Carprofen br Mechanisms of GPCR internalization Like for oth
Mechanisms of GPCR internalization Like for other types of receptors, prolonged agonist stimulation often leads to GPCR internalization, which can occur via different pathways ∗, , , , . Of these pathways, clathrin-mediated Carprofen (CME) is the best characterized and arguably most relevant one (Fig. 1) ∗, , , , . The first molecular event involved in GPCR internalization is the binding of a family of G protein-coupled receptor kinases (GRKs) to an agonist-occupied receptor, which phosphorylate multiple intracellular serine and threonine residues located in the 3rd intracellular loop or at the C-terminus of the receptor , , , . This is followed by binding of arrestins to the phosphorylated receptor, which plays a major role in both fast signal desensitization and receptor internalization , . On the one hand, arrestins compete with G proteins for binding to the receptor, thus leading to signal desensitization. On the other hand, they promote receptor internalization via interacting with key proteins involved in the assembly of clathrin-coated pits (CCPs) such as the clathrin heavy chain and the clathrin adapter protein AP2 , . This leads to the recruitment of GPCRs into CCPs, which detach form the plasma membrane in a process that requires the small GTPase dynamin . Receptors are then rapidly transferred to early endosomes, from where they can follow either of two main trafficking pathways , . Some GPCRs are sorted out in the endosomal compartment, where they are dephosphorylated, to be then recycled back to the plasma membrane. Others are directed to lysosomes where they are degraded, leading to receptor downregulation , .
Role of receptor internalization in MAPK signaling While rapid desensitization was shown to occur before receptor internalization and be mediated by receptor phosphorylation and β-arrestin recruitment, it soon began to emerge that β-arrestin recruitment and receptor internalization might also exert other functions. In experiments using a dominant-negative dynamin mutant, Daaka et al. showed that receptor internalization is required for efficient ERK activation in response to β2-adrenergic receptor stimulation . Subsequently, it was shown that β-arrestins can bind several components of mitogen-activated protein kinase (MAPK) pathways , , thus promoting G protein-independent MAPK signaling. Since some GPCR are found on early endosomes in complex with β-arrestins, it has been suggested that these events result in endosomal MAPK signaling (Fig. 1) . Intriguingly, the activation of arrestin-bound ERK has been shown to favor cytoplasmic vs. nuclear effects of MAPK activation by preventing ERK translocation to the nucleus , . However, the β-arrestin dependent activation of MAPKs can also occur while the receptors are still located on the plasma membrane. Thus, it remains to be clarified what is the relative contribution of cell surface vs. endosomal MAPK signaling. Moreover, some GPCRs that are poorly internalized are nevertheless able to efficiently induce MAPK signaling. This can be at least partially explained by the existence of other mechanisms leading to MAPK activation. Yet another possible explanation for these findings comes from a recent study on the β1-adrenergic receptor − which internalizes poorly upon agonist stimulation − indicating that receptor activation can lead to recruitment of β-arrestin to CCPs and MAPK signaling from CCPs in the absence of receptors .
New paradigm of GPCR signaling from intracellular compartments Although classical, G protein-dependent signaling has long been believed to be restricted to the plasma membrane, studies performed in the last ten years have provided strong evidence that internalized GPCRs can continue signaling on intracellular membranes (Fig. 1). A first indication came from experiments on the Ste2 receptor, which is implicated in pheromone signaling in yeast . Subsequently, our group and that of Jean-Pierre Vilardaga independently showed that the TSH and PTH receptors induce a persistent phase of cAMP production after internalization, which could be prevented by interfering with CME ∗, ∗. Signaling by internalized TSH receptors was shown to differ from the one occurring at the plasma membrane in that it was required for efficient phosphorylation of the vasodilator-stimulated phosphoprotein (VASP) and actin depolymerization in response to TSH, which is involved in thyroglobulin reuptake and, thus, in thyroid hormone release . In the case of the PTH receptor, signaling was shown to be turned off by retromer − which mediates retrograde trafficking from endosomes to the trans-Golgi network − and endosomal acidification , . These findings challenged the classical model of GPCR signaling by indicating that G protein signaling can also occur on intracellular membranes. They also pointed to early endosomes, in the case of the PTH receptor, and the Golgi/trans-Golgi network, in the case of the TSH receptor, as likely sites of intracellular GPCR signaling (Fig. 1).