Investigating the requirement of MRAP2 for ghrelin function

项目总结 Ghrelin是一种循环中的食欲激素，主要由胃在禁食期间分泌，以发出信号 大脑的饥饿感。Ghrelin通过作用于其受体生长来调节食物摄入量和能量消耗 激素促分泌素受体1a(GHSR1a)。Ghrelin对食物摄入量和能量平衡的影响 是通过激活AGRP/NPY神经元以及Ghrelin的整体或中心缺失而集中介导的 受体可防止饮食引起的肥胖。出于这个原因，GHSR1a是一个很有希望的治疗高血压的靶点 肥胖症和Ghrelin受体的拮抗剂已经为此目的而开发，然而，它们的混杂 或者较低的生物利用度使其无法在临床上使用。具有改进的药学作用的新化合物- 化学性质和更好的体内疗效很可能通过使用更相关的药物发现来确定 战略。改进筛查战略将需要更好地了解GHSR1a法规。我们 最近发现Ghrelin的促食欲素效应在黑素皮质素受体 删除辅助蛋白2(MRAP2)。我们的初步结果表明，MRAP2与 GHSR1a，并强烈增强Ghrelin刺激的受体下游信号。此外，我们还展示了 MRAP2在Ghrelin反应性AGRP/NPY神经元中表达。与这些发现一致的是，我们表明 来自MRAP2 KO小鼠的AGRP神经元不能对饥饿做出反应。建议进行的实验 该项目将利用几种动物模型来实现对MRAP2的靶向调控 Ghrelin信号在特定神经元中的表达和测定。本提案的目标是1)确定 MRAP2在促进Ghrelin信号转导和AGRP神经元饥饿反应中的作用 MRAP2增强GHSR1a信号效应的机制。目标1中的研究将测试 AGRP神经元MRAP2表达调控改变饥饿感觉和Ghrelin作用的假说 在大脑里。Aim 2中的研究将检验MRAP2增强GHSR1a信号的假设，两者 体内和体外，通过干扰受体的脱敏。这项研究具有重要意义，因为 成功完成将提供有关MRAP2的作用和作用机制的基本信息 由于AgRP神经元与Ghrelin功能有关，因此加深了我们对AGRP神经元调控和 下丘脑控制能量动态平衡。这项研究具有创新性，因为完成这项研究将产生新的 对辅助蛋白调节G蛋白偶联受体的认识及鉴定MRAP2是一种 AGRP神经元中的新型能量传感器。我们还将使用在我们的 实验室以组织特有的方式准确地检测和遗传调节MRAP2的表达。 最终，该项目的完成将为监管GHSR1a提供关键的见解，确定 该受体的必需辅助蛋白，并使针对Ghrelin的药物发现策略的设计成为可能 与之前的尝试相比，这将发出更具生理相关性、更有可能成功的信号。

PROJECT SUMMARY Ghrelin is a circulating orexigenic hormone mainly secreted by the stomach during fasting periods to signal hunger to the brain. Ghrelin regulates food intake and energy expenditure by acting at its receptor the Growth Hormone Secretagogue Receptor 1a (GHSR1a). The effect of ghrelin on food intake and energy homeostasis are mediated centrally through the activation of AGRP/NPY neurons, and global or central deletion of the ghrelin receptor protects from diet-induced obesity. For this reason, GHSR1a is a promising target for the treatment of obesity and antagonists of the ghrelin receptor have been developed for this purpose, however, their promiscuity or low bioavailabilty precludes them from being used in the clinic. New compounds with improved pharmaco- chemical properties and better in-vivo efficacy could likely be identified by using more relevant drug discovery strategies. Improvement of screening strategies will require a better understanding of GHSR1a regulation. We have recently discovered that the orexigenic effect of ghrelin is lost in mice in which the Melanocortin Receptor Accessory Protein 2 (MRAP2) was deleted. Our preliminary results demonstrate that MRAP2 interacts with GHSR1a and strongly potentiates ghrelin-stimulated signaling downstream of the receptor. Additionally, we show that MRAP2 is expressed in the ghrelin responsive AGRP/NPY neurons. Consistent with those findings, we show that AGRP neurons from MRAP2 KO mice fail to activate in response to starvation. Experiments proposed in this project will take advantage of several animal models that allow the targeted modulation of MRAP2 expression and the measurement of ghrelin signaling in specific neurons. The goal of this proposal is to 1) Identify the role of MRAP2 in promoting ghrelin signaling and starvation response of AGRP neurons; 2) Identify the mechanisms through which MRAP2 enhances to efficacy of GHSR1a signaling. The studies in aim 1 will test the hypothesis that the modulation of MRAP2 expression in AGRP neurons alters hunger sensing and ghrelin actions in the brain. The studies in aim 2 will test the hypothesis that MRAP2 potentiates GHSR1a signaling, both in-vivo and in-vitro, by interfering with the desensitization of the receptor. This research is significant because successful completion will provide fundamental information on the role and mechanism of action of MRAP2 in AGRP neurons as it pertains to ghrelin functions, advance our understanding of AGRP neurons regulation and hypothalamic control of energy homeostasis. This research is innovative because completion will generate novel knowledge on the regulation of G-Protein Coupled receptors by accessory proteins and identify MRAP2 as a novel energy sensor in AGRP neurons. We will also be using innovative mouse model generated in our laboratory to accurately detect and genetically modulate the expression of MRAP2 in a tissue specific manner. Ultimately, the completion of this project will provide critical insights in the regulation of GHSR1a, identify an obligatory accessory protein of this receptor and enable the design of drug discovery strategies targeting ghrelin signaling that will be far more physiologically relevant and more likely to succeed than previous attempts.