Department for Molecular and Cellular Mechanisms of Neurodegeneration, University of Leipzig, Paul Flechsig Institute of Brain Research, Leipzig, Germany.
Neuropathol Appl Neurobiol. 2014 Dec;40(7):815-32. doi: 10.1111/nan.12163.
Neurodegeneration in Alzheimer's disease (AD) is characterized by pathological protein aggregates and inadequate activation of cell cycle regulating proteins. Recently, Smad proteins were identified to control the expression of AD relevant proteins such as APP, CDK4 and CDK inhibitors, both critical regulators of cell cycle activation. This might indicate a central role for Smads in AD pathology where they show a substantial deficiency and disturbed subcellular distribution in neurones. Still, the mechanisms driving relocation and decrease of neuronal Smad in AD are not well understood. However, Pin1, a peptidyl-prolyl-cis/trans-isomerase, which allows isomerization of tau protein, was recently identified also controlling the fate of Smads. Here we analyse a possible role of Pin1 for Smad disturbances in AD.
Multiple immunofluorescence labelling and confocal laser-scanning microscopy were performed to examine the localization of Smad and Pin1 in human control and AD hippocampi. Ectopic Pin1 expression in neuronal cell cultures combined with Western blot analysis and immunoprecipitation allowed studying Smad level and subcellular distribution. Luciferase reporter assays, electromobility shift, RNAi-technique and qRT-PCR revealed a potential transcriptional impact of Smad on Pin1 promoter.
We report on a colocalization of phosphorylated Smad in AD with Pin1. Pin1 does not only affect Smad phosphorylation and stability but also regulates subcellular localization of Smad2 and supports its binding to phosphorylated tau protein. Smads, in turn, exert a negative feed-back regulation on Pin1.
Our data suggest both Smad proteins and Pin1 to be elements of a vicious circle with potential pathogenetic significance in AD.
阿尔茨海默病（AD）中的神经退行性变的特征是病理性蛋白聚集和细胞周期调节蛋白的激活不足。最近，Smad 蛋白被鉴定为控制 AD 相关蛋白（如 APP、CDK4 和 CDK 抑制剂）的表达，这些蛋白都是细胞周期激活的关键调节剂。这表明 Smad 在 AD 病理中具有核心作用，在神经元中它们表现出明显的缺乏和紊乱的亚细胞分布。然而，导致 AD 中神经元 Smad 重定位和减少的机制尚不清楚。然而，最近发现一种肽基脯氨酰顺/反式异构酶 Pin1 ，它允许 tau 蛋白的异构化，也控制 Smad 的命运。在这里，我们分析了 Pin1 对 AD 中 Smad 紊乱的可能作用。
通过多重免疫荧光标记和共聚焦激光扫描显微镜检查，研究了 Smad 和 Pin1 在人对照和 AD 海马中的定位。神经元细胞培养中的异位 Pin1 表达与 Western blot 分析和免疫沉淀相结合，允许研究 Smad 水平和亚细胞分布。荧光素酶报告基因分析、电泳迁移率变动分析、RNAi 技术和 qRT-PCR 揭示了 Smad 对 Pin1 启动子的潜在转录影响。
我们报告了 AD 中磷酸化 Smad 与 Pin1 的共定位。Pin1 不仅影响 Smad 的磷酸化和稳定性，还调节 Smad2 的亚细胞定位，并支持其与磷酸化 tau 蛋白的结合。Smad 反过来对 Pin1 施加负反馈调节。
我们的数据表明 Smad 蛋白和 Pin1 都是一个恶性循环的元素，在 AD 中具有潜在的发病意义。
