Trends in Pharmacological Sciences
ReviewSignaling by internalized G-protein-coupled receptors
Introduction
Cells respond to environmental cues and communicate with each other through the activation of receptors located on the cell surface. G-protein-coupled receptors (GPCRs) form the largest family of such receptors. They mediate effects of neurotransmitters, hormones, ions, odorants and light. Their signals are essentially mediated via the activation of heterotrimeric G proteins and their effectors (e.g. adenylyl cyclase, phospholipase C, potassium and calcium channels). Because of their involvement in a large number of physiological and pathological processes, GPCRs have been subject to intensive investigation and represent major targets for current pharmacological intervention [1].
Similar to other classes of receptors, prolonged stimulation of GPCRs often leads to their internalization into endosomes, presumably via more than one internalization pathway 1, 2, 3. Although originally considered as a major mechanism of signal desensitization, the results of several studies performed over the past 15 years suggest other functions for receptor internalization [4], most notably receptor resensitization 1, 5, 6, 7 and signaling to the mitogen-activated protein kinase (MAPK) cascade [8]. In addition, proteolytic fragments of internalized Frizzled GPCRs have been shown to translocate to the nucleus where they might activate gene transcription [9]. In spite of these observations, it is generally believed that GPCR signaling to classical G-protein-dependent pathways, such as the Gs-dependent activation of adenylyl cyclase, occurs exclusively at the cell surface. This view has been challenged by three recent studies, which suggest that a previously unrecognized type of persistent GPCR signaling to cAMP can occur after ligand–receptor internalization 10, 11, 12. Here, we will review the evidence for the existence of GPCR–cAMP signaling pathways on endosomes, and their possible pathophysiological and pharmacological implications.
Section snippets
GPCR internalization and desensitization
The molecular mechanisms of clathrin-dependent GPCR internalization have been extensively investigated (reviewed in Refs. 1, 2, 3, 13, 14). The trigger for receptor internalization is the conformational change induced by agonist binding, which, apart from initiating G-protein-dependent signaling, transforms receptors into substrates of the G-protein-coupled receptor kinases (GRKs). As a result, the ligand-occupied receptors become phosphorylated at cytosolic Ser/Thr residues. Ligand-occupied
Lessons from receptor tyrosine kinase signaling
Endosomes possess several characteristics that, at least in principle, make them ideal intracellular signaling platforms; among others, they have a high surface-to-volume ratio, which would favor ligand–receptor interactions, they have a unique lipid (high phosphatidylinositol 3-phosphate content) and protein composition permitting selective recruitment of signaling components, they move centripetally, thus potentially allowing the dissemination of short-range signals to compartments distant
‘Non-classical’ GPCR signaling at intracellular membranes
In analogy with the findings on RTKs, GPCR signaling to MAPKs has also been proposed to occur at endosomes. The first evidence came from experiments where a dynamin dominant negative mutant was shown to inhibit ERK activation stimulated by the β2-adrenergic receptor [8]. Subsequently, it was shown that certain GPCRs remain associated with β-arrestins in endosomes [31] and that β-arrestins can bind to several components of the MAPK pathways 32, 33. In light of these findings, it has been
Persistent GPCR signaling to cAMP at endosomes
Very recently, the emerging concept of ‘non-classical’ endosomal GPCR signaling has been complemented by findings also suggesting classical, G-protein-dependent signaling of intracellular GPCRs. Data from three groups provide strong evidence for persistent signaling to adenylyl cyclase by internalized GPCRs.
To monitor GPCR–cAMP signaling directly in living cells, we have recently developed a transgenic mouse [10] with ubiquitous expression of a fluorescent reporter for cAMP [42]. The thyroid
Functional consequences of GPCR–cAMP signaling on endosomes
Although these new findings clearly support the existence of GPCR–cAMP signaling pathways on endosomes, our understanding of their functional relevance is still limited. What appears clear is that, differently from what is occurring at the cell surface, GPCR–cAMP signaling on endosomes is persistent. This phenomenon might be particularly relevant in vivo, where the access to ligands is often limited and can vary over time. This is the case for several hormones, including TSH [46], that are
Concluding remarks
Based on these recent results we propose a new model of GPCR signaling (Figure 4). Several GPCRs are internalized together with their ligands (and perhaps with G proteins and adenylyl cyclases) in endosomes or other intracellular compartments. Here, at least some of them, namely TSHR, PTHR and S1P1 receptors, find the machinery required for cAMP production. As both ligands and receptors remain confined in endosomes for some time, this mechanism permits prolonged signaling even after removal of
Acknowledgement
Research by the authors referred to in this publication is supported by grants from the European Research Council (Advanced Grant TOPAS) and the Deutsche Forschungsgemeinschaft (SFB487).
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