Endocytic Trafficking of G-Protein-Coupled Receptors as a Novel Regulator of Inflammation

这个项目将基于我们在受体药理学理解方面的最新进展，调查和发展对实现炎症消退所需过程的基本理解。炎症是受伤或感染后帮助组织恢复动态平衡的自然过程。然而，当炎症变得不受控制时，它可能会危害健康。例如，脓毒症是指身体的正常炎症反应非常严重，无法消除，并对身体器官造成损害。这是重症监护病房中最常见的死亡原因，在英国每年约有3.5万人死亡。因此，了解炎症在健康和特定疾病中如何自然调节和分解的分子机制对于健康和开发更有效的治疗方法至关重要[1]。炎症的一个关键调节因子是G蛋白偶联受体家族；甲酰肽受体(FPR)，特别是FPR2，被认为负责分解。这些受体调节多种反应，从白细胞的激活和迁移，到细胞的凋亡和吞噬。然而，GPCRs究竟是如何参与这些过程的分子机制以及它们在脓毒症治疗中的治疗潜力仍不清楚。令人兴奋的是，我们最近的工作发现了新的见解[2]，表明FPR2的内吞运输在炎症的消退中起着关键作用。胞内转运控制可用的功能性受体的数量，允许细胞在响应外部刺激时增加或减少信号幅度[3]。我们发现，中断FPR2的运输会导致细胞凋亡增加；这是解决问题的关键要求。这一发现为了解炎症和治疗的发展提供了一个新的分子靶点。以这些发现为基础，该项目将由3部分组成：1)研究FPR2的分子药理学和细胞内转运2)开发FPR2内吞的多肽抑制剂3)利用基因编辑技术分析FPR2功能改变的功能后果，以产生在体内显示效果的新型小鼠模型这个专业的学生将利用并提供各种现代科学技术方面的培训，特别是我们将使用尖端成像技术(超分辨率、实时共聚焦显微镜、荧光生物传感器)来研究和确定甲酰肽受体经历细胞内转运的机制，以及这如何调节信号、细胞因子的产生、迁移和吞噬来确定炎症反应的强度。我们将根据甲酰肽受体的氨基酸序列开发肽抑制剂，该受体将在室内合成，用于扰乱运输。最后，新的基因编辑技术将被用来产生表达运输受损受体的小鼠系，以在体内验证这一过程在炎症动物模型中的重要性。这个跨学科和协作的项目致力于BBSRC的世界级生物科学支撑战略，承诺识别调控炎症的基本机制，验证新的靶点，并可能为未来治疗炎症的开发确定新的小分子抑制剂1)Ortega-Gómez A，Perretti M，Soehnlein O.EMBO Mol Med。2013年5月；5(5)：661-74.2)Thompson D，McArthur S，Hislop JN，Flow RJ，Perretti M(2014)J Biol Chem。26；289(52)：36166-78.3Hislop JN，von Zastrow M.2011；12：137-48。

This project will investigate and develop the basic understanding of the processes required to bring about the resolution of inflammation based on our recent novel advances in the understanding of receptor pharmacology. Inflammation is a natural process that helps restore tissue homeostasis after injury or infection. However, when inflammation becomes uncontrolled it can be hazardous to health. For example, sepsis is when the body's normal inflammatory response is so severe that it fails to resolve and causes injury to the body's organs. This is the most common cause of death within intensive care, accounting for ~35,000 deaths in the UK each year. Understanding the molecular mechanisms of how inflammation is regulated and resolved naturally in health and in specific disorders is therefore critical to well-being and for the development of more effective therapies [1]. One critical regulator of inflammation is the family of G-protein-coupled receptors; the formyl peptide receptors (FPR), and in particular, FPR2, thought to be responsible for resolution. These receptors modulate a variety of responses, from leukocyte activation and migration, to apoptosis and phagocytosis. However, the molecular mechanisms underlying exactly how GPCRs are involved in these processes and thus, their therapeutic potential in the treatment of sepsis remains unclear. Excitingly, our recent work has uncovered new insight [2], indicating that endocytic trafficking of FPR2 plays a critical role in resolution of inflammation. Endocytic trafficking controls the number of functional receptors available, allowing the cell to increase or decrease signalling amplitude in response external stimuli [3]. We have discovered that disruption of FPR2 trafficking leads to increased cellular apoptosis; a key requirement for resolution. This finding provides a novel molecular target for understanding of inflammation and the development of therapeutics. Building on these findings, the project will consist of 3 parts:1) Investigate the molecular pharmacology and endocytic trafficking of FPRs2) Develop peptide inhibitors of endocytic trafficking3) Analyse the functional consequences of altered FPR2 function by the use of gene editing to generate a novel mouse model to show effects in vivo This studentship will employ and provide training in a diverse array of modern scientific techniques, specifically we will use cutting edge imaging techniques (super-resolution, real-time confocal microscopy, fluorescent biosensors) to investigate and identify the mechanism by which the formyl peptide receptor undergoes endocytic trafficking and how this regulates signalling, cytokine production, migration and phagocytosis to determine the strength of the inflammatory response. We will develop peptide inhibitors based on the amino acid sequence of the formyl peptide receptor, which will be synthesized in house, and used to perturb trafficking. Finally new gene editing techniques will be used to generate a mouse line expressing trafficking impaired receptors to validate, in vivo, the importance of this process in animal models of inflammation. This interdisciplinary and collaborative project addresses the BBSRC strategy of world-class underpinning bioscience by promising to identify fundamental mechanisms regulating inflammation and validate new targets and potentially identify new small molecule inhibitors for the future development for the treatment of inflammation.1) Ortega-Gómez A, Perretti M, Soehnlein O. EMBO Mol Med. 2013 May;5(5):661-74.2) Thompson D, McArthur S, Hislop JN, Flower RJ, Perretti M (2014) J Biol Chem. 26;289(52):36166-78.3) Hislop JN, von Zastrow M. Traffic. 2011;12:137-48.