Introduction
G protein-coupled receptors (GPCRs) are encoded by approximately 800 different genes and form the largest family of membrane proteins in the human genome. GPCRs mediate the effects of a variety of extracellular cues, from neurotransmitters to photons, and consequently, are fundamental regulators of physiological homeostasis (Calebiro and Godbole, 2018). It is therefore unsurprising that around one third of current pharmaceuticals target this receptor family (Calebiro and Godbole, 2018). Despite this, several important aspects of GPCR signalling remain insufficiently understood. In the classical view of GPCR signalling, ligand-activated GPCRs signal at the plasma membrane via heterotrimeric G proteins, often rapidly desensitise, and undergo arrestin-mediated internalisation (Calebiro and Godbole, 2018). Signalling competence can then be restored by GPCR re-sensitisation and recycling back to the plasma membrane (Calebiro and Godbole, 2018). However, evidence gathered over the last decade indicates that select GPCRs can also signal through heterotrimeric G proteins at distinct intracellular sites and that the resulting signals may be important for physiological functions (Irannejad et al., 2013, Eichel and von Zastrow, 2018, Calebiro et al., 2009, Godbole et al., 2017, Yarwood et al., 2017). Evidence of intracellular GPCR signalling has been largely obtained through the application of genetically encoded fluorescence/bioluminescence resonance energy transfer (FRET/BRET)-based sensors that measure dynamic protein–protein and intramolecular interactions in real-time and in live cells. Such methodologies have a much higher temporal resolution than endpoint biochemical assays and have substantially improved our understanding of GPCR signalling. Above all, they have revealed that GPCR signalling is not limited to the plasma membrane, but instead is a highly regulated event that works in combination with GPCR trafficking to enhance the specificity of signalling in response to distinct physiological cues.