Gαs regulates the post-endocytic sorting of G protein-coupled receptors

Rosciglione, Stéphanie; Thériault, Catherine; Boily, Marc-Olivier; Paquette, Marilène; Lavoie, Christine

doi:10.1038/ncomms5556

Cited by 43 publications

(44 citation statements)

References 68 publications

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“…2, A and B). A similar extent of colocalization between EEA1 and the ␦-opioid and CXCR4 receptors was previously reported (32). These data indicate that internalized PAR4 is sorted into early endosomal compartments between 15 and 30 min after agonist stimulation.…”

Section: Internalized Par4 Is Trafficked Through Early Endosomessupporting

confidence: 70%

Protease-activated Receptor-4 Signaling and Trafficking Is Regulated by the Clathrin Adaptor Protein Complex-2 Independent of β-Arrestins

Smith

Coronel

et al. 2016

Journal of Biological Chemistry

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Protease-activated receptor-4 (PAR4) is a G protein-coupled receptor (GPCR) for thrombin and is proteolytically activated, similar to the prototypical PAR1. Due to the irreversible activation of PAR1, receptor trafficking is intimately linked to signal regulation. However, unlike PAR1, the mechanisms that control PAR4 trafficking are not known. Here, we sought to define the mechanisms that control PAR4 trafficking and signaling. In HeLa cells depleted of clathrin by siRNA, activated PAR4 failed to internalize. Consistent with clathrin-mediated endocytosis, expression of a dynamin dominant-negative K44A mutant also blocked activated PAR4 internalization. However, unlike most GPCRs, PAR4 internalization occurred independently of ␤-arrestins and the receptor's C-tail domain. Rather, we discovered a highly conserved tyrosine-based motif in the third intracellular loop of PAR4 and found that the clathrin adaptor protein complex-2 (AP-2) is important for internalization. Depletion of AP-2 inhibited PAR4 internalization induced by agonist. In addition, mutation of the critical residues of the tyrosine-based motif disrupted agonist-induced PAR4 internalization. Using Dami megakaryocytic cells, we confirmed that AP-2 is required for agonist-induced internalization of endogenous PAR4. Moreover, inhibition of activated PAR4 internalization enhanced ERK1/2 signaling, whereas Akt signaling was markedly diminished. These findings indicate that activated PAR4 internalization requires AP-2 and a tyrosine-based motif and occurs independent of ␤-arrestins, unlike most classical GPCRs. Moreover, these findings are the first to show that internalization of activated PAR4 is linked to proper ERK1/2 and Akt activation.Thrombin is a coagulant protease that plays an essential role in hemostasis and thrombosis in response to tissue injury (1).Thrombin is locally generated at sites of vascular injury, where it initiates responses in various cell types including platelets, endothelial, and smooth muscle cells. Thrombin elicits cellular responses through members of the protease-activated receptor (PAR) 5 family of G protein-coupled receptors (GPCRs). Unlike classic GPCRs, which are activated by a diffusible ligand, PARs are activated by the proteolytic cleavage of the receptor N-terminal domain (2, 3). The newly exposed N terminus, termed the "tethered ligand," then binds intramolecularly to the receptor's second extracellular loop, inducing a conformational change that facilitates coupling to heterotrimeric G proteins and downstream signaling. Studies have shown that a six-amino acid synthetic peptide mimic of a PAR's tethered ligand is sufficient to activate the receptor and evoke cellular responses similar to those resulting from proteolytic cleavage of the receptor (4). Because of the irreversible mechanism of PAR1 activation with the generation of a tethered ligand that cannot diffuse away, the mechanisms that regulate PAR1 trafficking are important for maintaining proper signaling and appropriate cellular responses (5).After activa...

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Section: Internalized Par4 Is Trafficked Through Early Endosomessupporting

confidence: 70%

Protease-activated Receptor-4 Signaling and Trafficking Is Regulated by the Clathrin Adaptor Protein Complex-2 Independent of β-Arrestins

Smith

Coronel

et al. 2016

Journal of Biological Chemistry

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“…Furthermore, G α subunits may regulate membrane trafficking through a role in addition to that of signaling. Interaction with growth and differentiation factor-associated serum protein-1 and dysbindin has been suggested to regulate the sorting of GPCRs into multivesicular endosomes via the ESCRT pathway (139). Hence, membrane trafficking appears to be a common regulator of signaling for RTKs, receptor serine/threonine kinases, and GPCRs.…”

Section: G Protein-coupled Receptorsmentioning

confidence: 98%

Spatial and Temporal Regulation of Receptor Tyrosine Kinase Activation and Intracellular Signal Transduction

Bergeron

Guglielmo

Dahan

et al. 2016

Annu. Rev. Biochem.

105

100

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Epidermal growth factor (EGF) and insulin receptor tyrosine kinases (RTKs) exemplify how receptor location is coupled to signal transduction. Extracellular binding of ligands to these RTKs triggers their concentration into vesicles that bud off from the cell surface to generate intracellular signaling endosomes. On the exposed cytosolic surface of these endosomes, RTK autophosphorylation selects the downstream signaling proteins and lipids to effect growth factor and polypeptide hormone action. This selection is followed by the recruitment of protein tyrosine phosphatases that inactivate the RTKs and deliver them by membrane fusion and fission to late endosomes. Coincidentally, proteinases inside the endosome cleave the EGF and insulin ligands. Subsequent inward budding of the endosomal membrane generates multivesicular endosomes. Fusion with lysosomes then results in RTK degradation and downregulation. Through the spatial positioning of RTKs in target cells for EGF and insulin action, the temporal extent of signaling, attenuation, and downregulation is regulated.

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“…First, the GASP machinery is subject to control by conventional signaling proteins such as heterotrimeric G proteins (Rosciglione et al, 2014), suggesting that this machinery may afford additional integration between receptor signaling and trafficking. Second, there is evidence that GASP controls various noncanonical signaling processes, such as through binding to specific transcription factors (Abu-Helo and Simonin, 2010) or regulating autophagy (He et al, 2013a).…”

Section: D-opioid Receptor Traffickingmentioning

confidence: 99%