Pathological conformations involving the amino terminus of tau occur early in Alzheimer's disease and are differentially detected by monoclonal antibodies.
Pathological conformations involving the amino terminus of tau occur early in Alzheimer's disease and are differentially detected by monoclonal antibodies.
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DOI：
10.1016/j.nbd.2016.05.016
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发表时间：
2016-10
影响因子：
6.1
通讯作者：
Kanaan NM
中科院分区：
文献类型：
--
作者：
Combs B;Hamel C;Kanaan NM
涉及tau蛋白氨基末端的构象变化是阿尔茨海默病和其他tau病中与tau病理相关的最早的变化之一。Tau的这一区域包含一个磷酸酶激活结构域(PAD)，该结构域在蛋白的病理形式中异常暴露，这一事件与顺行快速轴突运输的中断有关。我们利用四种识别tau、TNT1、TNT2(一种新抗体)、Tau12和Tau13氨基端的抗体来进一步研究这一重要区域。通过扫描重组tau蛋白中的丙氨酸突变，我们提纯了每个抗体的表位。我们检查了抗体在非变性和变性试验中特异性标记病理tau的相对能力，以深入了解PAD暴露的一些机制细节。然后，我们测定了不同疾病阶段的人类海马区切片中每种抗体标记的tau病理模式，以便在疾病进展的背景下表征PAD暴露的特征。抗体的反应性特征分为两组。TNT1和TNT2识别氨基酸7-12中的表位，并以构象依赖的方式特异性地识别阿尔茨海默病脑中的重组tau聚集体和病理性tau。这些抗体标记了Braak早期神经元的早期缠绕前病理，并与典型的神经纤维缠绕中的噻嗪红(一种纤维病理的标志)共同定位。然而，TNT1和TNT2的晚期缠结为阴性，这表明在缠结进化的后期阶段TNT1和TNT2的表位丢失。相比之下，Tau12和Tau13在氨基末端都发现了不连续的表位，在生化和组织免疫组织学检测中无法区分正常和病理的tau。尽管这些表位非常接近，但抗体识别tau病变的能力明显不同，这表明并不是所有的N端tau抗体都能检测到涉及PAD暴露的构象变化，并且N端相对离散的区域(即氨基酸7-12，TNT1和TNT2表位)是正常tau和病理性tau之间差异的核心。早期Tau病理中PAD的出现和晚期缠结中PAD的消失表明，毒性形式的Tau与最早形式的Tau沉积有关。总之，这些发现表明，TNT抗体是PAD早期构象显示的有用标记物，并提供了有关构象变化的信息，这些信息在tau病理的毒性机制中具有潜在的意义。
Conformational changes involving the amino terminus of the tau protein are among the earliest alterations associated with tau pathology in Alzheimer’s disease and other tauopathies. This region of tau contains a phosphatase-activating domain (PAD) that is aberrantly exposed in pathological forms of the protein, an event that is associated with disruptions in anterograde fast axonal transport. We utilized four antibodies that recognize the amino terminus of tau, TNT1, TNT2 (a novel antibody), Tau12, and Tau13, to further study this important region. Using scanning alanine mutations in recombinant tau proteins, we refined the epitopes of each antibody. We examined the antibodies’ relative abilities to specifically label pathological tau in non-denaturing and denaturing assays to gain insight into some of the mechanistic details of PAD exposure. We then determined the pattern of tau pathology labeled by each antibody in human hippocampal sections at various disease stages in order to characterize PAD exposure in the context of disease progression. The characteristics of reactivity for the antibodies fell into two groups. TNT1 and TNT2 recognized epitopes within amino acids 7–12 and specifically identified recombinant tau aggregates and pathological tau from Alzheimer’s disease brains in a conformation-dependent manner. These antibodies labeled early pre-tangle pathology from neurons in early Braak stages and colocalized with thiazine red, a marker of fibrillar pathology, in classic neurofibrillary tangles. However, late tangles were negative for TNT1 and TNT2 indicating a loss of the epitope in later stages of tangle evolution. In contrast, Tau12 and Tau13 both identified discontinuous epitopes in the amino terminus and were unable to differentiate between normal and pathological tau in biochemical and tissue immunohistological assays. Despite the close proximity of these epitopes, the antibodies demonstrated remarkably different abilities to identify pathological changes in tau indicating that detection of conformational alterations involving PAD exposure is not achieved by all N-terminal tau antibodies and that a relatively discrete region of the N-terminus (i.e., amino acids 7–12, the TNT1 and TNT2 epitope) is central to the differences between normal and pathological tau. The appearance of PAD in early tau pathology and its disappearance in late-stage tangles suggest that toxic forms of tau are associated with the earliest forms of tau deposits. Collectively, these findings demonstrate that the TNT antibodies are useful markers for early conformational display of PAD and provide information regarding conformational changes that have potential implications in the toxic mechanisms of tau pathology.