Novel E3 Ligases in Nerve Degeneration and Regeneration

描述（由申请人提供）：在神经退行性疾病中，轴突变性通常先于神经元细胞体死亡，因此抑制轴突变性代表了对抗这些疾病的潜在新干预点。轴突变性现在被认为是一个活跃的、自我毁灭的程序，其中泛素蛋白体途径起着重要作用。我们已经确定了一种新的E3泛素连接酶，ZNRF1在神经损伤后的雪旺细胞中被诱导。轴突损伤后，ZNRF1也在神经元中特异性上调，并与泛素一起上调。ZNRF1和另一种相关的E3连接酶ZNRF2在发育和成年期间都在整个神经系统中高度表达。通过体外实验，我们发现ZNRF蛋白活性是正常轴突变性所必需的。在本提案中，我们概述了实验来定义ZNRF蛋白如何影响轴突变性。首先，我们将分析znrf缺陷小鼠的表型，以确定这些蛋白质的生理作用。特别是，我们将重点分析机械性和代谢性损伤后的神经变性/再生过程以及突触发生。我们还旨在鉴定znrf介导的泛素化靶向蛋白。将进行蛋白质组学分析，从缺乏ZNRF蛋白的小鼠脑组织或从含有显性阴性ZNRF突变体和ZNRF靶点的亲和纯化复合物中鉴定ZNRF靶点。这些研究将提高我们对轴突病和神经元变性的认识，并为开发神经退行性疾病的新疗法提供基础

DESCRIPTION (provided by applicant): Axonal degeneration often precedes the death of neuronal cell bodies in neurodegenerative diseases, thus inhibiting axonal degeneration represents a potential novel point of intervention in combating these diseases. Axonal degeneration is now regarded as an active, self-destructive program in which the ubiquitin proteosome pathway plays an important role. We have identified a novel E3 ubiquitin ligase, ZNRF1 that is induced in Schwann cells after nerve injury. ZNRF1 is also specifically upregulated in neurons, along with ubiquitin, after axonal damage. ZNRF1 and another related E3 ligase, ZNRF2, are both highly expressed throughout the nervous system during development and in adulthood. Using in vitro assays we have found that ZNRF protein activity is required for normal axonal degeneration. In this proposal, we have outlined experiments to define how ZNRF proteins influence axonal degeneration. First, we will analyze the phenotypes of ZNRF-deficient mice in order to determine physiological roles of these proteins. In particular, the will focus our analysis on the processes of nerve degeneration/regeneration after mechanical and metabolic injury as well as synaptogenesis. We also aim to identify proteins targeted by ZNRF-mediated ubiquitination. A proteomic analysis will be performed to identify ZNRF targets either from brain tissues of mice deficient for ZNRF proteins or from affinity-purified complex containing dominant negative ZNRF mutants and ZNRF targets. These studies will improve our understanding of axonopathy and neuronal degeneration, and provide a basis for developing novel treatments for neurodegenerative diseases