Homo- and hetero-oligomerization of G protein-coupled receptors

Lee, Samuel P.; O’Dowd, Brian F.; George, Susan R.

doi:10.1016/j.lfs.2003.09.028

Cited by 86 publications

(61 citation statements)

References 28 publications

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“…Using human embryonic kidney HEK-293 cells stably expressing wild-type or mutated forms of CXCR4, we demonstrated that STAT3 phosphorylation requires the N-terminal part of the third intracellular loop (ICL3) and the tyrosine 157 present at the end of the second intracellular loop (ICL2) of CXCR4. In contrast, neither the conserved Tyr 135 in the DRY motif at the N terminus of ICL2 nor the Tyr 65 and Tyr 76 in the first intracellular loop (ICL1) are involved in this activation. ICL3, which does not contain any tyrosine residues, is needed to activate Jak2.…”

mentioning

confidence: 87%

“…Mutants-To investigate the role of intracellular tyrosines of CXCR4 in Jak2/ STAT3 activation pathway after SDF-1␣ binding, CXCR4 mutants containing single amino acid substitutions in which Tyr 65 and Tyr 135 were replaced by a phenylalanine and Tyr 76 and Tyr 157 by alanine were constructed and stably transfected in HEK-293 cells. We also used previously constructed HEK-293 cells expressing CXCR4 mutants in which each entire intracellular loop was modified (CXCR4.ICL1m, CXCR4.ICL2m, and CXCR4.ICL3m) and a HEK-293 cell line expressing a truncated form of CXCR4 (CXCR4.7TM) (54).…”

Section: Construction and Expression Of The Cxcr4mentioning

confidence: 99%

“…Tyrosines, residues that are phosphorylated during this activation pathway, are present in ICL1 (Tyr 65 and Tyr 76 ) and ICL2 (Tyr 135 and Tyr 157 ). Neither ICL3 nor the C-terminal domain of CXCR4 contains any tyrosine.…”

Section: The Third Intracellular Loop and The Tyrosine 157 Of Cxcr4mentioning

confidence: 99%

“…To address this question, we used previously constructed stable HEK-293 cells that express mutated forms of CXCR4 in which each intracellular loop (ICL) was replaced by a scrambled amino acid sequence of ICL1 that does not contain serine, threonine and tyrosine, or a CXCR4 form truncated at position 308 (54). We also constructed stable HEK-293 cells that express point mutation CXCR4 receptors in which each intracellular tyrosine, Tyr 65 (ICL1), Tyr 76 (ICL1), Tyr 135 (ICL2), or Tyr 157 (ICL2) was replaced by a nonphosphorylatable amino acid and deleted forms of ICL3. We demonstrate here that SDF-1␣-mediated Jak2/ STAT3 signaling is transduced through the N terminus of ICL3 and Tyr 157 of CXCR4.…”

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confidence: 99%

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Identification of the Cytoplasmic Domains of CXCR4 Involved in Jak2 and STAT3 Phosphorylation

Ahr

Denizot

Robert-Hebmann

et al. 2005

Journal of Biological Chemistry

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The chemokine SDF-1␣ transduces G i -dependent and -independent signals through CXCR4. Activation of Jak2/STAT3, a G i -independent signaling pathway, which plays a major role in survival signals, is known to be activated after SDF-1␣ binding to CXCR4 but the domains of CXCR4 involved in this signaling remain unexplored. Using human embryonic kidney HEK-293 cells stably expressing wild-type or mutated forms of CXCR4, we demonstrated that STAT3 phosphorylation requires the N-terminal part of the third intracellular loop (ICL3) and the tyrosine 157 present at the end of the second intracellular loop (ICL2) of CXCR4. In contrast, neither the conserved Tyr 135 in the DRY motif at the N terminus of ICL2 nor the Tyr 65 and Tyr 76 in the first intracellular loop (ICL1) are involved in this activation. ICL3, which does not contain any tyrosine residues, is needed to activate Jak2. These results demonstrate that two separate domains of CXCR4 are involved in Jak2/ STAT3 signaling. The N-terminal part of ICL3 is needed to activate Jak2 after SDF-1␣ binding to CXCR4, leading to phosphorylation of only one cytoplasmic Tyr, present at the C terminus of ICL2, which triggers STAT3 activation. This work has profound implications for the understanding of CXCR4-transduced signaling.

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mentioning

confidence: 87%

Section: Construction and Expression Of The Cxcr4mentioning

confidence: 99%

Section: The Third Intracellular Loop and The Tyrosine 157 Of Cxcr4mentioning

confidence: 99%

mentioning

confidence: 99%

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Identification of the Cytoplasmic Domains of CXCR4 Involved in Jak2 and STAT3 Phosphorylation

Ahr

Denizot

Robert-Hebmann

et al. 2005

Journal of Biological Chemistry

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“…M any G protein-coupled receptors (GPCRs) have been shown to assemble as homodimers, heterodimers, as well as larger oligomers (1,2). The existence of such oligomeric entities raises questions as to their functional consequences as well as their physiological relevance.…”

mentioning

confidence: 99%

The active metabolite of Clopidogrel disrupts P2Y12 receptor oligomers and partitions them out of lipid rafts

Savi

Zachayus

Delesque-Touchard

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

279

221

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P2Y12, a G protein-coupled receptor that plays a central role in platelet activation has been recently identified as the receptor targeted by the antithrombotic drug, clopidogrel. In this study, we further deciphered the mechanism of action of clopidogrel and of its active metabolite (Act-Met) on P2Y12 receptors. Using biochemical approaches, we demonstrated the existence of homooligomeric complexes of P2Y12 receptors at the surface of mammalian cells and in freshly isolated platelets. In vitro treatment with Act-Met or in vivo oral administration to rats with clopidogrel induced the breakdown of these oligomers into dimeric and monomeric entities in P2Y12 expressing HEK293 and platelets respectively. In addition, we showed the predominant association of P2Y12 oligomers to cell membrane lipid rafts and the partitioning of P2Y12 out of rafts in response to clopidogrel and Act-Met. The raft-associated P2Y12 oligomers represented the functional form of the receptor, as demonstrated by binding and signal transduction studies. Finally, using a series of receptors individually mutated at each cysteine residue and a chimeric P2Y12͞P2Y13 receptor, we pointed out the involvement of cysteine 97 within the first extracellular loop of P2Y12 in the mechanism of action of Act-Met. mechanism of action ͉ platelet ͉ antiaggregant M any G protein-coupled receptors (GPCRs) have been shown to assemble as homodimers, heterodimers, as well as larger oligomers (1, 2). The existence of such oligomeric entities raises questions as to their functional consequences as well as their physiological relevance. Heterologous expression systems have provided a variety of answers concerning ligand-dependent regulation of GPCR oligomeric states. Ligand binding, depending on the GPCR studied, can promote (3-10) or inhibit (11-13) dimer formation, as well as having no effect on preexisting constitutive homo-or heterodimers (14-25). The fact that heterodimerization may alter the pharmacological properties of a GPCR along with its internalization and signal transduction behavior is of critical importance (26, 27).Clustering, even for nonheptahelical receptors, now appears as a common feature of cell signaling. Specialized structures such as clathrin-coated pits, caveolae, and lipid rafts contain high concentrations of signaling molecules. Rafts represent dynamic assemblies of proteins and lipids, mostly sphingolipids and cholesterol (28,29). Proteins such as glycophosphatidylinositol-anchored proteins, nonreceptor tyrosine kinases, G␣ subunits of heterotrimeric G proteins, and palmitoylated proteins appear to localize to these microdomains (30). In addition, recent studies have shown that partitioning of proteins in and out of rafts can depend on their state of activation or dimerization (31-33). A variety of GPCR have also been identified in caveolae or rafts. These include ␣ and ␤-adrenergic receptors (34, 35), adenosine A1 receptor (36), angiotensin II type 1 receptor (37), muscarinic receptor (38), EDG1 receptor (39), bradykinin B1 and B2 receptor...

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Ethological resolution of behavioral topography and D2-like vs. D1-like agonist responses in congenic D4 dopamine receptor “knockouts”: Identification of D4:D1-like interactions

O’Sullivan

Kinsella²,

Grandy

et al. 2005

Synapse

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To clarify the involvement of dopamine D4 receptors in behavioral regulation, the phenotypic ethogram of congenic D4 "knockout" mice was studied in terms of (i) course of exploration and habituation, and (ii) topographical responsiveness to the selective D2-like agonist RU 24213 and the selective D1-like agonists A 68930, SK&F 83959 and SK&F 83822. Congenic D4 knockouts were characterized by a small reduction in exploratory sniffing with delayed habituation of sifting. The magnitude and topographical specificity of these effects indicated that any functional role for D4 receptors in exploratory processes is subtle. Induction of stereotyped, ponderous locomotion by RU 24213 was reduced in D4-null mice consistent with an involvement of D4 receptors in the topographical expression of stereotypy. Induction of grooming and, at higher doses, seizures by A 68930, which stimulates both adenylyl cyclase (AC) and phospholipase C (PLC), were unaltered in congenic D4 knockouts. In contrast, induction of grooming by SK&F 83959, which stimulates PLC but not AC and fails to induce seizures, was reduced in D4-null mice; this indicates that D4 receptors interact with PLC-coupled D1-like receptors in regulating D1-like-mediated grooming. Conversely, induction of seizures by SK&F 83822, which stimulates AC but not PLC and fails to induce grooming, was reduced in congenic D4 knockouts; this indicates that D4 receptors interact with AC-coupled D1-like receptors in regulating D1-like-mediated seizures. These studies identify novel functional roles for the D4 receptor that are distinct from those of closely related D2-like family members and involve interactions with their D1-like counterparts.

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Homo- and hetero-oligomerization of G protein-coupled receptors

Cited by 86 publications

References 28 publications

Identification of the Cytoplasmic Domains of CXCR4 Involved in Jak2 and STAT3 Phosphorylation

Identification of the Cytoplasmic Domains of CXCR4 Involved in Jak2 and STAT3 Phosphorylation

The active metabolite of Clopidogrel disrupts P2Y12 receptor oligomers and partitions them out of lipid rafts

Ethological resolution of behavioral topography and D2-like vs. D1-like agonist responses in congenic D4 dopamine receptor “knockouts”: Identification of D4:D1-like interactions

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