Regulation of the Synaptic Localization of the nAChR

描述（由申请人提供）：突触高度专门用于有效的突触传递，突触前神经末梢中的递质释放位点与神经递质受体的突触后浓度对齐。在发育中的神经肌肉接头处，一种称为聚集蛋白的神经衍生因子指导乙酰胆碱受体的突触后定位，我们的目标是了解聚集蛋白作用的分子机制。具体来说，我们将测试的假设，聚集蛋白调节AChR的本地化诱导AChR的磷酸化，改变其与细胞内支架蛋白，rapsyn。我们的具体目标如下：（1）明确聚集蛋白诱导的β亚基磷酸化在AChR定位中的作用。我们将产生一个β亚基缺乏磷酸化位点的转基因小鼠，然后确定这种突变是否会损害AChR的突触定位。我们还将确定磷酸化是否特异性地调节突触后膜中受体的锚定、聚集和/或稳定。(2)明确rapsyn-mediated AChR定位的分子机制。在这里，我们将定义rapsyn的结合位点（S）的乙酰胆碱受体，并确定结合是否受聚集蛋白。此外，我们将研究如何agrin诱导rapsyn/AChR协会的变化介导受体定位。这项工作与先天性和自身免疫性肌无力综合征等神经肌肉疾病直接相关，这些疾病的突触AChR水平严重不足，导致突触功能受损和肌肉无力。我们的工作将有助于确定受体定位所涉及的机制，并可能提出稳定其在肌无力综合征或其他形式的神经肌肉疾病患者中的定位的策略。

DESCRIPTION (provided by applicant): Synapses are highly specialized for efficient synaptic transmission, with the transmitter release sites in the presynaptic nerve terminal being aligned with postsynaptic concentrations of the neurotransmitter receptor. At the developing neuromuscular junction, a nerve-derived factor called agrin directs the postsynaptic localization of the acetylcholine receptor, and our goal is to understand the molecular mechanisms that underlie agrin's action. Specifically, we will test the hypothesis that agrin regulates AChR localization by inducing phosphorylation of the AChR and altering its association with the intracellular scaffolding protein, rapsyn. We have the following specific aims: (1) To define the role of agrin-induced phosphoryation of the beta subunit in AChR localization. We will generate transgenic mice with a beta subunit lacking the phosphorylation site, and then determine whether this mutation impairs the synaptic localization of the AChR. We will also determine whether phosphorylation specifically regulates the anchoring, clustering, and/or stabilization of the receptor in the postsynaptic membrane. (2) To define the molecular mechanism of rapsyn-mediated AChR localization. Here, we will define rapsyn's binding site(s) on the AChR and determine whether binding is regulated by agrin. Moreover, we will investigate how agrin-induced changes in rapsyn/AChR association mediate receptor localization. This work is directly relevant to neuromuscular disorders such as congenital and autoimmune myasthenic syndromes, where there are severe deficiencies in AChR levels at the synapse, leading to impaired synaptic function and debilitating muscle weakness. Our work will help define the mechanisms involved in receptor localization and may suggest strategies to stabilize its localization in patients with myasthenic syndromes or other forms of neuromuscular disease.