总而言之，我们的研究 将斑马鱼的创新检测与体内技术相结合，确定逆行的具体作用 神经营养因子信号在轴突生长中的作用。我们的工作将进一步揭示 神经营养因子信号被转化为调节生长锥动态和轴突的细胞反应 分机。

During development or following injury, axon terminals must navigate through a complex environment to form functional connections. Ret is a neurotrophin receptor which is present at the tips of growing axons. Ret activation and internalization induces retrograde transport of the activated receptor to the cell body triggering a transcriptional response, which in turn promotes axon outgrowth. Despite its essential role during nervous system development, it is not known how activated Ret receptor is trafficked in axons, the nature of the transcriptional response induced by the retrograde Ret signaling, and how these transcriptional targets ultimately promote axon growth. To address these questions, we are using the unique advantages of zebrafish, including live imaging and genetic approaches, to identify the molecular mechanisms that govern retrograde transport of a neurotrophin receptor Ret and the subsequent transcriptional response. In our preliminary studies, we discovered that retrograde transport of Ret depends on binding to the scaffold protein, Jip3, which links cargo to the retrograde motor for transport. Both ret and jip3 mutants display truncated sensory axons, and live imaging revealed abnormal growth cone dynamics and reduced axon terminal elaboration. Using RNA sequencing, we identified a number of factors that are transcriptionally induced in response to Ret-Jip3 retrograde signaling and are putative regulators of actin-based protrusive behavior in this context. Based on this data, we hypothesize that Jip3-dependent retrograde transport of Ret induces factors that promote growth cone dynamics required for sensory axon extension. We will test this hypothesis in the three specific aims. In Aim 1, we will define the molecular mechanisms of Ret retrograde transport and the role of Jip3 in this process. Experiments in the second aim will define the defects in growth cone dynamics that lead to the failure of axon extension in ret and jip3 mutants and determine the role of the Ret retrograde signaling in this process. The last aim will investigate the transcriptional response elicited by the retrograde Ret signaling and how factors that are regulated by this transcriptional program promote growth cone dynamics. Altogether, our study combines innovative assays in zebrafish with in vivo techniques to determine the specific role of retrograde neurotrophin signaling in axon outgrowth. Our work will further uncover the mechanisms by which long-range neurotrophin signals are transduced into cellular responses that regulate growth cone dynamics and axon extension.