Regulation of NMDAR-Mediated Synaptic Signaling

项目摘要/摘要 我们建议研究离子亲性谷氨酸受体(IGluRs)的NMDA亚型(NMDAR)及其 NRAP-1是第一个被发现的NMDAR特异性辅助蛋白，我们最近在一个 线虫NMDAR介导行为修饰物的遗传筛选。NMDAR在进化上 它们在突触可塑性，即长时程增强(LTP)中的作用是保守的和众所周知的；它们的重要性 学习和记忆的细胞模型；以及它们直接或间接参与许多神经学和 精神障碍。尽管NRAP-1改变了突触后NMDAR的功能，但我们发现它是 由突触前谷氨酸能神经元释放。这一发现提供了我们在概念上的重大进步 对NMDAR介导的突触信号调节的理解，以及对控制 对于突触强度和某些涉及NMDAR的临床疾病。在初步实验中，我们 成功地获得了重组生产的NRAP-1的晶体，并测定了其晶体结构为1.9 解决方案。阐明NRAP-1的结构为了解NRAP-1如何 修改NMDAR函数。通过对脊椎动物和线虫NMDAR的研究，我们还展示了 NMDAR氨基末端结构域(ATD)在受体门控和 NRAP-1的作用机制。我们现在计划在这项基础性工作的基础上再接再厉，询问NRAP-1如何 调节NMDAR功能的功能，即NRAP-1和NMDAR之间的相互作用，以及 这些相互作用如何改变受体的动力学？与NMDAR的过度表达相反，我们发现 NRAP-1在体内的过表达显著增加了NMDAR介导的电流和行为。这 对控制突触可塑性有重要意义。此外，我们还发现NRAP-1积极地 沿着神经突起运输。总之，这些发现表明，调节NRAP-1的分泌可能 是一种用于调节突触强度的活性依赖变化的机制。因此，我们将解决 NRAP-1转运和分泌的分子要求。我们提议的研究的相关性 是因为NMDAR介导的信号传导障碍与突触病变有关 神经退行性疾病以及精神分裂症和抑郁症等精神健康疾病。 突触分子在进化上是保守的，我们对调控机制的理解 突触信号从基于遗传学的无脊椎动物研究中受益匪浅，如果蝇和 线虫。值得注意的是，NMDAR和NRAP-1类蛋白似乎是共同进化的，这表明脊椎动物 NMDARs可能受辅助蛋白的调节。因此，我们预计我们计划中的研究将有所帮助。 为以蛋白质-蛋白质为中心的NMDARs的新机制理解提供了一个框架 互动。

PROJECT SUMMARY/ABSTRACT We propose to study the NMDA-subtype (NMDARs) of ionotropic glutamate receptors (iGluRs) and their regulation by NRAP-1, the first identified NMDAR-specific auxiliary protein, which we recently discovered in a genetic screen for modifiers of NMDAR-mediated behavior in C. elegans. NMDARs are evolutionarily conserved and well known for their role in synaptic plasticity, i.e., long-term potentiation (LTP); their importance for cellular models of learning and memory; and their direct or indirect involvement in many neurological and psychiatric disorders. Although NRAP-1 modifies the function of postsynaptic NMDARs, we showed that it was released by presynaptic glutamatergic neurons. This discovery provided a major conceptual advance in our understanding of the regulation of NMDAR-mediated synaptic signaling, with implications for both the control of synaptic strength and for certain clinical disorders involving NMDARs. In preliminary experiments, we successfully obtained crystals of recombinantly produced NRAP-1 and determined the crystal structure at 1.9 Å resolution. Elucidating the structure of NRAP-1 has provided important new insight into how NRAP-1 modifies NMDAR function. By studying vertebrate and C. elegans NMDARs, we have also demonstrated a fundamental importance for the NMDAR amino-terminal domain (ATD) with respect to both receptor gating and to the mechanism of action of NRAP-1. We now plan to build on this foundational work and ask how NRAP-1 functions to modulate NMDAR function, i.e., what are the interactions between NRAP-1 and NMDARs, and how do these interactions change receptor kinetics? In contrast to overexpression of NMDARs, we discovered that overexpression of NRAP-1 in vivo significantly increased NMDAR-mediated currents and behavior. This has important implications for the control of synaptic plasticity. Furthermore, we found that NRAP-1 is actively transported along neural processes. Together, these findings suggest that modulating NRAP-1 secretion might be a mechanism used to regulate activity dependent changes in synaptic strength. Therefore, we will address the molecular requirements for the transport and secretion of NRAP-1. The relevance of our proposed studies is high because disorders of NMDAR-mediated signaling are implicated in synaptopathies associated with neurodegenerative disorders as well as for mental health illnesses such as schizophrenia and depression. Synaptic molecules are evolutionarily conserved, and our understanding of the mechanisms that regulate synaptic signaling has greatly benefited from genetics-based studies in invertebrates such as Drosophila and C. elegans. Notably, NMDARs and NRAP-1-like proteins appear to have co-evolved suggesting that vertebrate NMDARs are likely regulated by auxiliary proteins. We therefore anticipate that our planned studies will help provide a framework for a new mechanistic understanding of NMDARs centered on protein-protein interactions.