Characterization of a caspase-cleavage resistant tau knock-in mouse

 描述(申请人提供)：阿尔茨海默病(AD)是世界上最常见的老年依赖痴呆的原因，与大脑淀粉样斑块和神经原纤维缠结(NFT)有关。淀粉样斑块主要由A？多肽组成，而NFTs则由细胞骨架蛋白tau过度磷酸化形式的异常聚集体组成。A？多肽是由淀粉样前体蛋白(APP)的双裂解产生的。BACE1裂解产生C-末端片段-Ctf，该片段被-分泌酶加工成几种A？亚型。遗传数据表明，APP处理的调节与AD有关。目前治疗阿尔茨海默病的药物发现方法主要集中在防止Aü形成或消除现有的淀粉样蛋白沉积上。以淀粉样蛋白为靶点的化合物/生物制品在临床试验中一再失败，既可以归因于纳入的AD患者，他们的疾病进展太晚，无法从治疗干预中受益，也可以归因于A？不是AD病理生理的主要原因。AD最先进的替代假说是tau毒性假说。与这一假设一致，减少内源性tau的表达可以防止表达带有家族性AD突变的人APP的转基因小鼠的行为缺陷，而不会改变Aü水平。我们的初步数据表明，tau还介导了家族性丹麦痴呆(FDDKI小鼠)的行为缺陷，FDDKI小鼠是一种与APP处理BRI2/ITM2B调节因子突变相关的AD样痴呆，也以tau病为特征。一些证据表明，半胱氨酸天冬氨酸氨基转移酶在AD进展的早期被激活，并可能在神经元丢失和NFT病理中发挥作用。Tau在D421(Tau)被刽子手caspase切割。在caspase裂解之后，tau促进了依赖于核的细丝的形成，并采用了被早期病理tau标记mc1识别的构象变化。NFTTau可被抗体PHF-1磷酸化并识别。在AD大脑中，Tau与NFTs的标志物相关，并与认知能力下降相关。此外，-Tau可引发NFT的形成，并可产生毒性效应。这些发现导致了一种假设，即Tau是神经退行性变的关键毒性部分。为了直接验证这一理论，我们产生了敲入小鼠，在这些小鼠中，编码D421的第12外显子中的内源tau密码子GAC突变为AAC，现在AAC编码天冬酰胺(N)。这些被称为TauD421N的敲入小鼠表达一种tau突变体taucas/res，这种突变体不能被caspase切割，因此不能产生tau。TauD421N小鼠将被用来确定在AD和FDD这两种神经退行性疾病的动物模型中，Tau是否介导突触可塑性和记忆缺陷。我们的研究将探讨调节Tau的形成是否是治疗AD和其他由神经毒性Tau形式介导的神经退行性疾病的潜在策略。

 DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is the most common cause of ageing-dependent dementia in the world and is associated with cerebral amyloid plaques and neurofibrillary tangles (NFTs). Amyloid plaques are mostly composed of Aß peptides while NFTs are composed of abnormal aggregates of hyperphosphorylated forms of the cytoskeletal protein tau. Aß peptides are produced by a double cleavage of the amyloid precursor protein (APP). BACE1 cleavage produces the C-terminal fragment, ß-CTF, which is then processed into several Aß isoforms by -secretase. Genetic data suggest that regulation of APP processing contributes to AD. Current drug discovery approaches in AD have focused on preventing Aß formation or removing existing amyloid deposits. The repeated failures of compounds/biologics targeting amyloid in clinical trials can either be attributed to the inclusion of AD patients who were too advanced in their disease progression to benefit from therapeutic intervention, or to the fact that Aß is not the main cause of AD patho-physiology. The most advanced alternative hypothesis of AD is that of tau toxicity. Consistent with this hypothesis, reducing endogenous tau expression prevents behavioral deficits in transgenic mice expressing human APP with familial AD mutations, without altering Aß levels. Our preliminary data indicate that tau also mediates behavioral deficits in Familial Danish Dementia (FDDKI mice), an AD-like dementia associated with mutations in the regulator of APP processing BRI2/ITM2B, also characterized by tauopathy. Some evidence suggests that caspases are activated early in the progression of AD and may play a role in neuronal loss and NFT pathology. Tau is cleaved at D421 (Tau) by executioner caspases. Following caspase-cleavage, Tau facilitates nucleation-dependent filament formation and adopts a conformational change recognized by MC1, an early pathological tau marker. Tau can be phosphorylated and recognized by the NFT antibody PHF-1. In AD brains, Tau associates with markers of NFTs and correlates with cognitive decline. Furthermore, Tau initiates NFT formation and can exert toxic effects. These findings have led to the hypothesis that Tau is a critical toxic moiety underlying neurodegeneration. To directly test this theory, we have generated knock-in mice in which the endogenous tau codon GAC in exon 12, encoding for D421, has been mutated into AAC, which now encodes for an asparagine (N). These knock-in mice, called TauD421N, express a tau mutant, taucas/res that cannot be cleaved by caspases and therefore cannot generate Tau. TauD421N mice will be used to determine whether Tau mediates synaptic plasticity and memory deficits in animal models of AD and FDD, two neurodegenerative disorders in which tau plays a pathogenic role. Our studies will speak to whether modulation of Tau formation is a potential strategy for the treatment of AD and other neurodegenerative disorders mediated by neuro-toxic tau forms.