线粒体蛋白CHCHD10稳定突触的作用及机制研究

Motoneuron controls skeletal muscle through neuromuscular junction (NMJ). Mitochondria enrich in the synapse and are required for normal neurotransmission. ALS is caused by the loss of controls between motoneuron and muscle. Synaptic impairment occurs earlier than neuron loss. Mitochondria CHCHD10 is a new ALS risk gene. The roles and mechanisms of CHCHD10 in regulating NMJ remain unclear. We found that CHCHD10 specifically expresses in skeletal muscle but not in spinal cord or sciatic nerve. Loss of CHCHD10 in muscle cell leads to mitochondria fragmentation and ATP reduction. AChR cannot be clustered efficiently by agrin in cultured myotube, and NMJ cannot be stabilized in the CHCHD10 ablated muscle in vivo. These results suggest that CHCHD10 regulates synaptic mitochondria and NMJ homeostasis. To further clarify the underlying mechanisms, with a combination of strategies with molecular biology, cell biology, morphology, and physiology, we would use conditional knockout mouse to study the roles of CHCHD10 in the postsynaptic receptor stabilization and motoneuron survival by regulating Agrin signal. Our research would clarify the regulatory mechanisms in mitochondria and NMJ maintenance. It would also help to understand pathological mechanisms of neuromuscular disorders caused by mitochondria impairment.

运动神经元通过神经肌肉接头NMJ控制骨骼肌运动，NMJ障碍是ALS的早期症状。已知线粒体富集在NMJ突触后膜区域。肌萎缩脊髓侧索硬化症ALS新基因CHCHD10编码线粒体蛋白，其在调节NMJ神经递质传递的作用和机制，目前尚不清楚。我们发现CHCHD10特异定位在骨骼肌突触后膜，参与线粒体形态和功能维持。在肌肉细胞中敲除CHCHD10，可以抑制Agrin引起的AChR聚集。小鼠骨骼肌中敲除CHCHD10，可以导致NMJ破碎。这些结果提示：CHCHD10通过突触后膜线粒体稳定NMJ。为进一步揭示CHCHD10作用机制，我们将利用基因敲除小鼠，结合分子生物学，细胞生物学，形态和生理学手段，阐明其通过Agrin信号促进AChR聚集，维持突触后膜稳定和运动神经元存活。本课题将有助于理解线粒体和NMJ稳定的关系，为治疗NMJ无法稳定的神经退行性疾病提供新的思路。

运动神经元通过神经肌肉接头NMJ控制骨骼肌运动，NMJ障碍是ALS的早期症状。已知线粒体富集在NMJ突触后膜区域。肌萎缩脊髓侧索硬化症ALS新基因CHCHD10编码线粒体蛋白，其在调节NMJ神经递质传递的作用和机制，目前尚不清楚。我们主要通过免疫组化、电生理、电镜以及行为学等实验技术手段来探究骨骼肌中CHCHD10对NMJ稳态的调控作用。我们发现CHCHD10特异性定位在骨骼肌突触后膜，参与线粒体形态和功能维持。骨骼肌细胞内条件性敲除CHCHD10的小鼠，表现出抓力下降，NMJ神经信号传递功能障碍。沉默或敲除CHCHD10基因能引起细胞或组织ATP水平下降，提示ATP 的产生依赖于CHCHD10的表达。进一步实验证实了ATP能促进Agrin诱导的AChR聚集，并促进转录因子GABPα与AChR亚基基因的启动子结合。这提示着ATP通过促进AChR亚基基因表达，进而促使Agrin诱导的AChR聚集。 该研究提示CHCHD10是维持线粒体结构和产生ATP所必需的，并且ATP通过增强GABPα介导的AChR亚基基因表达来促进Agrin诱导的AChR聚集，从而维持NMJ正常的功能。本课题提示了突触后CHCHD10对调节NMJ的稳态起着重要作用，加深了我们对线粒体和NMJ稳定关系的理解，为治疗NMJ无法稳定的神经退行性疾病提供新的思路。

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