Dynamic tuneable G protein-coupled receptor monomer-dimer populations

Patricia M Dijkman,Anthony Watts,Juan C Munoz-Garcia,Mark I Wallace,Oliver K Castell,Chris De Graaf,Alan D Goddard

doi:10.1038/s41467-018-03727-6

Abstract

G protein-coupled receptors (GPCRs) are the largest class of membrane receptors, playing a key role in the regulation of processes as varied as neurotransmission and immune response. Evidence for GPCR oligomerisation has been accumulating that challenges the idea that GPCRs function solely as monomeric receptors; however, GPCR oligomerisation remains controversial primarily due to the difficulties in comparing evidence from very different types of structural and dynamic data. Using a combination of single-molecule and ensemble FRET, double electron–electron resonance spectroscopy, and simulations, we show that dimerisation of the GPCR neurotensin receptor 1 is regulated by receptor density and is dynamically tuneable over the physiological range. We propose a “rolling dimer” interface model in which multiple dimer conformations co-exist and interconvert. These findings unite previous seemingly conflicting observations, provide a compelling mechanism for regulating receptor signalling, and act as a guide for future physiological studies.

Highlights

G protein-coupled receptors (GPCRs) are the largest class of membrane receptors, playing a key role in the regulation of processes as varied as neurotransmission and immune response
To gain insight into dynamics of neurotensin receptor 1 (NTS1) dimerisation, single-molecule Förster resonance energy transfer (FRET) experiments were conducted on receptors fluorescently labelled at the intracellular end of TM4 (T186C4.42, where the superscript refers to Ballesteros-Weinstein numbering for GPCRs, see Supplementary Fig. 1), which has previously been proposed to sit at the NTS1 dimerisation interface[24]
In agreement with recent observations for other GPCRs23, the data presented here support the notion of a dynamic transient NTS1 dimer with a half-life (t1/2 = 1.2 s) consistent with previous reports on other receptors (~0.1–5 s)[15,16,17]

Summary

Introduction

G protein-coupled receptors (GPCRs) are the largest class of membrane receptors, playing a key role in the regulation of processes as varied as neurotransmission and immune response. We propose a “rolling dimer” interface model in which multiple dimer conformations co-exist and interconvert These findings unite previous seemingly conflicting observations, provide a compelling mechanism for regulating receptor signalling, and act as a guide for future physiological studies. Receptor oligomerisation may be required to traffic receptors to the plasma membrane[6], and regulate receptor internalisation[7], but may affect ligand binding[8], and G protein activation[9] suggesting some role for dimerisation in biased signalling[10]. Detailed morphological data for class A GPCR dimers remain largely elusive and are often conflicting[2], with every receptor transmembrane segment having been implicated in dimerisation and even studies on the same receptor proposing different interfaces (Supplementary Table 1). In solution at low detergent concentrations, NTS1 dimerisation occurs in a concentrationdependent manner and dimers show ligand-binding cooperativity, but catalyse G protein nucleotide exchange with lower affinity than monomers[23]

Methods

Results

Conclusion