Generation of a Transgenic Mouse Model for Functional Study of a Novel Gene FG01

描述(申请人提供)：阿尔茨海默病(AD)是最常见的神经退行性疾病，影响10%的65岁以上的人和50%的85岁以上的人口。神经元外斑块由β-淀粉样蛋白(A？)多肽和由微管相关tau蛋白过度磷酸化的扭曲细丝组成的细胞内神经原纤维缠结(NFTs)是AD的两个主要病理特征。多条证据表明，A？大脑中的tau是AD发病的罪魁祸首，而过度磷酸化的tau在多种神经退行性疾病中被发现。因此，确定参与A？生成和tau过度磷酸化可能是疾病干预的关键。利用随机纯合子基因扰动(RHGP)技术，我们发现了一个新的基因fg01，并且发现FG01蛋白的过表达可以抑制A？生成和tau磷酸化。此外，我们还证明了其潜在的机制：即FG01可以与腺苷环化酶相互作用，促进cAMP的合成，上调PKA活性，使GSK-3磷酸化并失活，进而影响A？生成和tau磷酸化。我们现在正在培育转基因小鼠，这种小鼠在大脑中特异性地过度表达FG01。为了进一步研究FG01在体内的功能，我们提出了以下目标：(1)鉴定脑特异的FG01转基因小鼠；(2)将FG01转基因小鼠与3xTg-AD小鼠杂交，观察FG01对降低A？在体内，Tau的生成、tau的磷酸化、PKA和GSK-3的活性以及减轻其他AD样表型/异常的作用。这项工作将为新基因FG01参与AD致病事件的时空进展提供重要的洞察力，并可能阐明新的治疗靶点。公共卫生相关性：AD病理的两个主要标志是细胞外淀粉样斑块和神经元内神经原纤维缠结(NFTs)，其主要成分是A？和过度磷酸化的tau。A？相关基因的鉴定和鉴定生成和tau磷酸化是理解AD发病机制的关键。我们最近发现了一个新的基因fg01，它的过表达被发现在体外通过调节腺苷酸环化酶和PKA的活性来减少A2的产生、tau的磷酸化和GSK-3的活性。除了在AD的发病机制中起重要作用外，PKA和GSK-3在记忆/认知、糖尿病和肿瘤发生等多个病理/生理过程中都是至关重要的。因此，建立在脑内过表达FG01的转基因小鼠，并研究FG01在体内的生理功能，将为AD的病理过程提供重要的认识。此外，FG01转基因小鼠模型将有潜力开发多种疾病的新诊断和治疗方法，包括AD、糖尿病和癌症。

DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is the most common neurodegenerative disorder, affecting 10% of people over the age of 65 and 50% of the population over the age of 85. Extraneuronal plaques consisting of ?-amyloid (A?) peptides and intracellular neurofibrillary tangles (NFTs) composed of hyperphosphorylated twisted filaments of the microtubule-associated tau protein are two major pathological hallmarks of AD. Multiple lines of evidence suggest that overproduction/aggregation of A? in the brain is the prime culprit for AD pathogenesis, while hyperphosphorylated tau has been found in multiple neurodegenerative diseases. Therefore, identification of factors involved in modulation of A? generation and tau hyperphosphorylation may be crucial for disease intervention. Using the Random Homozygous Gene Perturbation (RHGP) technology, we have identified a novel gene, fg01, and found that overexpression of the FG01 protein can inhibit both A? generation and tau phosphorylation. Furthermore, we have demonstrated the underlying mechanism: i.e., FG01 can interact with adenylate cyclases to facilitate cAMP synthesis, upregulating PKA activity to phosphorylate and inactivate GSK-3, which in turn affects A? generation and tau phosphorylation. We are now generating transgenic mice that specifically overexpress FG01 in the brain. To further investigate the functions of FG01 in vivo, we propose the following aims: (1) to characterize brain-specific FG01 transgenic mice; and (2) to cross FG01 transgenic mice with 3XTg-AD mice and examine the effects of FG01 on reducing A? generation, tau phosphorylation and the activities of PKA and GSK-3 in vivo, as well as on alleviating other AD- like phenotypes/abnormalities. This work will provide major insight into the involvement of the novel gene FG01 in the temporal and spatial progression of AD pathogenic events and may potentially elucidate new therapeutic targets. PUBLIC HEALTH RELEVANCE: The two major hallmarks of AD pathologies are extracellular amyloid plaques and intraneuronal neurofibrillary tangles (NFTs), whose major components are A? and hyperphosphorylated tau, respectively. Identification and characterization of genes involved in A? generation and tau phosphorylation are critical for understanding AD pathogenesis. We have recently identified a novel gene, fg01, whose overexpression has been found to reduce A2 production, tau phosphorylation, and GSK-3 activity through modulating adenylate cyclase and PKA activity in vitro. In addition to being important to AD pathogenesis, both PKA and GSK-3 are critical for multiple patho/physiological processes such as memory/cognition, diabetes and tumorigenesis. Therefore, developing transgenic mice that overexpress FG01 in the brain and studying the physiological functions of FG01 in vivo will provide important insights into AD pathological processes. Moreover, the FG01 transgenic mouse model will have potential in developing novel diagnostic and therapeutic methods for multiple diseases including AD, diabetes and cancers.