Mutations in the ‘DRY’ motif of the CB1 cannabinoid receptor result in biased receptor variants

Gyombolai, Pál; Tóth, András; Tímár, Dániel; Turu, Gábor; Hunyady, László

doi:10.1530/jme-14-0219

Cited by 31 publications

(32 citation statements)

References 53 publications

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“…Investigating signaling from biased CB 1 receptors such as S426A/S430A and the DRY mutant (Asp-Arg-Tyr) together with the identification of biased ligands and the crystal structure of CB 1 receptors should provide important tools to elucidate the mechanisms and roles of CB 1 receptor signaling (Gyombolai et al, 2015; Delgado-Peraza et al, 2016; Hua et al, 2016). …”

Section: The Endocannabinoid System In the Cnsmentioning

confidence: 99%

Cannabinoid Receptors in the Central Nervous System: Their Signaling and Roles in Disease

Kendall

Yudowski

2017

Front. Cell. Neurosci.

258

208

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The identification and cloning of the two major cannabinoid (CB1 and CB2) receptors together with the discovery of their endogenous ligands in the late 80s and early 90s, resulted in a major effort aimed at understanding the mechanisms and physiological roles of the endocannabinoid system (ECS). Due to its expression and localization in the central nervous system (CNS), the CB1 receptor together with its endogenous ligands (endocannabinoids (eCB)) and the enzymes involved in their synthesis and degradation, has been implicated in multiple pathophysiological events ranging from memory deficits to neurodegenerative disorders among others. In this review, we will provide a general overview of the ECS with emphasis on the CB1 receptor in health and disease. We will describe our current understanding of the complex aspects of receptor signaling and trafficking, including the non-canonical signaling pathways such as those mediated by β-arrestins within the context of functional selectivity and ligand bias. Finally, we will highlight some of the disorders in which CB1 receptors have been implicated. Significant knowledge has been achieved over the last 30 years. However, much more research is still needed to fully understand the complex roles of the ECS, particularly in vivo and to unlock its true potential as a source of therapeutic targets.

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Section: The Endocannabinoid System In the Cnsmentioning

confidence: 99%

Cannabinoid Receptors in the Central Nervous System: Their Signaling and Roles in Disease

Kendall

Yudowski

2017

Front. Cell. Neurosci.

258

208

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“…The conserved DRY motif in the intracellular region of TM3 participates directly in G protein coupling (58) and is known to modulate G protein signaling by class A GPCRs (59,60). Mutation of the central Arg(3.50) residue of the DRY motif in other GPCRs (61,62), including CCR5 (63), has been shown to produce receptor bias wherein G protein signaling is depleted but arrestin interactions are preserved or enhanced. Consistent with these findings, the CXCR4 R134A mutation almost completely abrogated Gαi recruitment ( Fig.…”

Section: Quantifying the Apparent Pathway Bias In Mutation-induced Simentioning

confidence: 99%

Functional anatomy of the full length CXCR4-CXCL12 complex systematically dissected by quantitative model-guided mutagenesis

Stephens

Ngo

Kufareva

et al. 2020

Preprint

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One sentence summary: A systematic structure-guided mutagenesis study of chemokine receptor CXCR4 reveals novel insights into epitopes regulating ligand recognition, ligand specificity and CXCL12-mediated signaling.Abstract: Due to their prominent role in development and infamy in cancer and HIV, the chemokine receptor CXCR4 and its ligand, CXCL12, have been the subject of numerous structural and functional studies. Nevertheless, a high resolution structure of the CXCR4-CXCL12 complex has not been reported. Even with several alternative computational models of the complex at hand, the relative contributions of different interaction epitopes to ligand binding, ligand selectivity and signaling are not readily apparent. Here, building upon our latest structural model, we employed a systematic mutagenesis strategy to dissect the functional anatomy of the of CXCR4-CXCL12 complex. Key charge swap mutagenesis experiments supported pairwise interactions between oppositely charged residues in the receptor and chemokine, confirming the accuracy of the predicted orientation of the chemokine relative to the receptor, while also providing insight into ligand selectivity. Progressive deletion of N-terminal residues revealed an unexpected contribution of the receptor N-terminus to chemokine signaling; this finding challenges a longstanding "two-site" hypothesis about the essential features of the receptor-chemokine interaction where the N-terminus is purported to only contribute to binding affinity. The results suggest that while the interaction of the chemokine N-terminus with the receptor binding pocket is the key driver of signaling, the signaling amplitude depends on the extent to which the receptor N-terminus binds the chemokine. Along with systematic characterization of other epitopes, the current data allow us to propose a comprehensive experimentally-consistent structural model for how the chemokine binds CXCR4 and initiates signal transmission through the receptor TM domain. IntroductionChemokine receptors are members of the Class A family G protein-coupled receptors (GPCRs) that are best known for their role in controlling the migration of cells, particularly leukocytes in the context of immune system function. They are activated by chemokines, small 8-10 kDa secreted proteins, via a mechanism that has long been described as involving "two sites" or "two steps" (1-5). According to the two-site mechanism, the globular domain of the chemokine binds to the N-terminus of a receptor (referred to as chemokine recognition site 1, CRS1) and contributes primarily to the stability of the complex whereas the N-terminus of the chemokine binds in the transmembrane (TM) domain binding pocket of the receptor (chemokine recognition site 2, CRS2) to activate signaling (6). The distinction between these two sites arose from the general observation that mutations in chemokine N-termini produce a disproportionately large effect on receptor signaling efficacy compared to mutations in the chemokine globular domains (7,8), with correspondin...

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“…Biochemical/mutation data with cannabinergic ligands and information on the structures of other class-A GPCRs have been used to identify candidate residues potentially involved in ligand functional selectivity at CBRs. Similar to other GPCRs, double mutation of D213 3.49 and R214 3.50 in the highly conserved aspartic acid-arginine-tyrosine (“DRY”) motif that plays a pivotal role in regulating GPCR conformational/activity states caused CB1R to bias towards β-arrestin signaling and away from G-protein activation without significant loss in binding affinity of cannabinoid agonists [85]. It was also observed that C355 6.47 of the CWxP motif was critical for binding of CP55940 and other classical cannabinoids that are known to induce receptor internalization through β-arrestin-mediated pathways [85].…”

Section: Structural Basis For Signaling Bias At Cannabinoid Gpcrsmentioning

confidence: 99%

“…Similar to other GPCRs, double mutation of D213 3.49 and R214 3.50 in the highly conserved aspartic acid-arginine-tyrosine (“DRY”) motif that plays a pivotal role in regulating GPCR conformational/activity states caused CB1R to bias towards β-arrestin signaling and away from G-protein activation without significant loss in binding affinity of cannabinoid agonists [85]. It was also observed that C355 6.47 of the CWxP motif was critical for binding of CP55940 and other classical cannabinoids that are known to induce receptor internalization through β-arrestin-mediated pathways [85]. However, WIN55212, an aminoalkylindole derivative that does not induce receptor internalization, maintained high affinity for CB1R, even when C355 6.47 had been mutated.…”

Section: Structural Basis For Signaling Bias At Cannabinoid Gpcrsmentioning

confidence: 99%

Functional selectivity at G-protein coupled receptors: Advancing cannabinoid receptors as drug targets

Mallipeddi

Janero

Zvonok

et al. 2017

Biochemical Pharmacology

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The phenomenon of functional selectivity, whereby a ligand preferentially directs the information output of a G-protein coupled receptor (GPCR) along (a) particular effector pathway(s) and away from others, has redefined traditional GPCR signaling paradigms to provide a new approach to structure-based drug design. The two principal cannabinoid receptors (CBRs) 1 and 2 belong to the class-A GPCR subfamily and are considered tenable therapeutic targets for several indications. Yet conventional orthosteric ligands (agonists, antagonists/inverse agonists) for these receptors have had very limited clinical utility due to their propensity to incite on-target adverse events. Chemically distinct classes of cannabinergic ligands exhibit signaling bias at CBRs toward individual subsets of signal transduction pathways. In this review, we discuss the known signaling pathways regulated by CBRs and examine the current evidence for functional selectivity at CBRs in response to endogenous and exogenous cannabinergic ligands as biased agonists. We further discuss the receptor and ligand structural features allowing for selective activation of CBR-dependent functional responses. The design and development of biased ligands may offer a pathway to therapeutic success for novel CBR-targeted drugs.

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Mutations in the ‘DRY’ motif of the CB1 cannabinoid receptor result in biased receptor variants

Cited by 31 publications

References 53 publications

Cannabinoid Receptors in the Central Nervous System: Their Signaling and Roles in Disease

Cannabinoid Receptors in the Central Nervous System: Their Signaling and Roles in Disease

Functional anatomy of the full length CXCR4-CXCL12 complex systematically dissected by quantitative model-guided mutagenesis

Functional selectivity at G-protein coupled receptors: Advancing cannabinoid receptors as drug targets

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