Roles for receptor activity modifying proteins with non-traditional binding partners

The communication between the cells in the body influences how our tissues are constructed and how they function. Cells signal to each other in a variety of ways, but the vast majority of signalling involves receptors / proteins that can recognise specific hormone or growth factor signalling molecules. In recent years it has been discovered that a growing number of hormone molecules do not signal to a receptor only by switching it on or off but instead, the action of the receptor on binding its hormone is altered according to the presence of a so-called receptor activity modifying protein or RAMP. Different RAMPs can either change the presence of a receptor on the cell surface, alter the hormone that can bind to a receptor or change the consequences of hormone binding. We aim to determine the specific consequences on cell signalling induced by different combinations of receptor, hormone and RAMP and to identify novel receptors that are regulated by RAMPs. We will use a combination of techniques that are currently available in Sheffield and the results of our work will provide new insights into how hormones function in the body with implications for understanding of health and disease.

Research on Receptor Activity Modifying Proteins (RAMPs) has focused on their ability to modulate ligand specificity of the calcitonin (CT) family receptors. Co-expression of RAMPs with the CT receptor (CTR) or the CTR-like receptor (CRLR) gives rise to 7 different receptors with specific properties. RAMPs therefore control actions of CT, calcitonin gene related peptide, adrenomedullin and amylin, hormones with wide activity profiles. Recently, further mechanisms by which RAMPs influence function have been discovered. They interact with several additional G protein-coupled receptors (GPCRs). However, those studies have focused upon pharmacological properties of receptor/RAMP/ligand combinations so the consequences of these novel interactions are unclear. As RAMPs may regulate receptor trafficking, cell surface expression, receptor/ligand interactions and internalisation and processing, it is likely they have important roles beyond regulation of ligand specificity. This application is to determine consequences on signalling and function of RAMP interactions with receptors outside the CTR family. We will use radioligand binding assays and g-protein scintillation proximity analysis to determine the pharmacological characteristics of the known non-CTR family/RAMP combinations and measure the effects of RAMPs on their trafficking internalisation and recycling. To identify novel GPCRs interacting with RAMPs, we will use FRET analysis to measure directly the physical interactions of RAMPs with a minimum of 30 other GPCRs. Where we identify interacting GPCRs, we will characterise their pharmacology as before. Finally we will investigate the role of cell background on interactions by studying 1 known and 1 novel RAMP partner in 2 different cell lines. These studies will advance understanding of GPCR signalling and RAMP function. This is an industrial partnership award with Mantelum, a company whose focus is therapeutic applications of manipulation of GPCR signalling.