Targeting Individual GPCRs with Redesigned Nonvisual Arrestins

Giménez, Luis E.; Vishnivetskiy, Sergey A.; Gurevich, Vsevolod V.

doi:10.1007/978-3-642-41199-1_8

Cited by 7 publications

(6 citation statements)

References 109 publications

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“…All mutations were introduced on Ala87Val background, as described (7,27,30), which mimics more receptor-specific visual subtypes (34,35), reducing the flexibility of the N-domain (36,37). To generate all mutants, mutagenizing oligonucleotides were used as forward primers and an oligonucleotide downstream from the far restriction site to be used for subcloning was used as a reverse primer.…”

Section: Methodsmentioning

confidence: 99%

“…In contrast, two ubiquitously expressed non-visual subtypes, arrestin-2 (arr-2) and arrestin-3 (arr-3), are fairly promiscuous and interact with a huge range of GPCRs (3). Recent advances in our understanding of the molecular basis of receptor specificity of arrestin proteins pave the way to targeted construction of reengineered receptor-specific arrestins (7,8). …”

Section: Introductionmentioning

confidence: 99%

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Mutations in arrestin-3 differentially affect binding to neuropeptide Y receptor subtypes

Giménez

Babilon

Wanka

et al. 2014

Cellular Signalling

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Based on the identification of residues that determine receptor selectivity in arrestins and the phylogenetic analysis of the arrestin (arr) family, we introduced fifteen mutations of receptor-discriminator residues in arr-3, which were identified previously using mutagenesis, in vitro binding, and BRET-based recruitment assay in intact cells. The effects of these mutations were tested using neuropeptide Y receptors Y1R and Y2R. NPY-elicited arr-3 recruitment to Y1R was not affected by these mutations, or even alanine substitution of all ten residues (arr-3-NCA), which prevented arr-3 binding to other receptors tested so far. However, NCA and two other mutations prevented agonist-independent arr-3 pre-docking to Y1R. In contrast, eight out of 15 mutations significantly reduced agonist-dependent arr-3 recruitment to Y2R. NCA eliminated arr-3 binding to active Y2R, whereas Tyr239Thr reduced it ~7-fold. Thus, manipulation of key residues on the receptor-binding surface generates arr-3 with high preference for Y1R over Y2R. Several mutations differentially affect arr-3 pre-docking and agonist-induced recruitment. Thus, arr-3 recruitment to the receptor involves several mechanistically distinct steps. Targeted mutagenesis can fine-tune arrestins directing them to specific receptors and particular activation states of the same receptor.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Mutations in arrestin-3 differentially affect binding to neuropeptide Y receptor subtypes

Giménez

Babilon

Wanka

et al. 2014

Cellular Signalling

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“…To measure arrestin interactions with GPCRs co-expressed in the same cell, we used bioluminescence resonance energy transfer (BRET) between receptors C-terminally tagged with Renilla luciferase and arrestins N-terminally tagged with Venus, as described [10,23,26,30]. The middle-loop mutations were introduced in arrestin-3 carrying an Ala87Val mutation, which is expected to increase its receptor selectivity [23].…”

Section: Resultsmentioning

confidence: 99%

“…Thus, enhanced arrestins that do not require receptor phosphorylation can potentially be used in compensational therapy aiming at suppression of excessive G protein-mediated signaling [10,11]. Vertebrates have> 800 GPCRs [12,13] and only four different arrestins [14].…”

Section: Introductionmentioning

confidence: 99%

Differential manipulation of arrestin-3 binding to basal and agonist-activated G protein-coupled receptors

Prokop

Perry

Vishnivetskiy

et al. 2017

Cellular Signalling

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Non-visual arrestins interact with hundreds of different G protein-coupled receptors (GPCRs). Here we show that by introducing mutations into elements that directly bind receptors, the specificity of arrestin-3 can be altered. Several mutations in the two parts of the central “crest” of the arrestin molecule, middle-loop and C-loop, enhanced or reduced arrestin-3 interactions with several GPCRs in receptor subtype and functional state-specific manner. For example, the Lys139Ile substitution in the middle-loop dramatically enhanced the binding to inactive M2 muscarinic receptor, so that agonist activation of the M2 did not further increase arrestin-3 binding. Thus, the Lys139Ile mutation made arrestin-3 essentially an activation-independent binding partner of M2, whereas its interactions with other receptors, including the β2-adrenergic receptor and the D1 and D2 dopamine receptors, retained normal activation dependence. In contrast, the Ala248Val mutation enhanced agonist-induced arrestin-3 binding to the β2-adrenergic and D2 dopamine receptors, while reducing its interaction with the D1 dopamine receptor. These mutations represent the first example of altering arrestin specificity via enhancement of the arrestin-receptor interactions rather than selective reduction of the binding to certain subtypes.

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“…Several additional residues in the finger loop that binds the cytoplasmic cavity of active GPCRs also appear to serve the same purpose [105]. These data suggest the feasibility of narrowing receptor specificity of non-visual subtypes, constructing mutants that target few receptors, or possibly even individual GPCRs [148].…”

Section: Receptor Preference Of Arrestinsmentioning

confidence: 94%

Arrestins: A Small Family of Multi-functional Proteins

Gurevich

2024

Preprint

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The first member of the family, visual arrestin-1, was discovered in the late 1970s. Later the other three mammalian subtypes were identified and cloned. The first described function was regulation of GPCR signaling: arrestins bind active phosphorylated GPCRs, blocking their coupling to G proteins. It was later discovered that receptor-bound and free arrestins interact with numerous proteins, regulating GPCR trafficking and various signaling pathways, including those that determine cell fate. Arrestins have no enzymatic activity, they function by organizing multi-protein complexes and localizing their interaction partners to particular cellular compartments. Today we understand the molecular mechanism of arrestin interactions with GPCRs better than the mechanisms underlying other functions. However, even limited knowledge enabled the construction of signaling-biased arrestin mutants and extraction of biologically active monofunctional peptides from these multifunctional proteins. Manipulation of cellular signaling with arrestin-based tools has research and likely therapeutic potential: re-engineered proteins and their parts can produce effects that conventional small molecule drugs cannot.

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Targeting Individual GPCRs with Redesigned Nonvisual Arrestins

Cited by 7 publications

References 109 publications

Mutations in arrestin-3 differentially affect binding to neuropeptide Y receptor subtypes

Mutations in arrestin-3 differentially affect binding to neuropeptide Y receptor subtypes

Differential manipulation of arrestin-3 binding to basal and agonist-activated G protein-coupled receptors

Arrestins: A Small Family of Multi-functional Proteins

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