Roles of Phosphoinositides at the Synapse

描述（申请人提供）：磷酸肌醇，如PI（4，5）P2（PIP 2），调节肌动蛋白动力学和囊泡运输的各个方面。然而，PIP 2对囊泡运输、其定位和通过合成的调节的确切贡献还不清楚。突触为探索这些问题提供了一个很好的模型。因此，本项目的目标是探索PIP 2在突触囊泡运输和肌动蛋白动力学中的突触前作用。为此，我将研究改变它们在活突触中的可用性的后果。通过向七鳃鳗巨网状脊髓突触中注射特异性PIP 2结合蛋白模块以减少内源性PIP 2或通过注射干扰PIP 2代谢酶的试剂来改变PIP 2的可用性。荧光成像，电生理学和电子显微镜的组合将被用来检查这些试剂对肌动蛋白动力学和突触囊泡贩运在体内的后果。这种方法将首次全面分析PIP 2在突触处的功能。De Camilli实验室已经确定并表征了在突触处代谢PIP 2的酶。这些酶之一的遗传和急性损伤，PIP 2-磷酸酶synaptojanin，改变肌动蛋白细胞骨架和破坏突触中的突触囊泡再循环，表明PIP 2在这些过程中的重要作用。改变的磷脂酰肌醇代谢与阿尔茨海默病的发病机制以及多种人类高血压、葡萄糖代谢和脂质代谢紊乱有关。因此，我的研究可能对人类健康产生广泛的影响。

DESCRIPTION (provided by applicant): Phosphoinositides, such as PI(4,5)P 2 (PIP2), regulate various aspects of actin dynamics and vesicle trafficking. However, the precise contributions of PIP2 to vesicle trafficking, its localization, and regulation by synthesis is not well-understood. The synapse offers a good model in which to explore these questions. Therefore, the goal of this project is to explore the presynaptic roles of the PIP2 in synaptic vesicle trafficking and actin dynamics. To do so, I will examine the consequences of altering their availability in living synapses. PIP2 availability will be altered by injecting into lamprey giant reticulospinal synapses either specific, PIP2 binding protein modules to decrease endogenous PIP2 or by injecting reagents that perturb PIP2-metabolizing enzymes. A combination of fluorescence imaging, electrophysiology, and electron microscopy will be used to examine the consequences of these reagents on actin dynamics and synaptic vesicle trafficking in vivo. This approach will provide the first comprehensive analysis of PIP2 functions at the synapse. The De Camilli lab has already identified and characterized enzymes that metabolize PIP2 at the synapse. Genetic and acute impairment of one of these enzymes, the PIP2-phosphatase synaptojanin, alters actin cytoskeleton and disrupts synaptic vesicle recycling in synapses, suggesting an important role for PIP2 in these processes. Altered phosphoinositide metabolism has been implicated in the pathogenesis of Alzheimer's disease, as well as a variety of human disorders of hypertension, glucose metabolism, and lipid metabolism. Thus, my studies may have wide implications for human health.