Uncovering the mechanisms of Ephexin5 function in dendritic spine plasticity and Alzheimer's disease

摘要/项目总结 阿尔茨海默氏病（AD）与树突棘、显微树突棘和树突状细胞的进行性丧失有关。 作为突触交流的场所，对记忆的储存至关重要。水平 RhoGEF Ephexin 5是树突棘密度的调节因子，它似乎是促进AD诱导的细胞凋亡的关键。 树突棘缺失。关于Ephexin 5的底物特异性和Ephexin 5活性的调节 仍然未知。该项目的目标是确定Ephexin 5信号传导的机制， 发生在正常生理和疾病中。我的假设是E5经历了磷酸化- 底物特异性从RhoA到Rac 1/Cdc 42的依赖性转变，与其在调节脊柱可塑性中的作用有关 并对AD的生理起作用。使用双光子荧光寿命成像与基因 编码FRET生物传感器，我将光学探测活性的GTP酶在活组织。我将定义基板 在正常生理环境下和AD诱导的细胞损伤模型中，神经元E5的特异性。 使用质谱，我将确定Ephexin 5的磷酸化是否在神经元信号传导过程中发生变化， 在AD样细胞状态下，使用磷酸突变分析，我将研究磷酸化如何影响E5 底物特异性我的研究结果将进一步了解学习和记忆的分子机制 并有可能确定新的治疗AD的目标。 这个项目的培训将在加州大学戴维斯分校进行，我的导师和赞助商Karen Zito，一个 突触实时成像专家，共同赞助人James Trimmer，神经元蛋白质结构专家 和功能我将获得先进的活细胞成像和质谱分析的关键技术技能，这将 帮助完成这个研究项目。此外，我将磨练指导，管理， 沟通和写作。

ABSTRACT / PROJECT SUMMARY Alzheimer’s Disease (AD) is associated with a progressive loss of dendritic spines, microscopic dendritic structures that serve as the sites of synaptic communication and are essential for the storage of memory. Levels of the RhoGEF Ephexin5, a regulator of dendritic spine density, appear to be key in facilitating AD-induced dendritic spine loss. Much about the substrate specificity of Ephexin5 and the regulation of Ephexin5 activity remain unknown. The goal of this project is to determine the mechanisms by which Ephexin5 signaling occurs in normal physiology and disease. It is my hypothesis that E5 undergoes a phosphorylation- dependent shift in substrate specificity from RhoA to Rac1/Cdc42, relevant to its role in regulating spine plasticity and contributing to the physiology of AD. Using 2-photon fluorescence lifetime imaging coupled with genetically encoded FRET biosensors, I will optically probe the activity of GTPases in live tissue. I will define the substrate specificity of neuronal E5 under normal physiological contexts, and in a model of AD-induced cellular damage. Using mass spectrometry, I will determine if phosphorylation of Ephexin5 changes during neuronal signaling or in an AD-like cellular state, and using phosphomutant analysis I will examine how phosphorylation influences E5 substrate specificity. My results will further the knowledge of the molecular mechanisms of learning and memory and have the potential to identify novel therapeutic targets for AD. Training for this project will take place at UC Davis, with the support of my mentor and Sponsor, Karen Zito, an expert in the live-imaging of synapses, and Co-sponsor James Trimmer, and expert in neuronal protein structure and function. I will acquire key technical skills in advanced live-cell imaging and mass spectrometry, which will aid in the completion of this research project. Additionally I will hone skills of mentorship, management, communication, and writing.