Molecular Mechanisms of AgRP Signaling

项目摘要 在大脑中，控制代谢功能和进食行为的关键焦点是 黑皮质素受体MC 3R和MC 4 R。这些受体对两种配体有反应， α-黑素细胞刺激激素（α-MSH）和刺豚鼠相关蛋白（AgRP）， 它们以相反的方式起作用以促进负能量平衡和正能量平衡， 分别最近的研究表明，AgRP释放的深刻重要性 代谢和体重稳态的神经元。该计划的目标是 了解AgRP作用的分子基础，从而实现治疗的新策略 与肥胖和代谢性疾病有关的各种疾病。我们实验室的新发现 正在显著重塑我们对a-MSH和AgRP作用的理解。当前 范式假定这些分子的作用是刺激或抑制 cAMP第二信使。然而，使用蛋白质设计，我们证明了AgRP MCR结合核心以外片段的突变对 长期喂养，同时完全保留受体亲和力和cAMP抑制 相对于野生型无变化。此外，新的合作结果发现，AgRP 通过cAMP促进内向整流钾通道的开放 独立的机制，直接依赖于这些外周AgRP的作用 片段本申请的目的1将通过确定AgRP如何 序列和构象驱动钾通道电流。这将受到考验 通过蛋白质设计，核磁共振结构测定，以及与一个新的人类 通过全外显子组分析鉴定的AgRP肥胖相关突变体。目标2将研究如何 syndecan-3是一种带负电荷的膜结合蛋白聚糖， 信号这将通过生物物理实验和 比较研究，将设计的蛋白质给予野生型和 syndecan-3敲除小鼠，随后进行喂养试验和脑组织成像。目标3 将扩大AgRP设计工作，以生产稳定的蛋白质，以测试 蛋白水解抗性促进长期AgRP作用，并作为治疗 癌症恶病质，一种以极度食欲不振和消瘦为特征的消耗性疾病 组织降解。

Project Summary In the brain, key focal points for control of metabolic function and feeding behavior are melanocortin receptors MC3R and MC4R. These receptors respond to two ligands, alpha-melanocyte stimulating hormone (a-MSH) and the agouti-related protein (AgRP), which act in opposite ways to promote negative and positive energy balance, respectively. Recent research demonstrates the profound importance of AgRP releasing neurons in metabolism and body weight homeostasis. The goal of this program is to understand the molecular basis of AgRP action, thus enabling new strategies for treating diverse conditions linked to obesity and metabolic diseases. New findings from our lab are significantly reshaping our understanding of a-MSH and AgRP action. The current paradigm posits that these molecules act to stimulate or suppress production of the cAMP second messenger. However, using protein design, we demonstrated that AgRP mutations in segments outside of the MCR binding core exert a profound influence on long term feeding, while leaving receptor affinity and cAMP suppression completely unchanged relative to wild-type. Moreover, new collaborative results find that AgRP promotes the opening of inward rectifying potassium channels through a cAMP independent mechanism, an effect that is directly dependent on these peripheral AgRP segments. Aim 1 of this application will expand these studies by identifying how AgRP sequence and conformation drive potassium channel currents. This will be tested through protein design, NMR structure determination, as well as with a new human AgRP obesity-linked mutant identified by whole exome analysis. Aim 2 will examine how syndecan-3, a negatively charged, membrane bound proteoglycan, facilitates AgRP signaling. This will be accomplished with biophysical experiments and through comparison studies where designed proteins are administered to wild-type and syndecan-3 knockout mice, followed by feeding trials and brain tissue imaging. Aim 3 will expand on AgRP design efforts to produce stable proteins to test the role of proteolytic resistance in promoting long-term AgRP action, and as leads for treating cancer cachexia, a wasting condition characterized by extreme loss of appetite and lean tissue degradation.