Ubiquitin-mediated proteolysis at synaptic terminals

描述(申请人提供)：阐明控制突触小泡蛋白降解的调节机制是我们朝着了解突触功能如何在生理条件下调节以及突触终末如何在疾病中功能障碍和退化的目标迈出的关键一步。突触囊泡蛋白在神经递质释放中起关键作用，突触囊泡蛋白表达水平的变化有助于突触的可塑性，如学习和记忆。此外，突触小泡蛋白表达水平的改变与多种神经退行性疾病和包括药物成瘾在内的精神疾病有关。尽管突触小泡蛋白水平的调节在突触功能和功能障碍中很重要，但这种调节的分子机制仍然不清楚。另一方面，泛素-蛋白酶体蛋白分解途径已经成为细胞调节特定蛋白质表达水平的主要机制，泛素-蛋白酶体途径的异常与一些神经退行性疾病的发病机制有关。然而，人们对这一途径靶向的神经元蛋白底物知之甚少。申请者假设泛素-蛋白酶体途径通过特定的E3泛素-蛋白连接酶针对突触小泡蛋白进行降解。为了支持这一假说，我们的初步研究表明，SIAH-1和SIAH-2是一个哺乳动物Seven缺失的同源物家族，可能作为E3泛素蛋白连接酶来调节突触囊泡蛋白突触素的降解。在这个项目中，这一假设将得到彻底的检验。其具体目的是：1)表征Siah介导的突触素在神经末梢的泛素化和降解；2)研究Siah-1和Siah-2调节依赖泛素的突触素降解的分子机制；3)确定Siah蛋白的其他突触靶点；4)确定Siah蛋白在中枢神经系统中的分布和亚细胞定位；以及5)研究Siah介导的蛋白降解在神经细胞凋亡中的作用。这些研究将促进我们对突触终末蛋白质降解的理解，并为开发治疗神经系统功能障碍的治疗策略提供基础。

DESCRIPTION (provided by applicant): Elucidation of the regulation mechanisms that control the degradation of synaptic vesicle proteins represents a critical step towards our goal of understanding how synaptic function is regulated under physiologic conditions and how synaptic terminals become dysfunctional and degenerated in diseases. Synaptic vesicle proteins play a critical role in neurotransmitter release, and changes in the expression levels of synaptic vesicle proteins contribute to synaptic plasticity such as learning and memory. Furthermore, alterations in the expression levels of synaptic vesicle proteins are associated with a variety of neurodegenerative diseases and psychiatric disorders including drug addiction. Despite the importance of the regulation of synaptic vesicle protein levels in synaptic function and dysfunction, the molecular mechanisms underlying such regulation remain uncharacterized. On the other hand, the ubiquitin-proteasome proteolytic pathway has emerged as a major mechanism by which cells regulate the expression levels of specific proteins, and aberrations in the ubiquitin-proteasome pathway have been implicated in the pathogenesis of several neurodegenerative diseases. However, very little is known about neuronal protein substrates that are targeted by this pathway. The applicant hypothesizes that the ubiquitin-proteasome pathway targets synaptic vesicle proteins for degradation through specific E3 ubiquitin-protein ligases. In support of this hypothesis, our preliminary studies suggest that Siah-1 and Siah-2, a family of mammalian Seven in Absentia homologues, may act as E3 ubiquitin-protein ligases to regulate the degradation of synaptic vesicle protein synaptophysin. In this project, this hypothesis will be tested thoroughly. The specific aims are to: 1) characterize Siah-mediated ubiquitination and degradation of synaptophysin at nerve terminals; 2) investigate the molecular mechanisms by which Siah-1 and Siah-2 regulate the ubiquitin-dependent degradation of synaptophysin; 3) identify additional synaptic targets of Siah proteins; 4) determine the distribution and subcellular localization of Siah proteins in the central nervous system; and 5) examine the role of Siah-mediated protein degradation in neuronal apoptosis. These studies should advance our understanding of protein degradation at synaptic terminals, and provide a basis for the development of therapeutic strategies for treating malfunctions of the nervous system.