Vorobyeva Anna G, Saunders Aleister J
Department of Biology, Drexel University, Philadelphia, PA USA.
Cilia. 2018 Aug 17;7:5. doi: 10.1186/s13630-018-0059-y. eCollection 2018.
Primary cilia are small non-motile microtubule and cell membrane protrusions expressed on most vertebrate cells, including cortical and hippocampal neurons. These small organelles serve as sensory structures sampling the extracellular environment and reprogramming the transcriptional machinery in response to environmental change. Primary cilia are decorated with a variety of receptor proteins and are necessary for specific signaling cascades such as the Sonic hedgehog (Shh) pathway. Disrupting cilia structure or function results in a spectrum of diseases collectively referred to as ciliopathies. Common to human ciliopathies is cognitive impairment, a symptom also observed in Alzheimer's disease (AD). One hallmark of AD is accumulation of senile plaques composed of neurotoxic Amyloid-β (Aβ) peptide. The Aβ peptide is generated by the proteolytic cleavage of the amyloid precursor protein (APP). We set out to determine if Aβ affects primary cilia structure and the Shh signaling cascade.
We utilized in vitro cell-based assays in combination with fluorescent confocal microscopy to address our study goals. Shh signaling and cilia structure was studied using two different cell lines, mouse NIH3T3 and human HeLa cells. To investigate how Aβ levels affect Shh signaling and cilia structure in these cells, we utilized naturally secreted Aβ as well as synthetic Aβ. Effects on Shh signaling were assessed by luciferase activity while cilia structure was analyzed by fluorescent microscopy.
Here, we report that APP localizes to primary cilia and Aβ treatment results in distorted primary cilia structure. In addition, we demonstrate that Aβ treatment interrupts canonical Shh signal transduction.
Overall, our study illustrates that Aβ can alter primary cilia structure suggesting that elevated Aβ levels, like those observed in AD patients, could have similar effects on neuronal primary cilia in the brain. Additionally, our study suggests that Aβ impairs the Shh signaling pathway. Together our findings shed light on two novel targets for future AD therapeutics.
初级纤毛是大多数脊椎动物细胞（包括皮质和海马神经元）上表达的小型非运动性微管和细胞膜突出物。这些小细胞器作为感觉结构，对细胞外环境进行采样，并根据环境变化对转录机制进行重新编程。初级纤毛装饰有多种受体蛋白，是特定信号级联反应（如音猬因子（Shh）信号通路）所必需的。破坏纤毛结构或功能会导致一系列统称为纤毛病的疾病。人类纤毛病的共同特征是认知障碍，这也是阿尔茨海默病（AD）中观察到的一种症状。AD的一个标志是由神经毒性淀粉样β（Aβ）肽组成的老年斑的积累。Aβ肽是由淀粉样前体蛋白（APP）的蛋白水解切割产生的。我们着手确定Aβ是否影响初级纤毛结构和Shh信号级联反应。
我们利用基于细胞的体外试验结合荧光共聚焦显微镜来实现我们的研究目标。使用两种不同的细胞系，小鼠NIH3T3和人HeLa细胞，研究Shh信号传导和纤毛结构。为了研究Aβ水平如何影响这些细胞中的Shh信号传导和纤毛结构，我们使用了天然分泌的Aβ以及合成Aβ。通过荧光素酶活性评估对Shh信号传导的影响，同时通过荧光显微镜分析纤毛结构。
在此，我们报告APP定位于初级纤毛，Aβ处理导致初级纤毛结构扭曲。此外，我们证明Aβ处理会中断经典的Shh信号转导。
总体而言，我们的研究表明Aβ可以改变初级纤毛结构，这表明AD患者中观察到的Aβ水平升高可能对大脑中的神经元初级纤毛有类似影响。此外，我们的研究表明Aβ会损害Shh信号通路。我们的研究结果共同揭示了未来AD治疗的两个新靶点。
