[Contemporary Clinical Neuroscience] The G Protein-Coupled Receptors Handbook Volume 215 || Receptor Oligomerization and Trafficking

GPCR Dimers and Trafficking 309

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From: Contemporary Clinical Neuroscience: The G Protein-Coupled Receptors HandbookEdited by: L. A. Devi © Humana Press Inc., Totowa, NJ

14Receptor Oligomerization and Trafficking

Selena E. Bartlett and Jennifer L. Whistler

1. INTRODUCTION

G protein-coupled receptor (GPCR) dimerization is a mechanism forregulating the signaling from several classes of plasma membrane receptorsand has been a particularly well-studied mechanism for the regulation oftyrosine kinase (Trk) receptors (reviewed in ref. 1). As reported for Trk andcytokine receptors, some GPCRs may also dimerize in response to agonist(2–6). However, GPCRs can also form constitutive dimers—often as earlyas during their biosynthesis (7–9). Adding to this complexity, GPCRs canform not only homodimers but also heterodimers with altered properties.Ultimately, one main goal is to understand the functional consequences ofGPCR dimerization.

GPCR dimerization was first reported for the gonadotropin-releasing hor-mone receptor (10), then for the β2-adrenergic receptor (AR) (11,12), and isnow considered to be a common theme for many GPCRs (for review, seerefs. 13–15). GPCRs can form homo- and heterodimers as well as higherorder oligomeric structures such as trimers, tetramers, and pentamers(12,16–18). For the purposes of this chapter, we use the word “dimer” todescribe all of these oligomers because it is the smallest possible oligomericunit. GPCR homodimerization has been shown to occur with several recep-tors, including the D2 dopamine receptor (19), the μ-opioid peptide receptor(MOP-R; ref. 20), several of the somatostatin (SST) receptors (21), and thechemokine CCR5 receptors (2), to name just a few. Heterodimerization hasbeen reported not only between receptors within the same GPCR family,such as the μ and δ-opioid peptide receptor (DOP-R) (22), the m2 and m3muscarinic receptors (23), and the CCR2 and CCR5 chemokine receptors

310 Bartlett and Whistler

(2), but also between diverse GPCR families, such as opioid receptors andβ2-ARs (24,25), adenosine A2a receptors and D1 dopamine receptors (26),and SSTR5 and D2 dopamine receptors (27). Although the existence of thesediverse heterodimers has yet to be demonstrated in vivo, the potential fortheir existence has profound implications for the biology of GPCR signalingand for drug design. There is a growing body of literature describing variousGPCR dimers and dimerization’s effects on receptor maturation through thesecretory pathway, ligand binding, and signaling in vitro (for review, seeref. 28). In contrast, there are relatively few studies describing the effects ofdimerization on receptor trafficking—particularly postendocytic trafficking.This chapter summarizes the ways that dimerization can alter the traffickingproperties of various GPCRs.

2. SECRETIONGPCR dimerization has been demonstrated in the absence of receptor

activation by agonists (for review, see ref. 14). Thus, it has been proposedthat GPCR dimerization may be the result of a constitutive process that occursearly in the biosynthetic pathway and is necessary for trafficking receptorsfrom the endoplasmic reticulum (ER) to the plasma membrane. For someGPCRs, heterodimerization with highly homologous receptors within thesame family is actually a prerequisite for the expression of functional recep-tor units. The γ-aminobutyric acid-B (GABAB) receptor is composed of twosubunits, GABABR1 and GABABR2. GABABR1 is not expressed at the cellsurface because it contains an ER retention signal, whereas GABABR2 isefficiently transported to the plasma membrane but is nonfunctional (29).Co-expression of GABABR2 with GABABR1 allows transport of both sub-units, presumably because heterodimerization of GABABR2 to GABABR1masks the ER retention signal (30,31). Intriguingly, it has also been demon-strated that the γ2S-subunit of the ionotropic GABAA receptor interacts withGABABR1 and can promote its cell-surface expression, thereby substitutingfor GABABR2 for trafficking of GABABR1 to the membrane. It can alsointeract with the GABABR1–GABABR2 heterodimeric complex to signifi-cantly enhance GABAB receptor internalization in response to agonist (32).This is the first demonstration of an interaction between a GPCR and an ionchannel affecting receptor trafficking. Although a general role of dimeriza-tion in ER export has yet to be established, studies that use fluorescence and/or bioluminescence resonance energy transfer with cell fractionation havedemonstrated the existence of receptor dimers in the ER for several receptorclasses, including CCR5 chemokine receptors (7), vasopressin receptors andoxytocin receptors (8), the yeast α-factor receptor (33), and the C5a chemo-tactic receptor (34).


Once the receptors have reached the plasma membrane, the debate con-tinues regarding whether ligands alter the dimerization state of the recep-tors. To date, there is no clear answer. Ligand binding has been shown toboth promote (6,27,35–40) or inhibit (26,41,42) dimerization, and severalgroups report no effect of ligand, suggesting instead that dimerization is aconstitutive process (7,8,34,43-50).

3. DESENSITIZATION AND ENDOCYTOSIS

Following transport to the plasma membrane and activation by an agonistligand, GPCRs are regulated by several mechanisms, many of which havebeen shown to be altered by receptor homo- or heterodimerization, includ-ing receptor desensitization, receptor endocytosis, receptor recycling, andreceptor degradation. For the purposes of this chapter, we define receptordesensitization as any process that alters the functional coupling of a recep-tor to its G protein/second messenger-signaling pathway. Endocytosis/inter-nalization is defined as the translocation of receptors from the cell surface toan intracellular compartment. Receptor recycling or receptor resensitizationis defined as receptors returning to the cell surface following their endocy-tosis. Finally, receptor degradation or downregulation is defined as any pro-cess that decreases the number of ligand-binding sites. Signaling fromGPCRs is rapidly regulated by a well-characterized and highly conservedcascade of events involving G protein coupling and activation, receptorphosphorylation by GPCR kinases (GRKs), and subsequent β-arrestinrecruitment that has been extensively reviewed elsewhere (51). These pro-cesses contribute directly to receptor desensitization by facilitating the un-coupling of the receptor from its G protein. GRK- and β-arrestin-mediateddesensitization is a rapid process that often occurs within minutes of recep-tor activation. However, receptors can also be desensitized/uncoupled fromG proteins by GRK and β-arrestin-independent mechanisms (for review, seeref. 52).

It has been reported that some GPCR heterodimeric complexes havealtered desensitization properties. For example, co-expression of the SSTreceptors SSTR2A and SSTR3 results in a slower desensitization of theSSTR2A–SSTR3 heterodimer compared to cells expressing SSTR2 orSSTR3 homodimers (53). Similarly, heterodimerization of the MOP-R andthe SST2A receptor leads to cross-desensitization of receptor function (21).This pattern of cross-desensitization is mirrored by co-expression andheterodimerization of the adenosine A2a receptor with the D2 dopaminereceptor (54) as well as by heterodimerization of the MOP-R with the CCR5chemokine receptor (55).


Following desensitization by GRKs and β-arrestin, GPCRs are rapidlyendocytosed into an intracellular compartment. This process occurs follow-ing even brief agonist exposure and is independent of signal transduction(56,57). GPCR internalization is predominantly mediated via the recruitmentof β-arrestins 1 and/or 2 to agonist-activated phosphorylated receptors. GPCRdimerization can alter the complement of proteins—particularly arrestins—that are recruited during endocytosis, thereby affecting the endocytic proper-ties of the receptors. In fact, heterodimerization has been shown to both inhibitand facilitate endocytosis, depending on the pair of receptors expressed.Heterodimerization of the β1-AR (a poor internalizer) and the β2-AR (a goodinternalizer) results in inhibition of agonist-promoted internalization of theβ2-AR (58). Similarly, the κ-opioid peptide receptor (KOP-R), which is apoor internalizer, inhibits the endocytosis of both the DOP-R (12) and the β2-AR (24) when it heterodimerizes with these receptors.

On the other hand, there are several examples of co-internalization of bothreceptors in a heterodimer pair, even when agonist ligand is present for onlyone of the receptors. These include the A2a adenosine–D2 dopamine dimerpair (54), in which addition of either the adenosine agonist or the dopamineagonist promotes internalization of both receptors, and the SSTR2a–MOPreceptor pair, in which activation with either the SSTR2a agonist or the MOP-R agonist promotes internalization of both receptors (21).

As mentioned earlier, in both cases, co-internalization was associated withcross-desensitization of receptor-mediated signaling. Similarly, α1a- andα1b-AR heterodimerization results in co-internalization of both receptorsfollowing treatment with oxymetazoline, an α1a-specific agonist. In con-trast, α1b does not co-internalize with agonist-activated neurokinin (NK)1receptors or agonist-activated CCR5 chemokine receptors, and neither ofthese receptors heterodimerizes with the α1b-AR (49), demonstrating thatdimerization appears to be required for co-internalization.

Another example of dimer-facilitated endocytosis occurs with mutant andwild-type MOP-Rs. Wild-type MOP-Rs do not endocytose when activatedby morphine (59). However, co-expression of a mutant MOP-R, D-MOP-R(which does internalize in response to morphine [60]) leads to dimerizationof MOP-R with D-MOP-R and endocytosis of both the wild-type and mutantMOP-R in response to morphine (20).

In some cases, it has been shown that the co-internalization effects aremodulated by changes in recruitment of β-arrestin. This has been elegantlydemonstrated with the thyrotropin-releasing hormone receptors 1 and 2(TRHR1 and TRHR2) dimers 961 and also with MOP-R and NK1-R dimers(61,62). TRHR2 interacts preferentially with β-arrestin 2 when expressed


alone in HEK293 and simian fibroblast (COS) cells. However, co-expres-sion of TRHR1 leads to interaction of both TRHR1 and TRHR2 with β-arrestin 1. As a consequence, the internalization rates for both TRHR1 andTRHR2 are altered, with no effect on ligand binding or basal inositol phos-phate production (61).

Based on the pattern of arrestin recruitment, GPCRs can be divided intotwo categories: class A and class B (63). Class A receptors have a higheraffinity for β-arrestin 2 than β-arrestin 1, and, after initial recruitment, class Areceptors rapidly dissociate from β-arrestin. Class B receptors bind β-arrestin1 and 2 with equal affinities, and, unlike class A receptors, class B receptorsremain in a stable complex with arrestins during endocytosis and, therefore,are colocalized with arrestins in endosomes. Alterations in the pattern of β-arrestin 2 recruitment are seen with MOP-R–NK1-R heterodimers. In cellsexpressing only the MOP-R, β-arrestin 2 is recruited to the activated recep-tors; however, β-arrestin 2 does not co-internalize with the receptor classifiesMOP-Rs as class A. However, when the NK1-R is co-expressed with theMOP-R, the heterodimers recruit β-arrestin 2 in response to either the MOP-R agonist [D-Ala(2)-N-Me-Phe(4), Gly(5)-ol]-enkephalin (DAMGO) or theNK1-R agonist substance P, and β-arrestin 2 is then co-internalized with theMOP-R–NK1-R heterodimeric complex, switching the classification of theMOP-R to class B (62). Furthermore, in the case of the MOP-R–NK1-R pair,co-internalization of β-arrestin 2 with the heterodimer leads to a delay in bothMOP-R recycling and MOP-R resensitization kinetics (62). This observationleads to the conclusion that heterodimerization may also affect receptor traf-ficking—more specifically, postendocytic sorting.

4. POSTENDOCYTIC TRAFFICKING

Receptors that have been desensitized and rapidly endocytosed areuniquely poised to make an important decision that has substantial impacton future signal transduction. As mentioned earlier, following endocytosis,receptors can be recycled, thereby restoring the functional complement ofreceptors. Alternatively, receptors that have been endocytosed can be tar-geted for degradation, thereby decreasing the functional complement of re-ceptors and ultimately resulting in receptor downregulation. Althoughendocytosis and subsequent degradation of receptors is not the only meansto produce receptor downregulation, it can produce receptor downregulationrapidly, even following brief exposure to agonist (64). Apparent receptordownregulation can also be affected by alterations in rate of receptor syn-thesis and/or folding or secretion (65). Not all GPCRs are downregulatedfollowing their endocytosis. For example, although both the MOP-R and


DOP-R are class A receptors that are endocytosed via clathrin-coated pitsfollowing agonist-induced activation, GRK phosphorylation, and associa-tion with β-arrestins, they differ in their fate following endocytosis. WhereasMOP-Rs are recycled following their endocytosis, DOP-Rs are transporteddeeper into the endocytic pathway and are rapidly degraded by the lysosome(60,66) and hence downregulated.

As discussed earlier, β-arrestin 2 is recruited and co-internalized with theMOP-R–NK1-R heterodimeric complex in response to either the μ-opioidagonist DAMGO or the NK1 receptor agonist substance P. This leads to adelay in both MOP-R recycling and MOP-R resensitization kinetics (62),suggesting that heterodimerization can alter the post-endocytic traffickingproperties of the MOP-R.

Heterodimerization of GPCRs may also result in changes, not only to therate of postendocytic recycling but also to the actual postendocytic fate ofthe receptors. This has been beautifully demonstrated using the V1a vaso-pressin receptor (V1aR), which heterodimerizes with V2 vasopressin recep-tor (V2R) (8). V1aR activation results in the recruitment of β-arrestin.Following receptor internalization, β-arrestin dissociates from V1aR, and thereceptor is rapidly recycled to the plasma membrane, which classifies theV1aR as a class A GPCR. In contrast, V2Rs recruit β-arrestin, and followinginternalization, β-arrestin fails to dissociate from the receptor, resulting inV2R accumulation in endosomes in the perinuclear compartment (67); thisclassifies V2R as a class B receptor. The postendocytic fates of V1aR andV2R are altered when the receptors are co-expressed. Nonselective agonistactivation of both V1aR and V2R results in the co-internalization of bothV1aR and V2R with β-arrestin into endosomes, where the interaction inhib-its the recycling of both receptors to the plasma membrane, thereby switch-ing the V1aR from a class A to a class B receptor. Selective activation ofonly V1aR results in co-internalization of the V2R, without co-internaliza-tion of β-arrestin. This results in rapid recycling of both V1aR and V2R,thereby converting V2R to class A receptor (67). These data suggest that forthe vasopressin receptors, it is the activated receptor within the heterodimericcomplex that determines the overall postendocytic fate of all receptors withinthe complex. It remains to be determined whether this is true for all reportedreceptor heterodimers.

5. CLINICAL RELEVANCE AND/OR BENEFITS

When one begins to think of all the combinations and permutationswhereby GPCRs could homo- or heterodimerize, the implications for uniquedrug targets or “landing pads” becomes apparent and really quite over-


whelming. Already there are several examples of ways in whichheterodimerization, solely by affecting trafficking, affects phenomena im-portant for human disease.

5.1. Human Immunodeficiency Virus Infection

CCR5Δ32, the truncated form of the human immunodeficiency virus(HIV) coreceptor CCR5, conveys resistance to HIV infection even in indi-viduals in the heterozygous state (68–71). Co-expression of this truncatedCCR5 with the wild-type CCR5 leads to retention of both receptors in theER. Because CCR5 forms heterodimers, it has been suggested that by form-ing a heterodimer with the wild-type receptor, the mutant receptor delaystransport of CCR5 coreceptors to the surface of cells, thereby preventingHIV coreceptor function (72). The existence of naturally occurringchemokine receptor mutants that alter CCR5 trafficking may be an explana-tion for the subpopulations of patients with acquired immune deficiencysyndrome that display differential resistance to HIV infection. This may helpguide future therapeutic strategies for its treatment.

5.2. Attenuation of Morphine Tolerance and Dependence

The current evidence suggests that receptor oligomerization may openthe door to the discovery of new, more effective drug treatments for disease.One example is the amelioration of the development of morphine toleranceand dependence. This section outlines studies that suggest that targeting theMOP-R oligomeric complex may be helpful to reduce the development ofmorphine tolerance.

Although opioids such as morphine remain the analgesic of choice inmany cases, a major limitation to their long-term use is the development ofphysiological tolerance, which is a profound decrease in analgesic effectobserved in all patients during prolonged administration of opioid drug.Despite considerable progress, the molecular and cellular mechanisms me-diating the development of tolerance and withdrawal to morphine remaincontroversial. Multiple hypotheses exist to explain morphine tolerance.

In contrast to other opioids and many other agonists in general, morphinefails to promote endocytosis of the wild-type μ-opioid receptor in culturedcells (59,73) and native neurons (74,75), whereas endogenous peptideligands such as endorphins and several opioid drugs, such as methadone(76), readily drive receptor endocytosis (74,75,77). Morphine-activatedMOP-Rs are relatively unique because they are not GRK-phosphorylatednor do they efficiently recruit β-arrestin, although they are in an “active”receptor conformation (78–80). Hence, morphine-activated MOP-Rs gener-


ally elude an important, highly conserved regulatory mechanism designedto rapidly modulate receptor-mediated signaling.

Whistler et al. have proposed that the regulation of opioid receptors byendocytosis serves as a protective role in reducing the development of toler-ance and dependence to opioid drugs (20,81). According to this model, pro-moting morphine-induced endocytosis of the MOP-R should lead to anattenuation of the development of morphine tolerance and dependence. Thishas been shown to be true in a cell culture model, where mutations of theMOP-R that enhance morphine-induced endocytosis ameliorate a cellularhallmark of tolerance (60). Conversely, mutations that prevented endocyto-sis of the MOP-R in response to ligands (such as methadone) enhancedmethadone-mediated tolerance (60). These observations led to the predic-tion that one might be able to use the oligomeric state of the MOP-R to alterits endocytosis in response to morphine, thereby altering the developmentof morphine tolerance.

Briefly, could a MOP-R dimer in which one protomer was occupied withDAMGO (which facilitates endocytosis) “drag” the other morphine-occu-pied protomer into the cell? In fact, co-administration of a small dose ofDAMGO facilitated morphine-induced endocytosis of the MOP-R both incell culture models and in vivo (20). Importantly, facilitation of MOP-Rendocytosis also prevented the development of morphine tolerance (20).Pharmaceutical interventions that target heterodimeric MOP-Rs that facili-tate the endocytosis of the MOP-R in response to morphine may play impor-tant roles in the attenuation of morphine tolerance and dependence.

6. CONCLUSIONS

Determining whether opioid receptor heterodimers or opioid–other GPCRdimers exist in vivo is a true challenge. Advances in bivalent ligands thatselectively target heterodimers may provide the necessary tools (for review,see refs. 82 and 83). Additionally, the phenotypes of the opioid receptorknockout mice also provide hints that heterodimerization, at least amongthe opioid receptors, is a real phenomenon. For example, some μ-opioidreceptor-specific analgesia is lost in the δ-opioid receptor-deficient mice(84). One explanation for this observation could be that a μ–δ opioid recep-tor dimer has also been eliminated in the genetically modified animals. Mak-ing the assumption that GPCRs form dimers in vivo, it may be possible todesign therapeutics to specifically target these complexes, which thereforeprovide more specific drug treatments. Signaling from GPCRs will ulti-mately depend on the cell-type and its specific environment. Different sig-nals may be generated from the same receptor, depending on the intracellular


complement of other GPCRs available and/or the environment of the cell.Understanding these differences will be important for the design of drugsand the treatment of diseases.

ACKNOWLEDGMENTS

S. E. Bartlett is supported in part by the Sidney Baer Trust as a NARSADYoung Investigator. J. L. Whistler is supported by a National Institutes ofHealth grant R01 DA015232 and by funds provided by the State of Califor-nia for medical research on alcohol and substance abuse through UCSF. Wewould like to thank Dr. Maria Waldhoer for a critical reading of the manu-script.

REFERENCES

1. Hackel PO, Zwick E, Prenzel N, Ullrich A. Epidermal growth factor receptors:critical mediators of multiple receptor pathways. Curr Opin Cell Biol 1999;11:184–189.

2. Mellado M, Rodriguez-Frade JM, Vila-Coro AJ, et al. Chemokine receptorhomo- or heterodimerization activates distinct signaling pathways. EMBO J2001;20:2497–2507

3. AbdAlla S, Lother H, Quitterer U. AT1-receptor heterodimers show en-hanced G-protein activation and altered receptor sequestration. Nature2000;407:94–98.

4. AbdAlla S. Zaki E, Lother H, Quitterer U. Involvement of the amino terminusof the B(2) receptor in agonist-induced receptor dimerization. J Biol Chem1999;274:26,079–26,084

5. Horvat RD, Barisas BG, Roess DA. Luteinizing hormone receptors are self-associated in slowly diffusing complexes during receptor desensitization. MolEndocrinol 2001;15:534–542.

6. Horvat RD, Roess DA, Nelson SE, Barisas BG, Clay CM. Binding of agonistbut not antagonist leads to fluorescence resonance energy transfer betweenintrinsically fluorescent gonadotropin-releasing hormone receptors. MolEndocrinol 2001;15:695–703.

7. Issafras H, Angers S, Bulenger S, et al. Constitutive agonist-independent CCR5oligomerization and antibody-mediated clustering occurring at physiologicallevels of receptors. J Biol Chem 2002;277:34,666–34,673.

8. Terrillon S, Durroux T, Mouillac B, et al. Oxytocin and vasopressin V1a andV2 receptors form constitutive homo- and heterodimers during biosynthesis.Mol Endocrinol 2003;17:677–691.

9. Romano C, Yang WL, O’Malley KL. Metabotropic glutamate receptor 5 is adisulfide-linked dimer. J Biol Chem 1996;271:28,612–28,616.

10. Conn PM, Rogers DC, Stewart JM, Niedel J, Sheffield T. Conversion of a


gonadotropin-releasing hormone antagonist to an agonist. Nature1982;296:653–655.

11. Hebert TE, Moffett S, Morello JP, et al. A peptide derived from a beta2-adren-ergic receptor transmembrane domain inhibits both receptor dimerization andactivation. J Biol Chem 1996;271:16,384–16,392.

12. Jordan BA, Devi LA. G-protein-coupled receptor heterodimerization modu-lates receptor function. Nature 1999;399:697–700.

13. Terrillon S, Bouvier M. Roles of G-protein-coupled receptor dimerization.EMBO Rep 2004;5:30–34.

14. Bai M. Dimerization of G-protein-coupled receptors: roles in signal transduc-tion. Cell Signal 2004;16:175–186.

15. Gomes I, Filipovska J, Jordan BA, Devi LA. Oligomerization of opioid recep-tors. Methods 2002;27:358–365.

16. Lee SP, O’Dowd BF, Ng GY, et al. Inhibition of cell surface expression bymutant receptors demonstrates that D2 dopamine receptors exist as oligomersin the cell. Mol Pharmacol 2000;58:120–128.

17. Zawarynski P, Tallerico T, Seeman P, Lee SP, O’Dowd BF, George SR.Dopamine D2 receptor dimers in human and rat brain. FEBS Lett1998;441:383–386.

18. Klco JM, Lassere TB, Baranski TJ. C5a receptor oligomerization. I. Disulfidetrapping reveals oligomers and potential contact surfaces in a G protein-coupled receptor. J Biol Chem 2003;278:35,345–35,353.

19. Lee SP, O’Dowd BF, Rajaram RD, Nguyen T, George SR. D2 dopamine re-ceptor homodimerization is mediated by multiple sites of interaction, includ-ing an intermolecular interaction involving transmembrane domain 4.Biochemistry 2003;42:11,023–11,031.

20. He L, Fong J, von Zastrow M, Whistler JL. Regulation of opioid receptor traf-ficking and morphine tolerance by receptor oligomerization. Cell2002;108:271–282.

21. Pfeiffer M, Koch T, Schroder H, Laugsch M, Hollt V, Schulz S.Heterodimerization of somatostatin and opioid receptors cross-modulates phos-phorylation, internalization, and desensitization. J Biol Chem 2002;277:19,762–19,772.

22. Gomes I, Jordan BA, Gupta A, Trapaidze N, Nagy V, Devi LA.Heterodimerization of mu and delta opioid receptors: A role in opiate synergy.J Neurosci 2000;20:RC110.

23. Maggio R, Vogel Z, Wess J. Coexpression studies with mutant muscarinic/adrenergic receptors provide evidence for intermolecular “cross-talk” betweenG-protein-linked receptors. Proc Natl Acad Sci USA 1993;90:3103–3107.

24. Jordan BA, Trapaidze N, Gomes I, Nivarthi R, Devi LA. Oligomerization ofopioid receptors with beta 2-adrenergic receptors: a role in trafficking and mi-togen-activated protein kinase activation. Proc Natl Acad Sci USA2001;98:343–348.

25. Rios CD, Jordan BA, Gomes I, Devi LA. G-protein-coupled receptor dimer-ization: modulation of receptor function. Pharmacol Ther 2001;92:71–87.

26. Gines S, Hillion J, Torvinen M, et al. Dopamine D1 and adenosine A1 recep-


tors form functionally interacting heteromeric complexes. Proc Natl Acad SciUSA 2000;97:8606–8611.

27. Rocheville M, Lange DC, Kumar U, Patel SC, Patel RC, Patel YC. Receptorsfor dopamine and somatostatin: formation of hetero-oligomers with enhancedfunctional activity. Science 2000;288:154–157.

28. Devi LA. Heterodimerization of G-protein-coupled receptors: pharmacology,signaling and trafficking. Trends Pharmacol Sci 2001;22:532–537.

29. Margeta-Mitrovic M, Jan YN, Jan LY. A trafficking checkpoint controlsGABA(B) receptor heterodimerization. Neuron 2000;27:97–106.

30. Galvez T, Duthey B, Kniazeff J, et al. Allosteric interactions between GB1 andGB2 subunits are required for optimal GABA(B) receptor function. EMBO J2001;20:2152–2159.

31. Pagano A, Rovelli G, Mosbacher J, et al. C-terminal interaction is essential forsurface trafficking but not for heteromeric assembly of GABA(b) receptors. JNeurosci 2001;21:1189–1202.

32. Balasubramanian S, Teissere JA, Raju DV, Hall RA. Hetero-oligomerizationbetween GABA-A and GABA-B receptors regulates GABA-B receptor traf-ficking. J Biol Chem 2004;279:18,840–18,850.

33. Overton MC, Blumer KJ. The extracellular N-terminal domain and transmem-brane domains 1 and 2 mediate oligomerization of a yeast G protein-coupledreceptor. J Biol Chem 2002;277:41,463–41,472.

34. Floyd DH, Geva A, Bruinsma SP, Overton MC, Blumer KJ, Baranski TJ. C5areceptor oligomerization. II. Fluorescence resonance energy transfer studies ofa human G protein-coupled receptor expressed in yeast. J Biol Chem 2003;278:35,354–35,361.

35. Patel RC, Kumar U, Lamb DC, et al. Ligand binding to somatostatin receptorsinduces receptor-specific oligomer formation in live cells. Proc Natl Acad SciUSA 2002;99:3294–3299.

36. Rodriguez-Frade JM, Vila-Coro AJ, de Ana AM, Albar JP, Martinez AC,Mellado M. The chemokine monocyte chemoattractant protein-1 induces func-tional responses through dimerization of its receptor CCR2. Proc Natl AcadSci USA 1999;96:3628–3633

37. Cornea A, Janovick JA, Maya-Nunez G, Conn PM. Gonadotropin-releasinghormone receptor microaggregation. Rate monitored by fluorescence resonanceenergy transfer. J Biol Chem 2001;276:2153–2158.

38. Kroeger KM, Hanyaloglu AC, Seeber RM, Miles LE, Eidne KA. Constitutiveand agonist-dependent homo-oligomerization of the thyrotropin-releasing hor-mone receptor. Detection in living cells using bioluminescence resonance en-ergy transfer. J Biol Chem 2001;276:12,736–12,743.

39. Zhu CC, Cook LB, Hinkle PM. Dimerization and phosphorylation of thyrotro-pin-releasing hormone receptors are modulated by agonist stimulation. J BiolChem 2002;277:28,228–28,237.

40. Hunzicker-Dunn M, Barisas G, Song J, Roess DA. Membrane organization ofluteinizing hormone receptors differs between actively signaling and desensi-tized receptors. J Biol Chem 2003;278:42,744–42,749.

41. Cheng ZJ, Miller LJ. Agonist-dependent dissociation of oligomeric complexes


of G protein-coupled cholecystokinin receptors demonstrated in living cellsusing bioluminescence resonance energy transfer. J Biol Chem2001;276:48,040–48,047.

42. Latif R, Graves P, Davies TF. Ligand-dependent inhibition of oligomerizationat the human thyrotropin receptor. J Biol Chem 2002;277:45,059–45,067.

43. Overton MC, Blumer KJ. G-protein-coupled receptors function as oligomersin vivo. Curr Biol 2000;10:341–344.

44. Jensen AA, Hansen JL, Sheikh SP, Brauner-Osborne H. Probing intermolecu-lar protein–protein interactions in the calcium-sensing receptor homodimerusing bioluminescence resonance energy transfer (BRET). Eur J Biochem2002;269:5076–5087.

45. Babcock GJ, Farzan M, Sodroski J. Ligand-independent dimerization ofCXCR4, a principal HIV-1 coreceptor. J Biol Chem 2003;278:3378–3385.

46. Canals M, Marcellino D, Fanelli F, et al. Adenosine A2A-dopamine D2 recep-tor–receptor heteromerization: qualitative and quantitative assessment byfluorescence and bioluminescence energy transfer. J Biol Chem2003;278:46,741–46,749.

47. Dinger MC, Bader JE, Kobor AD, Kretzschmar AK, Beck-Sickinger AG.Homodimerization of neuropeptide y receptors investigated by fluorescenceresonance energy transfer in living cells. J Biol Chem 2003;278:10,562–10,571.

48. Guo W, Shi L, Javitch JA. The fourth transmembrane segment forms the in-terface of the dopamine D2 receptor homodimer. J Biol Chem2003;278:4385–4388.

49. Stanasila L, Perez JB, Vogel H, Cotecchia S. Oligomerization of the alpha 1a-and alpha 1b-adrenergic receptor subtypes. Potential implications in receptorinternalization. J Biol Chem 2003;278:40,239–40,251.

50. Trettel F, Di Bartolomeo S, Lauro C, Catalano M, Ciotti MT, Limatola C. Ligand-independent CXCR2 dimerization. J Biol Chem 2003;278:40,980–40,988.

51. Ferguson SS. Evolving concepts in G protein-coupled receptor endocytosis: therole in receptor desensitization and signaling. Pharmacol Rev 2001;53:1–24.

52. Liu JG, Anand KJ. Protein kinases modulate the cellular adaptations associ-ated with opioid tolerance and dependence. Brain Res Brain Res Rev2001;38:1–19.

53. Pfeiffer M, Koch T, Schroder H, et al. Homo- and heterodimerization of soma-tostatin receptor subtypes. Inactivation of sst(3) receptor function byheterodimerization with sst(2A). J Biol Chem 2001;276:14,027–14,036.

54. Hillion J, Canals M, Torvinen M, et al. Coaggregation, cointernalization, andcodesensitization of adenosine A2A receptors and dopamine D2 receptors. JBiol Chem 2002;277:18,091–18,097.

55. Chen C, Li J, Bot G, Szabo I, Rogers TJ, Liu-Chen LY. Heterodimerizationand cross-desensitization between the mu-opioid receptor and the chemokineCCR5 receptor. Eur J Pharmacol 2004;483:175–186.

56. Zaki PA, Keith DE, Jr., Thomas JB, Carroll FI, Evans CJ. Agonist-, antago-nist-, and inverse agonist-regulated trafficking of the delta-opioid receptor cor-


relates with, but does not require, G protein activation. J Pharmacol Exp Ther2001;298:1015–1020.

57. Remmers AE, Clark MJ, Liu XY, Medzihradsky F. Delta opioid receptor down-regulation is independent of functional G protein yet is dependent on agonistefficacy. J Pharmacol Exp Ther 1998;287:625–632.

58. Lavoie C, Mercier JF, Salahpour A, et al. Beta 1/beta 2-adrenergic receptorheterodimerization regulates beta 2-adrenergic receptor internalization andERK signaling efficacy. J Biol Chem 2002;277:35,402–35,410.

59. Keith DE, Murray SR, Zaki PA, et al. Morphine activates opioid receptors with-out causing their rapid internalization. J Biol Chem 1996;271:19,021–19,024.

60. Finn AK, Whistler JL. Endocytosis of the mu opioid receptor reduces toler-ance and a cellular hallmark of opiate withdrawal. Neuron 2001;32:829–839.

61. Hanyaloglu AC, Seeber RM, Kohout TA, Lefkowitz RJ, Eidne KA. Homo-and hetero-oligomerization of thyrotropin-releasing hormone (TRH) receptorsubtypes. Differential regulation of beta-arrestins 1 and 2. J Biol Chem2002;277:50,422–50,430.

62. Pfeiffer M, Kirscht S, Stumm R, et al. Heterodimerization of substance P andmu-opioid receptors regulates receptor trafficking and resensitization. J BiolChem 2003;278:51,630–51,637.

63. Oakley RH, Laporte SA, Holt JA, Caron MG, Barak LS. Differential affinitiesof visual arrestin, beta arrestin1, and beta arrestin2 for G protein-coupled re-ceptors delineate two major classes of receptors. J Biol Chem2000;275:17,201–17,210.

64. Tsao PI, von Zastrow M. Type-specific sorting of G protein-coupled receptorsafter endocytosis. J Biol Chem 2000;275:11,130–11,140.

65. Petaja-Repo UE, Hogue M, Bhalla S, Laperriere A, Morello JP, Bouvier M.Ligands act as pharmacological chaperones and increase the efficiency of deltaopioid receptor maturation. EMBO J 2002;21:1628–1637.

66. Whistler JL, Enquist J, Marley A, et al. Modulation of postendocytic sorting ofG protein-coupled receptors. Science 2002;297:615–620.

67. Terrillon S, Barberis C, Bouvier M. Heterodimerization of V1a and V2 vaso-pressin receptors determines the interaction with beta-arrestin and their traf-ficking patterns. Proc Natl Acad Sci USA 2004;101:1548–1553.

68. Dean M, Jacobson LP, McFarlane G, et al. Reduced risk of AIDS lymphoma inindividuals heterozygous for the CCR5-delta32 mutation. Cancer Res1999;59:3561–3564.

69. Samson M, Libert F, Doranz BJ, et al. Resistance to HIV-1 infection incaucasian individuals bearing mutant alleles of the CCR-5 chemokine receptorgene. Nature 1996;382:722–725.

70. Huang Y, Paxton WA, Wolinsky SM, et al. The role of a mutant CCR5 allele inHIV-1 transmission and disease progression. Nat Med 1996;2:1240–1243.

71. Michael NL, Chang G, Louie LG, et al. The role of viral phenotype and CCR-5 gene defects in HIV-1 transmission and disease progression. Nat Med1997;3:338–340.


72. Benkirane M, Jin DY, Chun RF, Koup RA, Jeang KT. Mechanism oftransdominant inhibition of CCR5-mediated HIV-1 infection by ccr5delta32. JBiol Chem 1997;272:30,603–30,606.

73. Arden JR, Segredo V, Wang Z, Lameh J, Sadee W. Phosphorylation and ago-nist-specific intracellular trafficking of an epitope-tagged mu-opioid receptorexpressed in HEK 293 cells. J Neurochem 1995;65:1636–1645.

74. Sternini C, Spann M, Anton B, et al. Agonist-selective endocytosis of mu opioidreceptor by neurons in vivo. Proc Natl Acad Sci USA 1996;93:9241–9246.

75. Keith DE, Anton B, Murray SR, et al. mu-Opioid receptor internalization: opi-ate drugs have differential effects on a conserved endocytic mechanism in vitroand in the mammalian brain. Mol Pharmacol 1998;53:377–384.

76. Garrido MJ, Troconiz IF. Methadone: a review of its pharmacokinetic/pharma-codynamic properties. J Pharmacol Toxicol Methods 1999;42:61–66.

77. Trapaidze N, Gomes I, Cvejic S, Bansinath M, Devi LA. Opioid receptor en-docytosis and activation of MAP kinase pathway. Brain Res Mol Brain Res2000;76:220–228.

78. Blake AD, Bot G, Freeman JC, Reisine T. Differential opioid agonist regula-tion of the mouse mu opioid receptor. J Biol Chem 1997;272:782–790.

79. Whistler JL, von Zastrow M. Morphine-activated opioid receptors eludedesensitization by beta-arrestin. Proc Natl Acad Sci USA 1998;95:9914–9919.

80. Zhang J, Ferguson SS, Barak LS, et al. Role for G protein-coupled receptorkinase in agonist-specific regulation of mu-opioid receptor responsiveness.Proc Natl Acad Sci USA 1998;95:7157–7162.

81. Whistler JL, Chuang HH, Chu P, Jan LY, von Zastrow M. Functional dissocia-tion of mu opioid receptor signaling and endocytosis: implications for the biol-ogy of opiate tolerance and addiction. Neuron 1999;23:737–746.

82. Portoghese PS. From models to molecules: opioid receptor dimers, bivalentligands, and selective opioid receptor probes. J Med Chem 2001;44:2259–2269.

83. Lutz RA, Pfister HP. Opioid receptors and their pharmacological profiles. JRecept Res 1992;12:267–286.

84. Zhu Y, King MA, Schuller AG, et al. Retention of supraspinal delta-like anal-gesia and loss of morphine tolerance in delta opioid receptor knockout mice.Neuron 1999;24:243–252.

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[Contemporary Clinical Neuroscience] The G Protein-Coupled Receptors Handbook Volume 215 || Receptor Oligomerization and Trafficking