运动神经元存活基因1（SMN1）编码的SMN蛋白以复合体的形式参与snRNP的组装，该基因突变导致一种儿童常见的隐性遗传病——脊肌萎缩症（SMA）。无义突变的致病机制通常为无义突变介导mRNA降解（NMD），而课题组前期发现SMN1无义突变p.Glu14X出现了NMD逃逸，很可能因影响SMN功能而致病。为明确p.Glu14X的致病机制，本课题通过体外构建野生型/突变型表达质粒，共转染HEK293-T细胞，应用免疫共沉淀、Western Blot等方法评价该突变对SMN自我寡聚化以及对SMN与复合体中其他蛋白结合力的影响；应用细胞免疫荧光等方法评价该突变对SMN亚细胞定位的影响。通过建立SMA患儿淋巴细胞系，应用脉冲示踪技术、体外转录、免疫沉淀等方法分别经体内和体外实验评价突变对snRNP组装活性的影响，以探讨p.Glu14X的致病机制，推进SMA的病因研究并为SMA个体化治疗提供理论依据。

The survival motor neuron gene 1 (SMN1) encodes the SMN protein, which forms a complex with other factors involved in the assembly of small nuclear ribonucleoprotein (snRNP). Spinal muscular atrophy (SMA) is a common autosomal-recessive hereditary disease in children, which caused by mutations in the SMN1. It was well known that nonsense mutation can cause many genetic disorders by nonsense mutation-mediated mRNA decay (NMD). We previously found that nonsense mutation p.Glu14X in SMN1 escaped NMD, therefore this mutation is possibly to affect the function of SMN protein rather than protein expression. In order to understand the pathogenic mechanism of p.Glu14X mutation, we construct plasmids expressing epitope-tagged wild-type or mutant SMN to transfect HEK293-T cells. Several approaches including co-immunoprecipitation and western blot techniques are used to determine the effects of this mutation on SMN self-oligomerization and its ability to bind to other proteins. Cytoimmunofluorescence is used to ascertain the impact of mutation on the subcellular localization of SMN. By establishing lymphocyte cell lines from SMA children and using the pulse-chase technology, in vitro transcription, co-immunoprecipitation, etc., the assembly activity of snRNP is measured in vivo and in vitro. In this study, we explored the pathogenic mechanisms of p. Glu14X mutation in the SMN1 with the purpose of promoting the understanding of pathogenesis and individualized treatment of SMA.