Chen Yanchun, Wang Qing, Wang Qiaozhen, Liu Huancai, Zhou Fenghua, Zhang Yawen, Yuan Meng, Zhao Chunyan, Guan Yingjun, Wang Xin
Department of Histology and Embryology, Weifang Medical UniversityWeifang, Shandong, PR China.
Department of Human Anatomy, Weifang Medical UniversityWeifang, Shandong, PR China.
Am J Transl Res. 2017 Oct 15;9(10):4627-4639. eCollection 2017.
Amyotrophic lateral sclerosis (ALS) is a chronic neurodegenerative disease characterized by progressive degeneration of motor neurons. The pathogenesis of ALS remains largely unknown. RNA helicase DDX3 is a multifunctional protein involved in several steps of gene expression. Casein kinase 1ε (CK1ε) is an important signal molecule of Wnt signaling pathway and is closely related to neurite growth. However, the roles of DDX3 and CK1ε in the pathogenesis of ALS remain unclear. In this study, we first investigated the expression of DDX3 and CK1ε in the spinal cord of SOD1-G93A ALS transgenic mice using RT-PCR, Western blot and immunohistochemical technique. Results showed that the altered expression of DDX3 and CK1ε was found in the spinal cord of ALS mice. DDX3 and CK1ε positive cells were mainly distributed in the anterior horn of spinal cord and co-localized with neurons not with glial cells, suggesting that the altered expression of DDX3 and CK1ε was closely related to motor neuron degeneration of ALS. Moreover, we selected NSC34 cell line and transfected pEGFP-G93A-SOD1 plasmid to further examine the mechanism. Knockdown of DDX3 that uses small interfering RNA (siRNA) decreased the mRNA and protein levels of CK1ε significantly and inhibited neurite outgrowth of SOD1 mutant NSC34 cells in vitro. Co-immunoprecipitation kit confirmed that DDX3 could band with CK1ε in vivo. Our data suggested that DDX3 binding with CK1ε was closely related to motor neuron degeneration of ALS by affecting neurite outgrowth. Thus, elucidating the underlying mechanisms of ALS is crucial for future development of ALS treatments.
肌萎缩侧索硬化症（ALS）是一种慢性神经退行性疾病，其特征在于运动神经元的进行性退化。ALS的发病机制在很大程度上仍然未知。RNA解旋酶DDX3是一种多功能蛋白质，参与基因表达的多个步骤。酪蛋白激酶1ε（CK1ε）是Wnt信号通路的重要信号分子，与神经突生长密切相关。然而，DDX3和CK1ε在ALS发病机制中的作用仍不清楚。在本研究中，我们首先使用RT-PCR、蛋白质免疫印迹和免疫组织化学技术研究了DDX3和CK1ε在SOD1-G93A ALS转基因小鼠脊髓中的表达。结果表明，在ALS小鼠的脊髓中发现了DDX3和CK1ε的表达改变。DDX3和CK1ε阳性细胞主要分布在脊髓前角，与神经元共定位，而非与神经胶质细胞共定位，这表明DDX3和CK1ε的表达改变与ALS的运动神经元退化密切相关。此外，我们选择了NSC34细胞系并转染了pEGFP-G93A-SOD1质粒以进一步研究其机制。使用小干扰RNA（siRNA）敲低DDX3可显著降低CK1ε的mRNA和蛋白质水平，并在体外抑制SOD1突变型NSC34细胞的神经突生长。免疫共沉淀试剂盒证实DDX3在体内可与CK1ε结合。我们的数据表明，DDX3与CK1ε结合通过影响神经突生长与ALS的运动神经元退化密切相关。因此，阐明ALS的潜在机制对于ALS治疗的未来发展至关重要。
