Mechanisms of Axonal Pathfinding Controlled by Netrin-1

项目概述：作为一名独立的科学家，我的职业目标是了解轴突伸长和寻路的细胞和分子过程，并确定这些过程在神经发育和精神健康障碍中的作用。在康奈尔大学威尔医学院Samie R. Jaffrey博士的实验室进行博士后培训期间，我研究了轴突内mRNA翻译和蛋白质降解的作用。这个训练补充了我在蛋白质生物化学和突触可塑性方面的博士训练。我计划将原代神经元的体外研究与小鼠遗传学相结合，研究控制轴突寻径的信号通路。本研究的目的是研究局部蛋白翻译和降解在netrin-1信号传导中的作用。我发现PAR复合体是netrin-1信号传导的关键效应体，编码PAR3的转录本（PAR复合体的中心成分）定位于轴突，并根据netrin-1进行翻译。根据我的初步数据，我提出了三个目标。首先，我将描述PAR复合物效应物，泛素e3连接酶Smurfl在netrin-1介导的轴突伸长和生长锥转动中的作用。其次，我将使用PARS翻译的荧光报告和轴突特异性RNA干扰来解决局部PAR3翻译在netrin-1信号传导中的作用。对于这些研究，我将开发新的技术来研究生长锥转向，以补充现有的轴突生长研究。第三，本应用描述了PAR3转录物在轴突上有条件定位的转基因小鼠的设计、生成和分析。这些小鼠将允许在轴突寻径中建立轴突PARS翻译的生理作用。这项研究将极大地促进我们对netrin-1信号的理解，并将为神经科学的细胞、分子和遗传方法提供培训。相关性：在发育过程中，未成熟大脑中的引导分子引导轴突经过漫长而复杂的通路到达目标区域；这一过程中的紊乱会导致神经发育障碍，如精神分裂症。这个应用程序试图解释轴突内部的机制，导致轴突向或远离引导分子生长。我希望我的研究将具有广泛的相关性，并可能确定治疗或预防神经发育障碍的可能目标。

DESCRIPTION (provided by applicant): Project Summary: My career goal as an independent scientist is to understand the cellular and molecular processes involved in axon elongation and pathfinding and to identify the role of these processes in neurodevelopmental and mental health disorders. During my postdoctoral training in the laboratory of Dr. Samie R. Jaffrey at Weill Medical College of Cornell University, I have studied the role of intra-axonal mRNA translation and protein degradation. This training complements my doctoral training in protein biochemistry and synaptic plasticity. I plan to combine in vitro studies in primary neurons with mouse genetics to study the signaling pathways governing axonal pathfinding. The objective of this research is to examine to role of local protein translation and degradation in netrin-1 signaling. I have found that the PAR complex is a critical effector of netrin-1 signaling, and that the transcript encoding PAR3, a central component of the PAR complex, is localized to axons and translated in response to netrin-1. Based on my preliminary data, I propose three aims. First, I will characterize the role of a PAR complex effector, the ubiquitin E3-ligase Smurfl, in netrin-1-mediated axon elongation and growth cone turning. Second, I will use a fluorescent reporter of PARS translation and axon-specific RNA interference to address the role of local PAR3 translation in netrin-1 signaling. For these studies, I will develop new techniques to study growth cone turning to complement existing ones for the study of axon growth. Third, this application describes the design, generation, and analysis of transgenic mice that exhibit conditional localization of the PAR3 transcript to axons. These mice will allow the physiological role of axonal PARS translation to be established in axonal pathfinding. This research will significantly advance our understanding of netrin-1 signaling, and it will provide training in cellular, molecular, and genetic approaches to neuroscience. Relevance: During development, guidance molecules in the immature brain direct axons to their target areas, following long and complex pathways; disturbances in this process cause neurodevelopmental disorders, such as schizophrenia. This application seeks to explain the mechanisms inside an axon causing it to grow towards or away from guidance molecules. I expect that my studies will have broad relevance and might identify possible targets for the treatment or prevention of neurodevelopmental disorders.