Dataset Information

Exploring GPCR-arrestin interfaces with genetically encoded crosslinkers

ABSTRACT: -arrestins (arr1 and arr2) are ubiquitous regulators of G protein-coupled receptor (GPCR) signalling. Available data suggest that -arrestins dock to different receptors in different ways. However, the structural characterization of arrestin-GPCR complexes is challenging and alternative approaches to study arrestin-GPCR complexes are needed. Here, starting from the finger loop as a major site for the interaction of arrestins with GPCRs, we genetically incorporate non-canonical amino acids for photo- and chemical crosslinking into arr1 and arr2 and explore binding topologies to GPCRs forming either stable or transient complexes with arrestin: the vasopressin receptor 2 (rhodopsin-like), the corticotropin releasing factor receptor 1 and the parathyroid hormone receptor 1 (both secretin-like). We show that each receptor leaves a unique footprint on arrestin, whereas the two -arrestins yield quite similar crosslinking patterns. Furthermore, we show that the method allows defining the orientation of arrestin respect to the GPCR. Finally, we provide direct evidence for the formation of arrestin oligomers in the cell.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Christian Ihling

LAB HEAD: Andrea Sinz

PROVIDER: PXD020418 | Pride | 2020-09-21

REPOSITORIES: Pride

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Publications

Exploring GPCR-arrestin interfaces with genetically encoded crosslinkers.

Böttke Thore T Ernicke Stefan S Serfling Robert R Ihling Christian C Burda Edyta E Gurevich Vsevolod V VV Sinz Andrea A Coin Irene I

EMBO reports 20200914 11

β-arrestins (βarr1 and βarr2) are ubiquitous regulators of G protein-coupled receptor (GPCR) signaling. Available data suggest that β-arrestins dock to different receptors in different ways. However, the structural characterization of GPCR-arrestin complexes is challenging and alternative approaches to study GPCR-arrestin complexes are needed. Here, starting from the finger loop as a major site for the interaction of arrestins with GPCRs, we genetically incorporate non-canonical amino acids for ...[more]

PMID: 32929862

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Project description:Bush2016 - Extended Carrousel model of GPCR-RGS This model is described in the article: Yeast GPCR signaling reflects the fraction of occupied receptors, not the number. Bush A, Vasen G, Constantinou A, Dunayevich P, Patop IL, Blaustein M, Colman-Lerner A. Mol. Syst. Biol. 2016 Dec; 12(12): 898 Abstract: According to receptor theory, the effect of a ligand depends on the amount of agonist-receptor complex. Therefore, changes in receptor abundance should have quantitative effects. However, the response to pheromone in Saccharomyces cerevisiae is robust (unaltered) to increases or reductions in the abundance of the G-protein-coupled receptor (GPCR), Ste2, responding instead to the fraction of occupied receptor. We found experimentally that this robustness originates during G-protein activation. We developed a complete mathematical model of this step, which suggested the ability to compute fractional occupancy depends on the physical interaction between the inhibitory regulator of G-protein signaling (RGS), Sst2, and the receptor. Accordingly, replacing Sst2 by the heterologous hsRGS4, incapable of interacting with the receptor, abolished robustness. Conversely, forcing hsRGS4:Ste2 interaction restored robustness. Taken together with other results of our work, we conclude that this GPCR pathway computes fractional occupancy because ligand-bound GPCR-RGS complexes stimulate signaling while unoccupied complexes actively inhibit it. In eukaryotes, many RGSs bind to specific GPCRs, suggesting these complexes with opposing activities also detect fraction occupancy by a ratiometric measurement. Such complexes operate as push-pull devices, which we have recently described. This model is hosted on BioModels Database and identified by: BIOMD0000000638. To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication for more information.

Project description:Bush2016 - Simplified Carrousel model of GPCR This model is described in the article: Yeast GPCR signaling reflects the fraction of occupied receptors, not the number. Bush A, Vasen G, Constantinou A, Dunayevich P, Patop IL, Blaustein M, Colman-Lerner A. Mol. Syst. Biol. 2016 Dec; 12(12): 898 Abstract: According to receptor theory, the effect of a ligand depends on the amount of agonist-receptor complex. Therefore, changes in receptor abundance should have quantitative effects. However, the response to pheromone in Saccharomyces cerevisiae is robust (unaltered) to increases or reductions in the abundance of the G-protein-coupled receptor (GPCR), Ste2, responding instead to the fraction of occupied receptor. We found experimentally that this robustness originates during G-protein activation. We developed a complete mathematical model of this step, which suggested the ability to compute fractional occupancy depends on the physical interaction between the inhibitory regulator of G-protein signaling (RGS), Sst2, and the receptor. Accordingly, replacing Sst2 by the heterologous hsRGS4, incapable of interacting with the receptor, abolished robustness. Conversely, forcing hsRGS4:Ste2 interaction restored robustness. Taken together with other results of our work, we conclude that this GPCR pathway computes fractional occupancy because ligand-bound GPCR-RGS complexes stimulate signaling while unoccupied complexes actively inhibit it. In eukaryotes, many RGSs bind to specific GPCRs, suggesting these complexes with opposing activities also detect fraction occupancy by a ratiometric measurement. Such complexes operate as push-pull devices, which we have recently described. This model is hosted on BioModels Database and identified by: BIOMD0000000637. To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication for more information.

Dataset Information

Exploring GPCR-arrestin interfaces with genetically encoded crosslinkers

Publications

Exploring GPCR-arrestin interfaces with genetically encoded crosslinkers.

Similar Datasets

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