Spatio-temporal assessment of β-arrestin-centred “signalosomes” – the impact of effector binding on the functional selectivity of protease-activated receptor 2.

细胞是生命最基本的实体。为了与周围环境沟通，细胞最外层的细胞膜上有许多不同的受体。在人体内，细胞表面受体控制着大多数生理过程，不仅能引起对激素和神经递质的反应，还能感知气味和光线。然而，受体激活后在细胞内展开的确切分子过程的很大一部分是未知的。根据最近的数据，我们可以假设活性受体周围的区域迅速充满了酶、支持适配器和效应器蛋白来诱导细胞功能。然后，这些具有信号能力的复合体被内化并在整个细胞内运输。然而，我们不知道这些复合体的确切蛋白质组成，它们是否会根据它们的亚细胞定位而发生变化，以及这对细胞信号传递有什么影响。在这里，该项目旨在研究细胞表面受体及其效应器蛋白在初始激活后如何聚集和表现，同时监测整个运输过程中的细胞信号机制。这项研究聚焦于蛋白酶激活受体2(PAR2)，作为不同肠道疾病中疼痛的重要中介，将探讨细胞信号传递的基本机制及其与疼痛感知的关系。利用诺丁汉大学和膜蛋白和受体中心(COMPARE)提供的尖端成像设备，可以跟踪受体激活后细胞内发生的事情。创新技术，如NanoBRET信号探针和单分子荧光显微镜，将被用来促进我们目前对受体激活后与其他信号伙伴联系在一起的“如何”、“何时何地”和“为什么”的理解。此外，其目的是提供一个具有高空间和动力学分辨率的细胞内信号“图谱”。预期的结果将增加我们对细胞信号机制的总体理解，同时可能揭示缓解疼痛的新的药理靶点。

Cells are the most fundamental entities of life. To communicate with their surroundings, cells possess a multitude of different receptors on their most outer lining, the plasma membrane. In the human body, cell surface receptors control most physiological processes and not only elicit responses to hormones and neurotransmitters, but also enable the sensing of odours and light. However, large parts of the exact molecular processes which unfold inside a cell after receptor activation are unknown. From recent data, we can assume that the area surrounding active receptors rapidly populates with enzymes, supporting adaptors and effector proteins to induce a cellular function. These signalling competent complexes are then internalised and transported throughout the cell. However, we do not know of the exact protein composition of these complexes, whether they are subject to change, depending on their subcellular localisation and which consequences this has for cellular signalling. Here, the project aims to investigate how cell surface receptors and their effector proteins cluster and behave after an initial activation, while monitoring the cellular signalling machinery throughout the entire trafficking process. Focusing on the protease-activated receptor 2 (PAR2), as an important mediator of pain in different bowel diseases, this study will examine fundamental mechanisms of cellular signalling and how this relates to pain perception. Taking advantage of the cutting-edge imaging facilities provided by the University of Nottingham and the Centre of Membrane Proteins and Receptors (COMPARE), it is possible to track what happens inside a cell after receptor activation. Innovative technologies, such as NanoBRET signalling probes and single-molecule fluorescence microscopy will be used to advance our current understanding of “how”, “when and where” and “why” receptors associate with other signalling partners after their activation. Moreover, the aim is to provide an intracellular signalling “map” with high spatial and kinetic resolution. The expected results will add to our general understanding of cellular signalling mechanisms, while possibly revealing new pharmacological targets for pain relief.