“…Furthermore, our findings that AMD3100 and anti-CXCR4 12G5 did not prevent β-arrestin 2 recruitment and internalization of CXCR4 upon PE stimulation indicate that this receptor cross-talk can occur when CXCR4 is in its ligand-free and in an antagonist-bound configuration. This assumption is supported by the recent observation that chemically-induced fusion of CXCR4 to β-arrestin 1/2 leads to internalization of the receptor, independent of agonist binding to and G protein coupling through CXCR4 21 . Figure 7 α 1 -AR regulate internalization of CXCR4 in human vascular smooth muscle cells and CXCR4-mediated chemotaxis.…”
Recently, we reported that chemokine (C-X-C motif) receptor (CXCR)4 and atypical chemokine receptor 3 regulate α1-adrenergic receptors (α1-AR) through the formation of hetero-oligomeric complexes. Whether α1-ARs also regulate chemokine receptor function within such heteromeric receptor complexes is unknown. We observed that activation of α1b-AR within the α1b-AR:CXCR4 heteromeric complex leads to cross-recruitment of β-arrestin2 to CXCR4, which could not be inhibited with AMD3100. Activation of CXCR4 did not cross-recruit β-arrestin2 to α1b-AR. A peptide analogue of transmembrane domain 2 of CXCR4 interfered with α1b-AR:CXCR4 heteromerization and inhibited α1b-AR-mediated β-arrestin2 cross-recruitment. Phenylephrine (PE) induced internalization of CXCR4 in HEK293 cells co-expressing CXCR4 and α1b-AR and of endogenous CXCR4 in human vascular smooth muscle cells (hVSMC). The latter was detectable despite blockade of CXCR4 with the neutralizing antibody 12G5. hVSMC migrated towards CXCL12 and PE, but not towards a combination of CXCL12 and PE. PE inhibited CXCL12-induced chemotaxis of hVSMC (IC50: 77 ± 30 nM). Phentolamine cross-inhibited CXCL12-induced chemotaxis of hVSMC, whereas AMD3100 did not cross-inhibit PE-induced chemotaxis. These data provide evidence for asymmetrical cross-regulation of CXCR4 by α1-adrenergic receptors within the heteromeric receptor complex. Our findings provide mechanistic insights into the function of α1-AR:CXCR4 heteromers and suggest alternative approaches to modulate CXCR4 in disease conditions.
“…Furthermore, our findings that AMD3100 and anti-CXCR4 12G5 did not prevent β-arrestin 2 recruitment and internalization of CXCR4 upon PE stimulation indicate that this receptor cross-talk can occur when CXCR4 is in its ligand-free and in an antagonist-bound configuration. This assumption is supported by the recent observation that chemically-induced fusion of CXCR4 to β-arrestin 1/2 leads to internalization of the receptor, independent of agonist binding to and G protein coupling through CXCR4 21 . Figure 7 α 1 -AR regulate internalization of CXCR4 in human vascular smooth muscle cells and CXCR4-mediated chemotaxis.…”
Recently, we reported that chemokine (C-X-C motif) receptor (CXCR)4 and atypical chemokine receptor 3 regulate α1-adrenergic receptors (α1-AR) through the formation of hetero-oligomeric complexes. Whether α1-ARs also regulate chemokine receptor function within such heteromeric receptor complexes is unknown. We observed that activation of α1b-AR within the α1b-AR:CXCR4 heteromeric complex leads to cross-recruitment of β-arrestin2 to CXCR4, which could not be inhibited with AMD3100. Activation of CXCR4 did not cross-recruit β-arrestin2 to α1b-AR. A peptide analogue of transmembrane domain 2 of CXCR4 interfered with α1b-AR:CXCR4 heteromerization and inhibited α1b-AR-mediated β-arrestin2 cross-recruitment. Phenylephrine (PE) induced internalization of CXCR4 in HEK293 cells co-expressing CXCR4 and α1b-AR and of endogenous CXCR4 in human vascular smooth muscle cells (hVSMC). The latter was detectable despite blockade of CXCR4 with the neutralizing antibody 12G5. hVSMC migrated towards CXCL12 and PE, but not towards a combination of CXCL12 and PE. PE inhibited CXCL12-induced chemotaxis of hVSMC (IC50: 77 ± 30 nM). Phentolamine cross-inhibited CXCL12-induced chemotaxis of hVSMC, whereas AMD3100 did not cross-inhibit PE-induced chemotaxis. These data provide evidence for asymmetrical cross-regulation of CXCR4 by α1-adrenergic receptors within the heteromeric receptor complex. Our findings provide mechanistic insights into the function of α1-AR:CXCR4 heteromers and suggest alternative approaches to modulate CXCR4 in disease conditions.
“…Controlling the interaction of GPCRs with β-arrestin using the CRY-CIB photoactivation system has several advantages. Previous methods to control the interaction of GPCRs with β-arrestin were based on a chemical approach 23 , 33 . A pair of chemical dimerizers (FKBP-FRB) were used to control the interaction of β-arrestin with vasopressin receptors or chemokine receptors.…”
Section: Discussionmentioning
confidence: 99%
“…Combination of a novel indicators and the present system will provide powerful approaches for investigation of the relevance between conformation of β-arrestin and intracellular trafficking on cell surface and endosomes. In addition, several studies have demonstrated that the rapamycin-induced interaction of β-arrestin with vasopressin receptors or chemokine receptors promotes the phosphorylation of ERK1/2 23 , 33 . Light-induced interaction of β-arrestin with V2R also activated the phosphorylation of ERK1/2.…”
Intracellular trafficking of G protein-coupled receptors (GPCRs) controls their localization and degradation, which affects a cell’s ability to adapt to extracellular stimuli. Although the perturbation of trafficking induces important diseases, these trafficking mechanisms are poorly understood. Herein, we demonstrate an optogenetic method using an optical dimerizer, cryptochrome (CRY) and its partner protein (CIB), to analyze the trafficking mechanisms of GPCRs and their regulatory proteins. Temporally controlling the interaction between β-arrestin and β2-adrenergic receptor (ADRB2) reveals that the duration of the β-arrestin-ADRB2 interaction determines the trafficking pathway of ADRB2. Remarkably, the phosphorylation of ADRB2 by G protein-coupled receptor kinases is unnecessary to trigger clathrin-mediated endocytosis, and β-arrestin interacting with unphosphorylated ADRB2 fails to activate mitogen-activated protein kinase (MAPK) signaling, in contrast to the ADRB2 agonist isoproterenol. Temporal control of β-arrestin-GPCR interactions will enable the investigation of the unique roles of β-arrestin and the mechanism by which it regulates β-arrestin-specific trafficking pathways of different GPCRs.
“…Chemokines are a family of cytokines that promote and regulate inflammation by inducing various inflammatory cell subtypes . CXC chemokines interact with C‐X‐C chemokine receptor types 1/2 (CXCR1/2) receptors, playing a critical role in neutrophil migration and activation in areas of inflammation . Inhibition of the CXCLs/CXCR1/2 pathway results in an analgesic effect associated with decreased neutrophil infiltration in various animal models of inflammation .…”
The results of this study indicate that blockade of CXCR1/2 may represent a promising therapeutic strategy for the treatment of sepsis-associated ALI through regulation of neuropeptides and necroptosis.
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