磷脂结合蛋白Annexin A11相变在肌萎缩性脊髓侧索硬化症中的作用及机制研究

肌萎缩性脊髓侧索硬化症ALS是一种致命性神经退行性疾病，它的治疗是个世界性难题。Annexin A11（A11）是我国ALS患者特异的关键致病蛋白。我们的前期研究发现，A11可通过相变连接核糖核蛋白（RNP）颗粒和细胞器膜结构，发挥向神经远端运输RNA的功能。我们推测，突变体A11的异常相变通过影响RNP-A11-细胞器互作网络，和神经细胞不同局部区域内RNP与多种细胞器的结合和释放，从而影响轴突末端的局部蛋白质翻译，并导致神经元死亡。本研究拟利用APEX蛋白组学研究突变体A11异常相变介导的RNP颗粒-A11-细胞器互作的变化，进而以突变后出现的新增互作为靶标库，筛选可降低突变体毒性的靶基因；并通过局部精准光控相变技术研究突变体A11异常相变对局部区域内RNP搭载和释放的影响及导致神经元死亡的途径，阐明我国特异突变体A11导致运动神经元死亡的分子机制，为ALS治疗提供新的靶标和理论依据。

ALS is a fatal neurodegenerative disease with poor prognosis and no effective treatment. Annexin A11 (A11) is a key protein mutated in Chinese ALS patients. Our latest research revealed that A11 is a phase separation protein and it can transport mRNA to axon terminal by tethering RNP granule to membrane of lysosome. Based on our findings, we hypothesize that abnormal phase separation of mutated A11 can change RNP-A11-organelle interactome and affect the tethering and release of RNP to/from organelle in different local regions within neuron, and thus inhibit mRNA transport and local protein synthesis in axon terminal, which leads to death of motor neuron. To prove our hypothesis and reveal molecular pathological mechanisms of Chinese specific A11 mutants, firstly, we will investigate protein networks involved in the RNP granule-A11-membrane-bound organelles interaction using APEX Quantitative Proteomics technique. Using new gained interactions induced by A11 mutants as small targets library, we will try to identify targets that can decrease toxicity induced by mutant A11 using CRISPR/Cas9 screening. Secondly, to investigate how phase separation of A11 in different parts of neuron affects RNP cargo loading/release from organelles, we will use optogenetic system to induce local phase separation of A11 within different local regions of neuron and check whether it can affect mRNA-long-distant-transport function of A11, local protein synthesis in axon terminal, and unravel the death pathway of motor neuron. The results from this research project would provide theoretical guidance and potential drug targets for further development of diagnostic methods and treatments for Chinese ALS.