Abnormal accumulation of misfolded and hyperphosphorylated tau protein in brain is the defining feature of several neurodegenerative diseases called tauopathies, including Alzheimer’s disease (AD). In AD, this pathological change is reflected by highly specific cerebrospinal fluid (CSF) tau biomarkers, including both phosphorylated and non-phosphorylated variants. Interestingly, despite tau pathology being at the core of all tauopathies, CSF tau biomarkers remain unchanged in certain tauopathies, e.g., progressive supranuclear palsy (PSP), Pick’s disease (PiD), and corticobasal neurodegeneration (CBD). To better understand commonalities and differences between tauopathies, we report a multiplex assay combining immunoprecipitation and high-resolution mass spectrometry capable of detecting and quantifying peptides from different tau protein isoforms as well as non-phosphorylated and phosphorylated peptides, including those carrying multiple phosphorylations. We investigated the tau proteoforms in soluble and insoluble fractions of brain tissue from subjects with autopsy-confirmed tauopathies, including sporadic AD (n = 10), PSP (n = 11), PiD (n = 10), and CBD (n = 10), and controls (n = 10). Our results demonstrate that non-phosphorylated tau profiles differ across tauopathies, generally showing high abundance of microtubule-binding region (MTBR)-containing peptides in insoluble protein fractions compared with controls; the AD group showed 12–72 times higher levels of MTBR-containing aggregates. Quantification of tau isoforms showed the 3R being more abundant in PiD and the 4R isoform being more abundant in CBD and PSP in the insoluble fraction. Twenty-three different phosphorylated peptides were quantified. Most phosphorylated peptides were measurable in all investigated tauopathies. All phosphorylated peptides were significantly increased in AD insoluble fraction. However, doubly and triply phosphorylated peptides were significantly increased in AD even in the soluble fraction. Results were replicated using a validation cohort comprising AD (n = 10), CBD (n = 10), and controls (n = 10). Our study demonstrates that abnormal levels of phosphorylation and aggregation do indeed occur in non-AD tauopathies, however, both appear pronouncedly increased in AD, becoming a distinctive characteristic of AD pathology.

中文翻译：

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tau蛋白病中的tau蛋白分析：一项人脑研究


大脑中错误折叠和过度磷酸化 tau 蛋白的异常积累是包括阿尔茨海默病 (AD) 在内的几种称为 tau 病的神经退行性疾病的定义特征。在 AD 中，这种病理变化通过高度特异性的脑脊液 (CSF) tau 生物标志物反映，包括磷酸化和非磷酸化变体。有趣的是，尽管 tau 病理是所有 tau 病的核心，但脑脊液 tau 生物标志物在某些 tau 病中保持不变，例如进行性核上性麻痹 (PSP)、皮克氏病 (PiD) 和皮质基底神经变性 (CBD)。为了更好地了解 tau 蛋白病之间的共性和差异，我们报告了一种结合免疫沉淀和高分辨率质谱的多重检测，能够检测和定量不同 tau 蛋白亚型的肽以及非磷酸化和磷酸化肽，包括那些携带多重磷酸化的肽。我们研究了尸检证实患有 tau蛋白病的受试者脑组织中可溶性和不溶性部分的 tau 蛋白，包括散发性 AD (n = 10)、PSP (n = 11)、PiD (n = 10) 和 CBD (n = 10) ）和对照（n = 10）。我们的结果表明，不同 tau 蛋白病的非磷酸化 tau 谱存在差异，与对照相比，不溶性蛋白组分中通常显示出高丰度的含有微管结合区 (MTBR) 的肽； AD 组的 MTBR 聚集体含量高出 12-72 倍。 tau 同工型的定量显示，3R 在 PiD 中更丰富，4R 同工型在不溶性组分中的 CBD 和 PSP 中更丰富。对二十三种不同的磷酸化肽进行了定量。 大多数磷酸化肽在所有研究的 tau蛋白病中都是可测量的。 AD 不溶部分中的所有磷酸化肽均显着增加。然而，即使在可溶性部分中，AD 中的双磷酸化肽和三磷酸化肽也显着增加。使用由 AD (n = 10)、CBD (n = 10) 和对照组 (n = 10) 组成的验证队列复制结果。我们的研究表明，异常水平的磷酸化和聚集确实发生在非AD tau蛋白病中，然而，两者在AD中均明显增加，成为AD病理学的一个显着特征。