肌萎缩侧索硬化症（ALS）是运动神经元退行性疾病，C9orf72基因非编码区的六核苷酸重复扩增是目前发现的最为常见的ALS/FTD的人类突变。但是C9orf72基因突变如何导致神经元发生退行的机制还需深入研究。本项目通过C9orf72-3*Flag基因敲入小鼠发现C9orf72蛋白可以结合DNA双链断裂修复中关键分子DNA-PKcs，并且C9orf72调控神经元中DNA双链断裂后修复通路的激活以及NHEJ修复途径的效率。通过建立小鼠的模型我们试图阐明C9orf72基因突变后DPR蛋白和C9orf72表达下降通过影响神经元中DNA损伤的产生和修复一起导致神经退行。最后我们试图在成熟神经元中敲除LIG4研究神经元中NHEJ修复途径缺失对于神经退行性疾病发生的作用。本项目的研究试图阐明DNA双链断裂在神经元中的积累可能是神经元从衰老走向退行的重要的原因。

Amyotrophic Lateral Sclerosis (ALS) is a progressive neurodegenerative disease, affecting both lower and upper motor neurons. The hexanucleotide GGGGCC repeat expansion in non-coding region of C9orf72 gene is the most frequent genetic cause of ALS. However, the pathological mechanisms of C9orf72 hexanucleotide expansion remain unclear. In this project, we take advantage of C9orf72-3*Flag knock-in mice to identify DNA-PKcs as a novel binding protein of C9orf72 by IP-MASS. DNA-PKcs is critical kinase for NHEJ repair pathway activation after DNA double-strand breaks induction. Activation of DNA repair pathway and NHEJ efficiency were significantly impaired in C9orf72-deficient neurons. Therefore, we propose that both gain of function and loss of function mechanisms may contribute to neurodegeneration through DNA damage induction and impairment of damage repair respectively. Finally, we will determine whether NHEJ repair is critical for neurodegeneration by deleting LIG4 gene in mature neurons. Our study will gain a better understanding of how aging neurons with accumulated DNA double-strand breaks eventually become degeneration.