LRP4 signaling in neuromuscular junction formation

LRP4 信号在神经肌肉接头形成中的作用

• 批准号: 9604664
• 负责人: Lin Mei
• 金额: $ 16.22万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9604664
• 关键词: LRP4; signaling; neuromuscular; junction; formation

DESCRIPTION (provided by applicant): The multi-PI proposal is to investigate how LRP4 regulates the formation of the neuromuscular junction (NMJ), a synapse that has contributed greatly to understanding of synaptogenesis and of neuromuscular disorders. NMJ formation requires precise interaction between motoneurons (MNs) and muscle fibers. In advance of MN arrival, muscle fibers are prepatterned with small, primitive AChR clusters, which are believed to be critical for NMJ formation although genetic evidence is lacking at the moment. In an established pathway, MNs release agrin to promote postsynaptic development. It binds to LRP4 which acts in cis to stimulate the receptor tyrosine kinase MuSK; and ensuing signaling events lead to AChR clustering. However, little is known of how signals are transduced from agrin to MuSK. In preliminary studies, we solved the first crystal structure of an agrin-LRP4 complex, which provides insight into the initial step of the agrin signaling cascade. Our studies of cell-specific knockout (KO) and double KO mice revealed novel functions of LRP4 in NMJ formation. For example, muscle LRP4 may be critical for presynaptic differentiation during initial steps of NMJ formation. On the other hand, MN LRP4 may serve as agrin's receptor in trans to induce AChR clusters. Many of these findings are unexpected and raise critical questions. How does LRP4 relay the signal from agrin to MuSK? Is MN LRP4 sufficient to induce NMJ formation? Is muscle fiber prepatterning critical for NMJ formation? Is the synaptogenic activity critical for NMJ formation? How does muscle LRP4 regulate motor nerve terminal differentiation? To address these questions, we will 1) understand how signal is transduced from LRP4 to MuSK by solving the structure of the agrin- LRP4-MuSK complex; 2) determine if MN LRP4 is sufficient to induce NMJ formation and if muscle fiber prepatterning is necessary for NMJ formation; and 3) investigate mechanisms by which LRP4 in muscle cells controls presynaptic differentiation. The research represents a synergistic strategy that leverages complementary expertise, strengths and existing resources of the two labs. Results will provide a better understanding of cellular as well as molecular mechanisms of mammalian NMJ formation. Pathogenesis of neuromuscular disorders is known to involve abnormal NMJ structure and function. In fact, mutations of agrin/LRP4/MuSK signaling proteins have been implicated in congenital myasthenic syndrome (CMS). Recent evidence from various laboratories including ours indicates that patients with myasthenia gravis (MG) develop antibodies against MuSK and LRP4. Therefore, our research will contribute to a better understanding of pathogenic mechanisms of these neuromuscular disorders.

描述（由申请人提供）：多 PI 提案旨在研究 LRP4 如何调节神经肌肉接头 (NMJ) 的形成，神经肌肉接头 (NMJ) 是一种突触，为理解突触发生和神经肌肉疾病做出了巨大贡献。 NMJ 的形成需要运动神经元 (MN) 和肌纤维之间的精确相互作用。在 MN 到达之前，肌肉纤维已预先形成小型原始 AChR 簇，尽管目前缺乏遗传证据，但人们认为这些簇对于 NMJ 的形成至关重要。在既定途径中，微核释放集聚蛋白以促进突触后发育。它与 LRP4 结合，LRP4 顺式作用，刺激受体酪氨酸激酶 MuSK；随后的信号事件导致 AChR 聚集。然而，人们对信号如何从 agrin 转导为 MuSK 知之甚少。在初步研究中，我们解析了集聚蛋白-LRP4复合物的第一个晶体结构，这提供了对集聚蛋白信号级联的初始步骤的深入了解。我们对细胞特异性敲除 (KO) 和双 KO 小鼠的研究揭示了 LRP4 在 NMJ 形成中的新功能。例如，肌肉 LRP4 可能对于 NMJ 形成的初始步骤中的突触前分化至关重要。另一方面，MN LRP4 可能作为反式集聚蛋白受体来诱导 AChR 簇。其中许多发现是出乎意料的，并提出了关键问题。 LRP4 如何将信号从 agrin 传递到 MuSK？ MN LRP4 是否足以诱导 NMJ 形成？肌纤维预模式化对于 NMJ 形成至关重要吗？突触活性对于 NMJ 形成至关重要吗？肌肉LRP4如何调节运动神经末梢分化？为了解决这些问题，我们将 1) 通过解析 agrin-LRP4-MuSK 复合物的结构，了解信号如何从 LRP4 转导到 MuSK； 2)确定MN LRP4是否足以诱导NMJ形成以及肌纤维预图案化对于NMJ形成是否是必要的； 3) 研究肌肉细胞中 LRP4 控制突触前分化的机制。该研究代表了一种协同策略，利用了两个实验室的互补专业知识、优势和现有资源。结果将有助于更好地理解哺乳动物 NMJ 形成的细胞和分子机制。已知神经肌肉疾病的发病机制涉及异常的 NMJ 结构和功能。事实上，agrin/LRP4/MuSK 信号蛋白的突变与先天性肌无力综合征 (CMS) 有关。来自包括我们在内的各个实验室的最新证据表明，重症肌无力 (MG) 患者会产生针对 MuSK 和 LRP4 的抗体。因此，我们的研究将有助于更好地了解这些神经肌肉疾病的致病机制。