Mechanisms of neuromuscular junction formation

神经肌肉接头（NMJ）是运动神经元和肌肉纤维之间形成的突触， 对理解突触发生的一般原理做出了巨大贡献， 神经肌肉紊乱的症状NMJ的形成需要运动神经元之间的密切相互作用 和肌肉纤维。例如，在顺行信号传导中，运动神经元释放聚集蛋白， LRP 4，LDL受体家族的成员，在肌肉细胞中激活受体酪氨酸 激酶MuSK，两者都是NMJ形成所需的。MuSK的下游情况不佳 理解，除了乙酰胆碱受体浓度绝对需要衔接蛋白rapsyn。 然而，确切地说，信号是如何从MuSK激活转导到AChR浓度的， 很好理解。另一方面，已知骨骼肌对发育至关重要， 运动神经元轴突终末。与顺行调节相反， 关于肌肉对突触前分化的逆向调节的分子机制 纤维在初步研究中，我们发现经典的衔接蛋白rapsyn是一个E3 连接酶携带酶促反应所必需的单个残基突变的敲入小鼠 活动不能形成NMJ。我们的研究结果表明rapsyn有助于AChR 通过促进neddylation聚类。这些观察发现了一个以前未被认识到的 rapsyn的酶活性和neddylation在突触形成中的作用。LRP 4的研究 表明它可以通过独立于MuSK的机制调节发育中的轴突。的 该提案将检验两个假设。首先，MuSK增加rapsyn酶活性， neddylation用于AChR聚类和NMJ形成。第二，肌肉调节突触前 通过LRP 4进行区分。这一建议的结果将提供一个更好的理解细胞作为 以及哺乳动物NMJ形成的分子机制，并有助于更好地 了解神经肌肉疾病的致病机制。

The neuromuscular junction (NMJ), a synapse formed between motoneurons and muscle fibers, has contributed greatly to understanding of the general principles of synaptogenesis, as well as of neuromuscular disorders. NMJ formation requires intimate interaction between motoneurons and muscle fibers. For example, in antegrade signaling, motoneurons release agrin that binds LRP4, a member of the LDL receptor family, in muscle cells to activate the receptor tyrosine kinase MuSK, both of which are required for NMJ formation. Downstream of MuSK was not well understood, except that AChR concentration absolutely requires the adapter protein rapsyn. However, exactly how signals are transduced from MuSK activation to AChR concentration is not well understood. On the other hand, skeletal muscles are known to be critical to the development of axon terminals of motoneurons. In contrast to antegrade regulation, much less is understood about molecular mechanisms of retrograde regulation of presynaptic differentiation by muscle fibers. In preliminary studies, we discovered that the classic adaptor protein rapsyn is an E3 ligase. Knockin mice carrying the mutation of a single residue necessary for the enzymatic activity are unable to form the NMJ. Our results suggest that rapsyn contributes to AChR clustering by promoting neddylation. These observations identify a previously unappreciated enzymatic activity of rapsyn and a role of neddylation in synapse formation. Our studies of LRP4 suggest that it could regulate developing axons by mechanisms independent of MuSK. The proposal will test two hypotheses. First, MuSK increases rapsyn enzyme activity to promote neddylation for AChR clustering and NMJ formation. Second, muscle regulates presynaptic differentiation via LRP4. Results of this proposal will provide a better understanding of cellular as well as molecular mechanisms of mammalian NMJ formation and contribute to a better understanding of pathogenic mechanisms of neuromuscular disorders.