Alzheimer disease-specific conformation of hyperphosphorylated paired helical filament-tau is polyubiquitinated through Lys-48, Lys-11, and Lys-6 ubiquitin conjugation
Alzheimer disease-specific conformation of hyperphosphorylated paired helical filament-tau is polyubiquitinated through Lys-48, Lys-11, and Lys-6 ubiquitin conjugation
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DOI：
10.1074/jbc.m512786200
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发表时间：
2006-04-21
影响因子：
4.8
通讯作者：
Yang, AJ
中科院分区：
文献类型：
--
作者：
Cripps, D;Thomas, SN;Yang, AJ
阿尔茨海默病（AD）的关键病理学标志之一是过度磷酸化的微管相关蛋白Tau的成对螺旋丝（PHF）的积累。采用串联质谱法来检查PHF-Tau的翻译后修饰，特别是蛋白磷酸化和泛素化，以阐明它们在阿尔茨海默病早期阶段的作用。通过MC 1单克隆抗体亲和纯化来自阿尔茨海默病脑的PHF-Tau，以分离人AD脑特有构象的PHF-Tau的可溶性级分。大量的磷酸化位点被确定采用数据依赖性中性丢失算法触发MS 3扫描的磷酸肽。结果发现，可溶性PHF-Tau在其微管结合结构域的残基Lys-254、Lys-311和Lys-353处被泛素化，这表明PHF-Tau的泛素化可能是比以前认为的更早的病理事件，并且泛素化可能在调节AD过程中微管的完整性中起调节作用。泛素本身的串联质谱数据表明PHF-Tau被三个多聚泛素键修饰，位于Lys-6、Lys-11和Lys-48。相对定量分析表明，赖氨酸-48连接的多聚泛素化是多聚泛素化的主要形式，小部分泛素连接在赖氨酸-6和赖氨酸-11。因为已知通过赖氨酸-48连接的多聚泛素链的修饰充当靶向蛋白以通过泛素-蛋白酶体系统降解的基本手段，并且已经报道在赖氨酸-6处的修饰抑制泛素依赖性蛋白质降解，所以泛素-蛋白酶体系统的失效可能在引发抗降解PHF缠结的形成中起作用。
One of the key pathological hallmarks of Alzheimer disease (AD) is the accumulation of paired helical filaments (PHFs) of hyperphosphorylated microtubule-associated protein Tau. Tandem mass spectrometry was employed to examine PHF-Tau post-translational modifications, in particular protein phosphorylation and ubiquitination, to shed light on their role in the early stages of Alzheimer disease. PHF-Tau from Alzheimer disease brain was affinity-purified by MC1 monoclonal antibody to isolate a soluble fraction of PHF-Tau in a conformation unique to human AD brain. A large number of phosphorylation sites were identified by employing a data-dependent neutral loss algorithm to trigger MS3 scans of phosphopeptides. It was found that soluble PHF-Tau is ubiquitinated at its microtubule-binding domain at residues Lys-254, Lys-311, and Lys-353, suggesting that ubiquitination of PHF-Tau may be an earlier pathological event than previously thought and that ubiquitination could play a regulatory role in modulating the integrity of microtubules during the course of AD. Tandem mass spectrometry data for ubiquitin itself indicate that PHF-Tau is modified by three polyubiquitin linkages, at Lys-6, Lys-11, and Lys-48. Relative quantitative analysis indicates that Lys-48-linked polyubiquitination is the primary form of polyubiquitination with a minor portion of ubiquitin linked at Lys-6 and Lys-11. Because modification by Lys-48-linked polyubiquitin chains is known to serve as the essential means of targeting proteins for degradation by the ubiquitin-proteasome system, and it has been reported that modification at Lys-6 inhibits ubiquitin-dependent protein degradation, a failure of the ubiquitin-proteasome system could play a role in initiating the formation of degradation-resistant PHF tangles.