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(Circulation Research. 2002;91:672.)
© 2002 American Heart Association, Inc.

Reviews

Regulation of G Protein–Coupled Receptor Signaling by Scaffold Proteins

Randy A. Hall, Robert J. Lefkowitz

From the Department of Pharmacology (R.A.H.), Emory University School of Medicine, Atlanta, Ga; and the Departments of Medicine and Biochemistry (R.J.L.), Howard Hughes Medical Institute, Duke University Medical Center, Durham, NC.

Correspondence to Robert Lefkowitz, MD, Duke University, Howard Hughes Medical Institute, Departments of Medicine and Biochemistry, Box 3821, Durham, NC 27710. E-mail lefko001{at}receptor-biol.duke.edu

	Abstract

Top Abstract Introduction GPCR Carboxyl-Terminal... GPCR Carboxyl-Terminal... GPCR Interactions With... Scaffold Versus Nonscaffold... Agonist-Dependence of... Problems in Studying... Concluding Thoughts References

 
The actions of many hormones and neurotransmitters are mediated through stimulation of G protein–coupled receptors. A primary mechanism by which these receptors exert effects inside the cell is by association with heterotrimeric G proteins, which can activate a wide variety of cellular enzymes and ion channels. G protein–coupled receptors can also interact with a number of cytoplasmic scaffold proteins, which can link the receptors to various signaling intermediates and intracellular effectors. The multicomponent nature of G protein–coupled receptor signaling pathways makes them ideally suited for regulation by scaffold proteins. This review focuses on several specific examples of G protein–coupled receptor-associated scaffolds and the roles they may play in organizing receptor-initiated signaling pathways in the cardiovascular system and other tissues.

Key Words: GPCR • adrenergic • heptahelical • arrestin • phosphorylation

	Introduction

Top Abstract Introduction GPCR Carboxyl-Terminal... GPCR Carboxyl-Terminal... GPCR Interactions With... Scaffold Versus Nonscaffold... Agonist-Dependence of... Problems in Studying... Concluding Thoughts References

 
Cardiovascular function is regulated by a wide variety of hormones and neurotransmitters. The vast majority of hormones and neurotransmitters in the cardiovascular system exert their cellular effects through activation of G protein–coupled receptors (GPCRs), which are cell surface receptors also known as heptahelical receptors, 7-transmembrane receptors, and serpentine receptors because of their characteristic transmembrane topology. Agonist stimulation of a GPCR results in a conformational change in the receptor, leading to receptor association with heterotrimeric G proteins within the plasma membrane. This interaction causes conformational changes within the G-protein subunits leading to GDP release and GTP binding to the G subunit. The G and Gß subunits then dissociate from each other and exert regulation over various effectors such as enzymes and ion channels. Due to the multicomponent nature of GPCR signaling pathways, which involve at least a receptor, a G-protein complex, and an effector, these pathways can be made substantially more efficient via localization of the various components in close proximity to each other.

GPCRs associate not only with G proteins, but with a variety of other proteins as well.^1–4 GPCR-associated proteins may play at least four distinct roles in receptor signaling. First, a GPCR-associated protein may directly mediate receptor signaling, as in the case of G proteins. Second, a GPCR-associated protein may regulate receptor signaling through controlling receptor localization and/or trafficking. Third, a GPCR-associated protein may act as an allosteric modulator of receptor conformation, altering receptor pharmacology and/or other aspects of receptor function. Finally, a GPCR-associated protein may act as a scaffold, physically linking the receptor to various effectors. These four roles are by no means mutually exclusive. Each GPCR-associated protein may fill one, two, three, or all four of these roles.

Scaffold proteins may be defined as proteins that associate with two or more partners to enhance the efficiency and/or specificity of cellular signaling pathways. Interest in scaffold proteins has increased dramatically over the past decade, due to an explosion of technological advances in the detection and analysis of protein-protein interactions. These advances have led to the realization that many proteins have multiple binding partners and may therefore function as scaffold proteins. There are many different classes of scaffold protein, and over the past several years there have been a number of reviews on this subject.^5–9

This review article will not attempt to exhaustively list all of the proteins that have been reported to associate with various G protein–coupled receptors.^1–4 Instead, the focus will be on describing a few specific cases where GPCR-associated proteins appear to be acting as scaffolds (Table). Furthermore, this review will not focus exclusively on GPCRs involved in regulating cardiovascular function. Several of the best examples of regulation of GPCR signaling by scaffold proteins come from outside the cardiovascular system, and these will be discussed as prototypes of GPCR/scaffold interactions. It is likely that there are many interactions between cardiovascular GPCRs and scaffold proteins that are physiologically important but that have not yet been characterized. Work on the prototypical examples of GPCR/scaffold interactions described in this review have helped to lay the foundation for understanding the structural determinants and functional importance of other GPCR/scaffold interactions that may be discovered in the future.

View this table: [in this window] [in a new window]   Table 1. List of GPCR/Scaffold Interactions

As illustrated in Figure 1, a GPCR can associate with a cytoplasmic scaffolding protein via one of three intracellular loops or via the receptor’s intracellular carboxyl-terminus. The distal portions of many GPCR carboxyl-termini are known to interact with scaffold proteins containing PDZ domains,⁹ and these interactions will be discussed in the first section of this review. The carboxyl-termini of many GPCRs are also known to be involved in associations with various non-PDZ scaffold proteins, and examples of these interactions will be discussed in the second section of this review. Finally, the third intracellular loops of most GPCRs contain key determinants for association with ß-arrestins, important scaffold proteins that will be discussed in the third section of this review. The two types of GPCR that have been studied most intensively as model systems of the regulation of GPCR function are rhodopsin and ß-adrenergic receptors,¹⁰ and therefore these two receptor classes will be discussed first.

View larger version (26K): [in this window] [in a new window]   Figure 1. Schematic of GPCR/scaffold interactions. Scaffold proteins that interact with GPCRs may be divided into three broad categories: (1) PDZ scaffolds, which associate with the distal portions of GPCR carboxyl-termini; (2) various non-PDZ scaffolds, which associate with GPCR carboxyl-termini or other GPCR cytoplasmic regions; and (3) ß-arrestins, which associate with many GPCRs and typically recognize determinants on GPCR third cytoplasmic loops.

	GPCR Carboxyl-Terminal Interactions With PDZ Scaffold Proteins

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Rhodopsin, the light receptor, is a G protein–coupled receptor found in the retina. Accurate vision requires the rapid processing of visual images, and the cellular signaling pathways underlying vision therefore must be extremely fast. Given the unique demands of the visual system, it is perhaps no surprise that rhodopsin offers one of the clearest examples of G protein–coupled receptor signaling made more rapid and more efficient by means of an associated scaffold protein.

Drosophila rhodopsin physically associates with a scaffold protein named InaD.^11,12 InaD is a large cytoplasmic protein with five PSD-95/Discs-large/ZO-1 homology (PDZ) domains, which are specialized domains for mediating interactions with the carboxyl-termini of other proteins.⁹ Two of the InaD PDZ domains mediate the interaction with rhodopsin,¹² whereas the other domains mediate interactions with a variety of other proteins involved in visual signaling. Mammalian visual signaling is quite different from that of Drosophila, and the role of rhodopsin-associated scaffold proteins in mammalian visual signaling is unclear at present.

Stimulation of Drosophila rhodopsin promotes coupling to a G_q protein that activates an isoform of phospholipase C, leading to increased intracellular calcium and the activation of protein kinase C as well as the opening of a calcium-regulated channel known as TRP. Most of the components of this signaling pathway (rhodopsin, phospholipase C, protein kinase C, and the calcium channel TRP) have been found to associate with InaD^8,11–18 (Figure 2A).With all of these molecules bound to the same scaffold and located in close spatial proximity to each other, visual signaling is less reliant on the slow speed of diffusion and can proceed with lightning quickness. As proof of the physiological importance of the scaffolding function of InaD, it has been shown that Drosophila mutants lacking InaD exhibit visual signaling that is both reduced in magnitude and dramatically slowed relative to wild-type flies: the amplitudes of quantum bumps in response to singe photons of light by mutants lacking InaD are less than one-fifth of the amplitudes observed in control flies, and the latencies of visual responses are slowed by more than 6-fold in the InaD mutants.¹⁹ .

View larger version (18K): [in this window] [in a new window]   Figure 2. Prototypical GPCR/PDZ interactions. PDZ domain–containing scaffold proteins are shown in blue, with the PDZ domain regions indicated in gray. A, Multi-PDZ protein InaD can link Drosophila rhodopsin to key effectors, such as phospholipase C (PLC), protein kinase C (PKC), and the TRP calcium channel. B, Multi-PDZ protein PSD-95 can link the mammalian ß1-adrenergic receptor (ß1AR) to key effectors such as NMDA-type glutamate receptor channels. C, Multi-PDZ protein NHERF can bind to the ß2-adrenergic receptor (ß2AR) as well as to other proteins such as the Na+-H+ exchanger 3 (NHE3) and the actin-associated protein ezrin. D, PDZ protein tamalin can link various metabotropic glutamate receptor (mGluR) subtypes to cytoplasmic signaling proteins such as the ADP-ribosylation factor (ARF) nucleotide binding site opener (ARNO).

ß-Adrenergic receptors (ßARs) exist as three distinct subtypes and mediate physiological responses to epinephrine, a key hormone involved in the regulation of cardiovascular function in response to stress. Like Drosophila rhodopsin, mammalian ßARs can associate with PDZ domain–containing scaffold proteins. The ß₁-adrenergic receptor associates with two related PDZ proteins, postsynaptic density protein 95 (PSD-95)^20–22 and membrane-associated guanylate kinase-like protein inverted-2 (MAGI-2).²¹ The ß₂-adrenergic receptor, in contrast, does not associate with PSD-95 or MAGI-2, but rather associates specifically with two other PDZ proteins, the Na⁺-H⁺ exchanger regulatory factor proteins NHERF-1 and NHERF-2.^23–25

There is good evidence that the ßAR-associated PDZ proteins can function as scaffolds. The association of the ß₁AR with PSD-95²⁰ has been shown to physically link the ß₁AR to effectors such as the N-methyl-D-aspartate (NMDA) class of glutamate receptor channels, which are known to be regulated by ß₁AR stimulation in neurons^26–28 (Figure 2B). The agonist-promoted association of the ß₂AR with the NHERF proteins,²³ in contrast, facilitates regulation of the Na⁺-H⁺ exchanger type 3 (NHE3), a cell surface transporter that is inhibited by NHERF proteins²⁹ (Figure 2C). Increases in cellular cAMP typically inhibit NHE3 activity, but ß₂AR stimulation, which increases cellular cAMP, paradoxically enhances NHE3 activity.²⁹ This enhancement of NHE3 activity is blocked if the ß₂AR cannot bind NHERF,²³ suggesting that either ß₂AR association with NHERF prevents NHERF regulation of NHE3 or that association with NHERF links ß₂AR to a cellular signaling pathway important for regulation of Na⁺-H⁺ exchange.

ßAR associations with PDZ proteins are dependent on specialized motifs at the receptors’ carboxyl-termini: E-S/T-x-V in the case of the ß₁AR^20,21 and D-S/T-x-L in the case of the ß₂AR.^23,24 Interestingly, the associations of ß₁AR with PSD-95²² and ß₂AR with NHERF-1²⁵ have both been shown to be disrupted via receptor phosphorylation by G protein–coupled receptor kinase 5 (GRK5). PDZ domain interactions are often critically dependent on a serine or threonine at the -2 position of the target protein’s carboxyl-terminus,⁹ and it is therefore possible that many G protein–coupled receptor interactions with PDZ proteins will be regulated by GRK5 phosphorylation. Phosphorylation-dependent regulation of receptor association with scaffolds represents a mechanism by which cellular responsiveness to certain hormones, neurotransmitters, and other stimuli can be rapidly and reversibly fine-tuned.

Glutamate is the primary excitatory neurotransmitter in the mammalian central nervous system, and many of glutamate’s physiological actions are mediated through a family of G protein–coupled receptors known as metabotropic glutamate receptors (mGluRs). The carboxyl-termini of metabotropic glutamate receptors can associate with a variety of PDZ domain–containing scaffold proteins. For example, mGluR1 and mGluR5 can interact directly with the PDZ domain of Shank proteins,³⁰ whereas mGluR1, mGluR2, mGluR3, and mGluR5 have been shown to associate with the PDZ protein tamalin³¹ (Figure 2D). The motif S-S/T-L at the mGluR carboxyl-terminus is critical for both of these interactions. Tamalin is a cellular binding partner of the ADP-ribosylation factor (ARF) nucleotide binding site opener (ARNO), and has been shown to act as a scaffold to facilitate the physical linkage of mGluRs to ARNO in cells.³¹ Finally, mGluR7 associates specifically with a PDZ protein referred to as the protein that interacts with C-kinase (PICK1).^32–35 The mGluR7 carboxyl-terminus ends in the motif L-V-I, which is critical for the interaction with PICK1^32,33 and is quite distinct from the mGluR1/2/3/5 motif that mediates interaction with tamalin. PICK1 has been shown to link mGluR7 to protein kinase C in cells, revealing a scaffolding function for this interaction.³³

	GPCR Carboxyl-Terminal Interactions With Non-PDZ Scaffold Proteins

Top Abstract Introduction GPCR Carboxyl-Terminal... GPCR Carboxyl-Terminal... GPCR Interactions With... Scaffold Versus Nonscaffold... Agonist-Dependence of... Problems in Studying... Concluding Thoughts References

 
The family of A-kinase anchoring proteins (AKAPs) was one of the first classes of proteins to be recognized as scaffold proteins.³⁶ These proteins associate with protein kinase A (PKA) and a variety of other proteins to organize cellular signaling pathways. ß-Adrenergic receptors and many other G protein–coupled receptors can couple to G_s to increase cAMP and activate PKA downstream, and thus, the association of AKAPs with G protein–coupled receptors would seem to be an attractive potential mechanism for enhancing the efficiency of G_s-mediated signaling. Two different AKAPs have been found to interact with ß-adrenergic receptors. AKAP250, also known as gravin, has been reported to bind to the ß₂AR carboxyl-terminus, promoting receptor association with PKA and regulating receptor desensitization (Figure 3A).^37,38 AKAP79 has also been shown to interact with the ß₂AR, promoting ß₂AR phosphorylation and downstream mitogenic signaling.^39,40 The AKAP-binding motif on the ß₂AR carboxyl-terminus has not been defined in detail, and thus it is not clear at present if interaction with AKAPs is unique to the ß₂AR or if other GPCRs may also associate with certain AKAPs to organize signaling pathways involving PKA.

View larger version (17K): [in this window] [in a new window]   Figure 3. Prototypical GPCR interactions with non-PDZ scaffolds. A, AKAPs such as gravin and AKAP79 can link the ß2-adrenergic receptor (ß2AR) to protein kinase A. B, Jak2 can facilitate the association of angiotensin AT1 receptors with STAT1. C, Scaffold protein Homer can couple various metabotropic glutamate receptor (mGluR) subtypes to the inositol triphosphate receptor (IP3R), which is found in the endoplasmic reticulum (ER). D, ß-Arrestins (ßArr) associate with many GPCRs, disrupting G-protein coupling and also acting as scaffold proteins to facilitate multiple interactions between GPCRs and cytoplasmic proteins.

Angiotensin II is a potent hemodynamic regulator that exerts most of its physiological actions through a receptor known as AT₁. Stimulation of the AT₁ receptor has been found to activate not only traditional G-protein pathways but also the Janus kinase (Jak)-signal transducers and activators of transcription (STAT) signaling pathway,^41–52 which is typically activated by cytokine or growth factor receptors but not by GPCRs. Jaks are tyrosine kinases and STATs are transcription factors that can shuttle between the cytoplasm and the nucleus to regulate the expression of various genes.⁵³ The ability of the AT₁ receptor to regulate Jak/STAT signaling has been found to be dependent on a direct interaction between the AT₁R and Jak2.^45,49,51,52 Association of Jak2 with the AT₁R not only promotes Jak2 phosphorylation of STAT1, but also leads to recruitment of STAT1 into a complex with AT₁R,^51,52 revealing a function of Jak2 as a scaffold protein (Figure 3B). The interaction of Jak2 with the AT₁R is dependent on a specialized motif (Y-I-P-P) found in the AT₁R carboxyl-terminus⁴⁵ but not present in the carboxyl-termini of most other GPCRs.

The metabotropic glutamate receptor subtypes mGluR1 and mGluR5 have been found to associate via a specialized carboxyl-terminal motif (P-P-x-x-F-R) with the Homer family of proteins.^54–64 Homer 1b, 2, and 3 contain coiled-coil domains that mediate Homer multimerization, whereas Homer 1a is an immediate early gene that does not contain coiled-coil domains and thus can disrupt Homer 1b, 2, and 3 multimeric complexes when its expression is induced.^55,57 Association with Homer proteins has significant consequences for mGluR1 and mGluR5 function, and many of these effects are likely due to actions of Homer proteins as scaffolds (Figure 3C). Homer proteins can bind to a variety of other proteins beyond themselves and mGluRs, including intracellular inositol triphosphate (IP3) receptors,⁵⁶ syntaxin 13,⁶⁵ and the Shank family of scaffold proteins.³⁰ Because mGluR1 and mGluR5 are both G_q-coupled receptors that lead to increased cellular levels of IP3 when stimulated, the Homer-dependent linkage of these receptors to IP3 receptors may be especially important for regulating their signaling.⁵⁶ Homer proteins also exert a strong effect on regulating the level of constitutive G-protein coupling to mGluR1 and mGluR5: association with Homer3 dampens down mGluR1/5 constitutive activity, whereas expression of Homer1a enhances constitutive activity of mGluR1 and mGluR5, probably by disrupting mGluR1/5 association with Homer3.⁶³ It is not certain at present if these effects on mGluR constitutive activity are dependent on the action of Homer proteins as scaffolds or if they are due instead to direct allosteric effects of Homer proteins on receptor conformation.

	GPCR Interactions With ß-Arrestins

Top Abstract Introduction GPCR Carboxyl-Terminal... GPCR Carboxyl-Terminal... GPCR Interactions With... Scaffold Versus Nonscaffold... Agonist-Dependence of... Problems in Studying... Concluding Thoughts References

 
All of the GPCR/scaffold interactions discussed thus far are dependent on the presence of specialized motifs in the GPCR carboxyl-termini. The requirement of a defined motif for receptor/scaffold interaction is of interest, because it reveals a molecular mechanism by which different subtypes of receptors may be specifically linked to different intracellular effectors via differential association with scaffold proteins. However, the requirement of a defined motif for scaffold interaction with GPCRs also limits the potential generality of each GPCR/scaffold interaction, because only a small number of GPCRs are likely to possess a specialized motif required for interaction with a particular scaffold protein.

The exception to the rule of specificity in GPCR/scaffold interactions is the ß-arrestin family of proteins. ß-Arrestin1 and ß-arrestin2 were first identified as proteins involved in the desensitization of ß-adrenergic receptors,^66,67 but it is now known that ß-arrestins can associate with the majority of G protein–coupled receptors. Agonist activation of most G protein–coupled receptors results in receptor phosphorylation by GRKs, which promotes receptor association with ß-arrestins and receptor uncoupling from G proteins.⁶⁸ Key determinants for interaction with ß-arrestins are found in the third cytoplasmic loops of many GPCRs, although determinants in other intracellular GPCR regions may also contribute to ß-arrestin association.⁶⁸ ß-arrestins are known to associate with proteins involved in endocytosis such as clathrin,⁶⁹ AP-2,^70,71 N-ethylmaleimide–sensitive factor (NSF),⁷² ARF6,⁷³ ARNO,⁷³ and Mdm2,⁷⁴ and these interactions facilitate agonist-promoted GPCR internalization into clathrin-coated pits. Thus, ß-arrestins act as scaffolds to physically link G protein–coupled receptors to the endocytic machinery inside the cell. Unlike all of the other interactions described above, the interactions of ß-arrestins with G protein–coupled receptors are not dependent on a specialized motif unique to one or several receptors, and it is therefore likely that ß-arrestins act as scaffold proteins for most G protein–coupled receptors (Figure 3D).

ß-Arrestins can associate with a variety of proteins other than G protein–coupled receptors and endocytic proteins. For example, ß-arrestin1 can associate with the tyrosine kinase Src.^75,76 ß-Arrestin–dependent recruitment of Src plays a key role in mitogenic signaling by the ß₂-adrenergic receptor⁷⁵ as well as in mitogenic signaling by other receptors such as neurokinin receptors,⁷⁷ interleukin receptors,⁷⁸ protease-activated receptors,⁷⁹ and endothelin receptors.⁸⁰ In light of these findings, ß-arrestins are no longer viewed simply as proteins involved in G protein–coupled receptor desensitization, but rather as multipurpose scaffolds that can turn off some receptor-initiated signaling pathways while simultaneously activating other pathways.

Other proteins known to associate with ß-arrestins include members of two distinct mitogen activated kinase (MAP) kinase cascades, c-Jun N-terminal kinase 3 (JNK3) and extracellular signal regulated kinases 1 and 2 (ERK1/2). In the case of JNK3, ß-arrestin2, but not ß-arrestin1, is a high-affinity binding partner of both JNK3 and apoptosis-stimulating kinase 1 (ASK1), a kinase upstream of JNK activation.^81,82 JNK3 activity is potently regulated by ß-arrestin2 as well as by the recruitment of ß-arrestin2 to activated angiotensin AT₁ receptors.⁸¹ In the case of ERK1/2, both ß-arrestins appear to be capable of scaffolding the ERKs in close proximity to various G protein–coupled receptors, facilitating receptor-mediated ERK activation.^77,79,83 Thus, ß-arrestin1 and ß-arrestin2 can have differential scaffolding activities with very different consequences for G protein–coupled receptor signaling.

	Scaffold Versus Nonscaffold Actions of Receptor-Associated Proteins

Top Abstract Introduction GPCR Carboxyl-Terminal... GPCR Carboxyl-Terminal... GPCR Interactions With... Scaffold Versus Nonscaffold... Agonist-Dependence of... Problems in Studying... Concluding Thoughts References

 
As mentioned earlier, a protein may be considered a scaffold if it associates with two or more members of a signaling pathway to help increase the efficiency and/or specificity of the pathway. Not all receptor-associated proteins act as scaffolds, and even for those that do, it can often be difficult to determine whether a particular effect of the associated protein on receptor function is due to a scaffolding action or not. For example, the interactions of both ß₁AR and ß₂AR with their PDZ binding partners have effects on receptor internalization,^20,21,25 and the interactions of mGluR1 and mGluR5 with Homer proteins have significant consequences for receptor trafficking^{52,53,55,56,59} and coupling to G proteins.⁵⁸ However, it is unclear if these effects are due to (1) a true scaffolding function of the receptor-associated proteins, (2) a direct allosteric action of the receptor-associated proteins on receptor conformation, or (3) a combination of scaffolding effects and direct allosteric effects. It is likely that many receptor-associated proteins act simultaneously as both scaffolds and direct allosteric modulators of receptor function, so these actions can therefore be difficult to tease apart experimentally.

	Agonist-Dependence of Receptor/Scaffold Interactions

Top Abstract Introduction GPCR Carboxyl-Terminal... GPCR Carboxyl-Terminal... GPCR Interactions With... Scaffold Versus Nonscaffold... Agonist-Dependence of... Problems in Studying... Concluding Thoughts References

 
One of the most interesting questions to ask about scaffold proteins associated with G protein–coupled receptors is whether or not their interactions with receptors are regulated by agonist stimulation. It is well-known that agonist binding causes significant alterations in G protein–coupled receptor conformation, leading to profoundly enhanced association of receptor intracellular domains with G proteins.¹⁰ Of the receptor/scaffold interactions discussed, several have been found to be enhanced by agonist stimulation, including ß₂AR/NHERF, ß₁AR/MAGI-2, AT₁R/Jak2, and the interaction of many receptors with ß-arrestins. In contrast, there is no evidence at present for agonist regulation of some of the other receptor/scaffold interactions discussed, including rhodopsin/InaD, ß₂AR/AKAP, ß₁AR/PSD-95, mGluR/Homer, and mGluR/PDZ. The extent of agonist regulation of a receptor/scaffold interaction probably plays a significant role in determining the way in which the scaffold protein can regulate receptor signaling, especially in determining the temporal characteristics of the regulation.

	Problems in Studying Receptor/Scaffold Interactions

Top Abstract Introduction GPCR Carboxyl-Terminal... GPCR Carboxyl-Terminal... GPCR Interactions With... Scaffold Versus Nonscaffold... Agonist-Dependence of... Problems in Studying... Concluding Thoughts References

 
There are several problems inherent in studying the biology of scaffold proteins. One problem is that measurements of the function of such proteins must always be indirect, because scaffold proteins are defined not by any intrinsic activity of their own but rather by their ability to enhance the interactions of other proteins. The activities of G proteins can be accurately assessed by measuring their GTPase activity, and the activities of kinases can be accurately quantified by measuring their phosphorylation of a substrate; unfortunately, however, there is no simple assay for the quantification of scaffolding activity. Rough estimates of scaffold protein efficiency can be derived only for signaling systems as a whole, examined in the absence or presence of a given scaffold protein. The speed of a signaling pathway is probably the easiest parameter to quantify, as in the case of rhodopsin signaling in the absence or presence of InaD.¹⁹

A second problem inherent in studying scaffold proteins is that the signaling pathways they organize can be very complex. For example, the PDZ domain-containing scaffold protein PSD-95 is known to associate with at least 50 distinct signaling proteins.⁹ Studying the association of even two proteins in isolation can be extremely complicated, and studying the association of more than two proteins introduces numerous extra layers of complexity with each protein that is added into the mix. Scaffolds bind to multiple proteins, by definition, and to fully understand the function of a scaffold protein, it is important to understand in detail such issues as whether or not the various binding partners can bind to the scaffold simultaneously in cells. If the partners cannot all bind simultaneously, it is important to understand the sequence of binding events and the time course of each association, as well as how the binding of one protein might influence the binding or dissociation of all the other proteins. Questions such as these can be difficult to address experimentally in a quantitative fashion. Recent advances in real-time imaging of multiprotein cellular signaling complexes are likely to be very helpful in helping to answer such questions about the cellular functions of scaffold proteins.

	Concluding Thoughts

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Agonist-stimulated G protein–coupled receptors initiate cellular signaling pathways involving multiple components. Some G protein–coupled receptors have been found to associate with scaffold proteins, which also interact with components involved in downstream receptor signaling. These scaffold proteins can have significant effects on the efficiency of receptor signaling. The differential distributions of various scaffold proteins may help to explain why certain receptors exhibit differential activity in distinct tissues. G protein–coupled receptors are extremely common targets for therapeutic pharmaceuticals in the treatment of cardiovascular diseases and other disorders, and disruption of receptor interactions with scaffold proteins represents an intriguing potential therapeutic approach to the treatment of various disease states.

	Acknowledgments

 
R.A.H. is supported by grants HL64713 and GM60982 from the NIH and by a Distinguished Young Investigator in Medical Sciences award from the W.M. Keck Foundation. R.J.L. is supported by grants HL16037 and HL70631 from the NIH and is an investigator of the Howard Hughes Medical Institute.

Received April 5, 2002; revision received August 27, 2002; accepted August 28, 2002.

	References

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1. Hall RA, Premont RT, Lefkowitz RJ. G protein–coupled receptor signaling: beyond the G protein paradigm. J Cell Biol. 1999; 145: 927–932.[Free Full Text]
2. Heuss C, Gerber U. G-protein–independent signaling by G-protein–coupled receptors. Trends Neurosci. 2000; 23: 469–475.[CrossRef][Medline] [Order article via Infotrieve]
3. Brzostowski JA, Kimmel AR. Signaling at zero G: G-protein–independent functions for 7-TM receptors. Trends Biochem Sci. 2001; 26: 291–297.[CrossRef][Medline] [Order article via Infotrieve]
4. Milligan G, White JH. Protein-protein interactions at G-protein–coupled receptors. Trends Pharmacol Sci. 2001; 22: 513–518.[CrossRef][Medline] [Order article via Infotrieve]
5. Pawson T, Scott JD. Signaling through scaffold, anchoring, and adaptor proteins. Science. 1997; 278: 2075–2080.[Abstract/Free Full Text]
6. Whitmarsh AJ, Davis RJ. Structural organization of MAP-kinase signaling modules by scaffold proteins in yeast and mammals. Trends Biochem Sci. 1998; 23: 481–485.[CrossRef][Medline] [Order article via Infotrieve]
7. Burack WR, Shaw AS. Signal transduction: hanging on a scaffold. Curr Opin Cell Biol. 2000; 12: 211–216.[CrossRef][Medline] [Order article via Infotrieve]
8. Huber A. Scaffolding proteins organize multimolecular protein complexes for sensory signal transduction. Eur J Neurosci. 2001; 14: 769–776.[CrossRef][Medline] [Order article via Infotrieve]
9. Sheng M, Sala C. PDZ domains and the organization of supramolecular complexes. Annu Rev Neurosci. 2001; 24: 1–29.[CrossRef][Medline] [Order article via Infotrieve]
10. Dohlman HG, Thorner J, Caron MG, Lefkowitz RJ. Model systems for the study of seven-transmembrane-segment receptors. Annu Rev Biochem. 1991; 60: 653–688.[CrossRef][Medline] [Order article via Infotrieve]
11. Chevesich J, Kreuz AJ, Montell C. Requirement for the PDZ domain protein, INAD, for localization of the TRP store-operated channel to a signaling complex. Neuron. 1997; 18: 95–105.[CrossRef][Medline] [Order article via Infotrieve]
12. Xu XZ, Choudhury A, Li X, Montell C. Coordination of an array of signaling proteins through homo- and heteromeric interactions between PDZ domains and target proteins. J Cell Biol. 1998; 142: 545–555.[Abstract/Free Full Text]
13. Huber A, Sander P, Gobert A, Bahner M, Hermann R, Paulsen R. The transient receptor potential protein (Trp), a putative store-operated Ca²⁺ channel essential for phosphoinositide-mediated photoreception, forms a signaling complex with NorpA, InaC and InaD. EMBO J. 1996; 15: 7036–7045.[Medline] [Order article via Infotrieve]
14. Tsunoda S, Sierralta J, Sun Y, Bodner R, Suzuki E, Becker A, Socolich M, Zuker CS. A multivalent PDZ-domain protein assembles signalling complexes in a G-protein–coupled cascade. Nature. 1997; 388: 243–249.[CrossRef][Medline] [Order article via Infotrieve]
15. van Huizen R, Miller K, Chen DM, Li Y, Lai ZC, Raab RW, Stark WS, Shortridge RD, Li M. Two distantly positioned PDZ domains mediate multivalent INAD-phospholipase C interactions essential for G protein–coupled signaling. EMBO J. 1998; 17: 2285–2297.[CrossRef][Medline] [Order article via Infotrieve]
16. Bahner M, Sander P, Paulsen R, Huber A. The visual G protein of fly photoreceptors interacts with the PDZ domain assembled INAD signaling complex via direct binding of activated G_q to phospholipase cß. J Biol Chem. 2000; 275: 2901–2904.[Abstract/Free Full Text]
17. Li HS, Montell C. TRP and the PDZ protein, INAD, form the core complex required for retention of the signalplex in Drosophila photoreceptor cells. J Cell Biol. 2000; 150: 1411–1122.[Abstract/Free Full Text]
18. Tsunoda S, Sun Y, Suzuki E, Zuker C. Independent anchoring and assembly mechanisms of INAD signaling complexes in Drosophila photoreceptors. J Neurosci. 2001; 21: 150–158.[Abstract/Free Full Text]
19. Scott K, Zuker CS. Assembly of the Drosophila phototransduction cascade into a signalling complex shapes elementary responses. Nature. 1998; 395: 805–808.[CrossRef][Medline] [Order article via Infotrieve]
20. Hu LA, Tang Y, Miller WE, Cong M, Lau AG, Lefkowitz RJ, Hall RA. ß₁-Adrenergic receptor association with PSD-95: inhibition of receptor internalization and facilitation of ß₁-adrenergic receptor interaction withN-methyl-D-aspartate receptors. J Biol Chem. 2000; 275: 38659–38666.[Abstract/Free Full Text]
21. Xu J, Paquet M, Lau AG, Wood JD, Ross CA, Hall RA. ß₁-Adrenergic receptor association with the synaptic scaffolding protein membrane-associated guanylate kinase inverted-2 (MAGI-2): differential regulation of receptor internalization by MAGI-2 and PSD-95. J Biol Chem. 2001; 276: 41310–41317.[Abstract/Free Full Text]
22. Hu LA, Chen W, Premont RT, Cong M, Lefkowitz RJ. G Protein–coupled receptor kinase 5 regulates ß₁-adrenergic receptor association with PSD-95. J Biol Chem. 2002; 277: 1607–1613.[Abstract/Free Full Text]
23. Hall RA, Premont RT, Chow CW, Blitzer JT, Pitcher JA, Claing A, Stoffel RH, Barak LS, Shenolikar S, Weinman EJ, Grinstein S, Lefkowitz RJ. The ß₂-adrenergic receptor interacts with the Na⁺/H⁺-exchanger regulatory factor to control Na⁺/H⁺ exchange. Nature. 1998; 392: 626–630.[CrossRef][Medline] [Order article via Infotrieve]
24. Hall RA, Ostedgaard LS, Premont RT, Blitzer JT, Rahman N, Welsh MJ, Lefkowitz RJ. A C-terminal motif found in the ß₂-adrenergic receptor, P2Y1 receptor and cystic fibrosis transmembrane conductance regulator determines binding to the Na⁺/H⁺ exchanger regulatory factor family of PDZ proteins. Proc Natl Acad Sci U S A. 1998; 95: 8496–8501.[Abstract/Free Full Text]
25. Cao TT, Deacon HW, Reczek D, Bretscher A, von Zastrow M. A kinase-regulated PDZ-domain interaction controls endocytic sorting of the ß₂-adrenergic receptor. Nature. 1999; 401: 286–290.[CrossRef][Medline] [Order article via Infotrieve]
26. Gean PW, Huang CC, Lin JH, Tsai JJ. Sustained enhancement of NMDA receptor-mediated synaptic potential by isoproterenol in rat amygdalar slices. Brain Res. 1992; 594: 331–334.[CrossRef][Medline] [Order article via Infotrieve]
27. Huang CC, Tsai JJ, Gean PW. Enhancement of NMDA receptor-mediated synaptic potential by isoproterenol is blocked by Rp-adenosine 3',5'-cyclic monophosphothioate. Neurosci Lett. 1993; 161: 207–210.[CrossRef][Medline] [Order article via Infotrieve]
28. Raman IM, Tong G, Jahr CE. ß-Adrenergic regulation of synaptic NMDA receptors by cAMP-dependent protein kinase. Neuron. 1996; 16: 415–421.[CrossRef][Medline] [Order article via Infotrieve]
29. Weinman EJ, Minkoff C, Shenolikar S. Signal complex regulation of renal transport proteins: NHERF and regulation of NHE3 by PKA. Am J Physiol Renal Physiol. 2000; 279: F393–F399.[Abstract/Free Full Text]
30. Tu JC, Xiao B, Naisbitt S, Yuan JP, Petralia RS, Brakeman P, Doan A, Aakalu VK, Lanahan AA, Sheng M, Worley PF. Coupling of mGluR/Homer and PSD-95 complexes by the Shank family of postsynaptic density proteins. Neuron. 1999; 23: 583–592.[CrossRef][Medline] [Order article via Infotrieve]
31. Kitano J, Kimura K, Yamazaki Y, Soda T, Shigemoto R, Nakajima Y, Nakanishi S. Tamalin, a PDZ domain-containing protein, links a protein complex formation of group 1 metabotropic glutamate receptors and the guanine nucleotide exchange factor cytohesins. J Neurosci.;. 2002; 22: 1280–1289.[Abstract/Free Full Text]
32. Boudin H, Doan A, Xia J, Shigemoto R, Huganir RL, Worley P, Craig AM. Presynaptic clustering of mGluR7a requires the PICK1 PDZ domain binding site. Neuron. 2000; 28: 485–497.[CrossRef][Medline] [Order article via Infotrieve]
33. Dev KK, Nakajima Y, Kitano J, Braithwaite SP, Henley JM, Nakanishi S. PICK1 interacts with and regulates PKC phosphorylation of mGLUR7. J Neurosci. 2000; 20: 7252–7257.[Abstract/Free Full Text]
34. El Far O, Airas J, Wischmeyer E, Nehring RB, Karschin A, Betz H. Interaction of the C-terminal tail region of the metabotropic glutamate receptor 7 with the protein kinase C substrate PICK1. Eur J Neurosci. 2000; 12: 4215–4221.[CrossRef][Medline] [Order article via Infotrieve]
35. Boudin H, Craig AM. Molecular determinants for PICK1 synaptic aggregation and mGluR7a receptor coclustering: role of the PDZ, coiled-coil, and acidic domains. J Biol Chem. 2001; 276: 30270–30276.[Abstract/Free Full Text]
36. Colledge M, Scott JD. AKAPs: from structure to function. Trends Cell Biol. 1999; 9: 216–221.[CrossRef][Medline] [Order article via Infotrieve]
37. Shih M, Lin F, Scott JD, Wang HY, Malbon CC. Dynamic complexes of ß₂-adrenergic receptors with protein kinases and phosphatases and the role of gravin. J Biol Chem. 1999; 274: 1588–1595.[Abstract/Free Full Text]
38. Fan G, Shumay E, Wang H, Malbon CC. The scaffold protein gravin (cAMP-dependent protein kinase-anchoring protein 250) binds the ß2-adrenergic receptor via the receptor cytoplasmic Arg-329 to Leu-413 domain and provides a mobile scaffold during desensitization. J Biol Chem. 2001; 276: 24005–24014.[Abstract/Free Full Text]
39. Fraser ID, Cong M, Kim J, Rollins EN, Daaka Y, Lefkowitz RJ, Scott JD. Assembly of an A kinase-anchoring protein-ß₂-adrenergic receptor complex facilitates receptor phosphorylation and signaling. Curr Biol. 2000; 10: 409–412.[CrossRef][Medline] [Order article via Infotrieve]
40. Cong M, Perry SJ, Lin FT, Fraser ID, Hu LA, Chen W, Pitcher JA, Scott JD, Lefkowitz RJ. Regulation of membrane targeting of the G protein–coupled receptor kinase 2 by protein kinase A and its anchoring protein AKAP79. J Biol Chem. 2001; 276: 15192–15199.[Abstract/Free Full Text]
41. Bhat GJ, Thekkumkara TJ, Thomas WG, Conrad KM, Baker KM. Angiotensin II stimulates sis-inducing factor-like DNA binding activity: evidence that the AT1A receptor activates transcription factor-Stat91 and/or a related protein. J Biol Chem. 1994; 269: 31443–31449.[Abstract/Free Full Text]
42. Bhat GJ, Thekkumkara TJ, Thomas WG, Conrad KM, Baker KM. Activation of the STAT pathway by angiotensin II in T3CHO/AT1A cells: cross-talk between angiotensin II and interleukin-6 nuclear signaling. J Biol Chem. 1995; 270: 19059–19065.[Abstract/Free Full Text]
43. Marrero MB, Schieffer B, Paxton WG, Heerdt L, Berk BC, Delafontaine P, Bernstein KE. Direct stimulation of Jak/STAT pathway by the angiotensin II AT1 receptor. Nature. 1995; 375: 247–250.[CrossRef][Medline] [Order article via Infotrieve]
44. Marrero MB, Paxton WG, Schieffer B, Ling BN, Bernstein KE. Angiotensin II signalling events mediated by tyrosine phosphorylation. Cell Signal. 1996; 8: 21–26.[CrossRef][Medline] [Order article via Infotrieve]
45. Ali MS, Sayeski PP, Dirksen LB, Hayzer DJ, Marrero MB, Bernstein KE. Dependence on the motif YIPP for the physical association of Jak2 kinase with the intracellular carboxyl tail of the angiotensin II AT1 receptor. J Biol Chem. 1997; 272: 23382–23388.[Abstract/Free Full Text]
46. McWhinney CD, Hunt RA, Conrad KM, Dostal DE, Baker KM. The type I angiotensin II receptor couples to Stat1 and Stat3 activation through Jak2 kinase in neonatal rat cardiac myocytes. J Mol Cell Cardiol. 1997; 29: 2513–2524.[CrossRef][Medline] [Order article via Infotrieve]
47. Pan J, Fukuda K, Kodama H, Makino S, Takahashi T, Sano M, Hori S, Ogawa S. Role of angiotensin II in activation of the JAK/STAT pathway induced by acute pressure overload in the rat heart. Circ Res. 1997; 81: 611–617.[Abstract/Free Full Text]
48. Kodama H, Fukuda K, Pan J, Makino S, Sano M, Takahashi T, Hori S, Ogawa S. Biphasic activation of the JAK/STAT pathway by angiotensin II in rat cardiomyocytes. Circ Res. 1998; 82: 244–250.[Abstract/Free Full Text]
49. Marrero MB, Venema VJ, Ju H, Eaton DC, Venema RC. Regulation of angiotensin II-induced JAK2 tyrosine phosphorylation: roles of SHP-1 and SHP-2. Am J Physiol. 1998; 275: C1216–C1223.[Medline] [Order article via Infotrieve]
50. McWhinney CD, Dostal D, Baker K. Angiotensin II activates Stat5 through Jak2 kinase in cardiac myocytes. J Mol Cell Cardiol. 1998; 30: 751–761.[CrossRef][Medline] [Order article via Infotrieve]
51. Ali MS, Sayeski PP, Bernstein KE. Jak2 acts as both a STAT1 kinase and as a molecular bridge linking STAT1 to the angiotensin II AT1 receptor. J Biol Chem. 2000; 275: 15586–15593.[Abstract/Free Full Text]
52. Sayeski PP, Ali MS, Frank SJ, Bernstein KE. The angiotensin II–dependent nuclear translocation of Stat1 is mediated by the Jak2 protein motif 231YRFRR. J Biol Chem. 2001; 276: 10556–10563.[Abstract/Free Full Text]
53. Schindler C, Darnell JE Jr. Transcriptional responses to polypeptide ligands: the JAK-STAT pathway. Annu Rev Biochem. 1995; 64: 621–651.[Medline] [Order article via Infotrieve]
54. Brakeman PR, Lanahan AA, O’Brien R, Roche K, Barnes CA, Huganir RL, Worley PF. Homer: a protein that selectively binds metabotropic glutamate receptors. Nature. 1997; 386: 284–288.[CrossRef][Medline] [Order article via Infotrieve]
55. Kato A, Ozawa F, Saitoh Y, Fukazawa Y, Sugiyama H, Inokuchi K. Novel members of the Vesl/Homer family of PDZ proteins that bind metabotropic glutamate receptors. J Biol Chem. 1998; 273: 23969–23975.[Abstract/Free Full Text]
56. Tu JC, Xiao B, Yuan JP, Lanahan AA, Leoffert K, Li M, Linden DJ, Worley PF. Homer binds a novel proline-rich motif and links group 1 metabotropic glutamate receptors with IP3 receptors. Neuron. 1998; 21: 717–726.[CrossRef][Medline] [Order article via Infotrieve]
57. Xiao B, Tu JC, Petralia RS, Yuan JP, Doan A, Breder CD, Ruggiero A, Lanahan AA, Wenthold RJ, Worley PF. Homer regulates the association of group 1 metabotropic glutamate receptors with multivalent complexes of homer-related, synaptic proteins. Neuron. 1998; 21: 707–716.[CrossRef][Medline] [Order article via Infotrieve]
58. Ciruela F, Soloviev MM, McIlhinney RA. Co-expression of metabotropic glutamate receptor type 1 with homer-1a/Vesl-1S increases the cell surface expression of the receptor. Biochem J. 1999; 341: 795–803.[CrossRef][Medline] [Order article via Infotrieve]
59. Roche KW, Tu JC, Petralia RS, Xiao B, Wenthold RJ, Worley PF. Homer 1b regulates the trafficking of group I metabotropic glutamate receptors. J Biol Chem. 1999; 274: 25953–25957.[Abstract/Free Full Text]
60. Ango F, Pin JP, Tu JC, Xiao B, Worley PF, Bockaert J, Fagni L. Dendritic and axonal targeting of type 5 metabotropic glutamate receptor is regulated by homer1 proteins and neuronal excitation. J Neurosci. 2000; 20: 8710–8716.[Abstract/Free Full Text]
61. Ciruela F, Soloviev MM, Chan WY, McIlhinney RA. Homer-1c/Vesl-1L modulates the cell surface targeting of metabotropic glutamate receptor type 1: evidence for an anchoring function. Mol Cell Neurosci. 2000; 15: 36–50.[CrossRef][Medline] [Order article via Infotrieve]
62. Kammermeier PJ, Xiao B, Tu JC, Worley PF, Ikeda SR. Homer proteins regulate coupling of group I metabotropic glutamate receptors to N-type calcium and M-type potassium channels. J Neurosci. 2000; 20: 7238–7245.[Abstract/Free Full Text]
63. Ango F, Prezeau L, Muller T, Tu JC, Xiao B, Worley PF, Pin JP, Bockaert J, Fagni L. Agonist-independent activation of metabotropic glutamate receptors by the intracellular protein Homer. Nature. 2001; 411: 962–965.[CrossRef][Medline] [Order article via Infotrieve]
64. Coutinho V, Kavanagh I, Sugiyama H, Tones MA, Henley JM. Characterization of a metabotropic glutamate receptor type 5-green fluorescent protein chimera (mGluR5-GFP): pharmacology, surface expression, and differential effects of Homer-1a and Homer-1c. Mol Cell Neurosci. 2001; 18: 296–306.[CrossRef][Medline] [Order article via Infotrieve]
65. Minakami R, Kato A, Sugiyama H. Interaction of Vesl-1L/Homer 1c with syntaxin 13. Biochem Biophys Res Commun. 2000; 272: 466–471.[CrossRef][Medline] [Order article via Infotrieve]
66. Lohse MJ, Benovic JL, Codina J, Caron MG, Lefkowitz RJ. ß-Arrestin: a protein that regulates ß-adrenergic receptor function. Science. 1990; 248: 1547–1550.[Abstract/Free Full Text]
67. Attramadal H, Arriza JL, Aoki C, Dawson TM, Codina J, Kwatra MM, Snyder SH, Caron MG, Lefkowitz RJ. ß-Arrestin2, a novel member of the arrestin/ß-arrestin gene family. J Biol Chem. 1992; 267: 17882–17890.[Abstract/Free Full Text]
68. Krupnick JG, Benovic JL. The role of receptor kinases and arrestins in G protein–coupled receptor regulation. Annu Rev Pharmacol Toxicol. 1998; 38: 289–319.[CrossRef][Medline] [Order article via Infotrieve]
69. Goodman OB Jr, Krupnick JG, Santini F, Gurevich VV, Penn RB, Gagnon AW, Keen JH, Benovic JL. ß-Arrestin acts as a clathrin adaptor in endocytosis of the ß₂-adrenergic receptor. Nature. 1996; 383: 447–450.[CrossRef][Medline] [Order article via Infotrieve]
70. Laporte SA, Oakley RH, Zhang J, Holt JA, Ferguson SS, Caron MG, Barak LS. The ß₂-adrenergic receptor/ß-arrestin complex recruits the clathrin adaptor AP-2 during endocytosis. Proc Natl Acad Sci U S A. 1999; 96: 3712–3717.[Abstract/Free Full Text]
71. Laporte SA, Oakley RH, Holt JA, Barak LS, Caron MG. The interaction of ß-arrestin with the AP-2 adaptor is required for the clustering of ß₂-adrenergic receptor into clathrin-coated pits. J Biol Chem. 2000; 275: 23120–23126.[Abstract/Free Full Text]
72. McDonald PH, Cote NL, Lin FT, Premont RT, Pitcher JA, Lefkowitz RJ. Identification of NSF as a ß-arrestin1-binding protein: implications for ß₂-adrenergic receptor regulation. J Biol Chem. 1999; 274: 10677–10680.[Abstract/Free Full Text]
73. Claing A, Chen W, Miller WE, Vitale N, Moss J, Premont RT, Lefkowitz RJ. ß-Arrestin–mediated ADP-ribosylation factor 6 activation and ß₂-adrenergic receptor endocytosis. J Biol Chem.;. 2001; 276: 42509–42513.[Abstract/Free Full Text]
74. Shenoy SK, McDonald PH, Kohout TA, Lefkowitz RJ. Regulation of receptor fate by ubiquitination of activated ß₂-adrenergic receptor and ß-arrestin. Science. 2001; 294: 1307–1313.[Abstract/Free Full Text]
75. Luttrell LM, Ferguson SS, Daaka Y, Miller WE, Maudsley S, Della Rocca GJ, Lin F, Kawakatsu H, Owada K, Luttrell DK, Caron MG, Lefkowitz RJ. ß-Arrestin–dependent formation of ß₂ adrenergic receptor-Src protein kinase complexes. Science. 1999; 283: 655–661.[Abstract/Free Full Text]
76. Miller WE, Maudsley S, Ahn S, Khan KD, Luttrell LM, Lefkowitz RJ. ß-Arrestin1 interacts with the catalytic domain of the tyrosine kinase c-SRC: role of ß-arrestin1–dependent targeting of c-SRC in receptor endocytosis. J Biol Chem. 2000; 275: 11312–11319.[Abstract/Free Full Text]
77. DeFea KA, Vaughn ZD, O’Bryan EM, Nishijima D, Dery O, Bunnett NW. The proliferative and antiapoptotic effects of substance P are facilitated by formation of a ß-arrestin–dependent scaffolding complex. Proc Natl Acad Sci U S A. 2000; 97: 11086–11091.[Abstract/Free Full Text]
78. Barlic J, Andrews JD, Kelvin AA, Bosinger SE, DeVries ME, Xu L, Dobransky T, Feldman RD, Ferguson SS, Kelvin DJ. Regulation of tyrosine kinase activation and granule release through ß-arrestin by CXCRI. Nat Immunol. 2000; 1: 227–233.[CrossRef][Medline] [Order article via Infotrieve]
79. DeFea KA, Zalevsky J, Thoma MS, Dery O, Mullins RD, Bunnett NW. ß-Arrestin–dependent endocytosis of proteinase-activated receptor 2 is required for intracellular targeting of activated ERK1/2. J Cell Biol. 2000; 148: 1267–1281.[Abstract/Free Full Text]
80. Imamura T, Huang J, Dalle S, Ugi S, Usui I, Luttrell LM, Miller WE, Lefkowitz RJ, Olefsky JM. ß-Arrestin–mediated recruitment of the Src family kinase Yes mediates endothelin-1–stimulated glucose transport. J Biol Chem.;. 2001; 276: 43663–43667.[Abstract/Free Full Text]<