title: β-Arrestin biosensors reveal a rapid, receptor-dependent activation/deactivation cycle.
abstract: (β-)Arrestins are important regulators of G-protein-coupled receptors (GPCRs). They bind to active, phosphorylated GPCRs and thereby shut off 'classical' signalling to G proteins, trigger internalization of GPCRs via interaction with the clathrin machinery and mediate signalling via 'non-classical' pathways. In addition to two visual arrestins that bind to rod and cone photoreceptors (termed arrestin1 and arrestin4), there are only two (non-visual) β-arrestin proteins (β-arrestin1 and β-arrestin2, also termed arrestin2 and arrestin3), which regulate hundreds of different (non-visual) GPCRs. Binding of these proteins to GPCRs usually requires the active form of the receptors plus their phosphorylation by G-protein-coupled receptor kinases (GRKs). The binding of receptors or their carboxy terminus as well as certain truncations induce active conformations of (β-)arrestins that have recently been solved by X-ray crystallography. Here we investigate both the interaction of β-arrestin with GPCRs, and the β-arrestin conformational changes in real time and in living human cells, using a series of fluorescence resonance energy transfer (FRET)-based β-arrestin2 biosensors. We observe receptor-specific patterns of conformational changes in β-arrestin2 that occur rapidly after the receptor-β-arrestin2 interaction. After agonist removal, these changes persist for longer than the direct receptor interaction. Our data indicate a rapid, receptor-type-specific, two-step binding and activation process between GPCRs and β-arrestins. They further indicate that β-arrestins remain active after dissociation from receptors, allowing them to remain at the cell surface and presumably signal independently. Thus, GPCRs trigger a rapid, receptor-specific activation/deactivation cycle of β-arrestins, which permits their active signalling.
authors.affiliation: Institute of Pharmacology and Toxicology, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.|Rudolf Virchow Center, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.
authors.forename: Susanne
authors.initials: S
authors.lastname: Nuber
authors.affiliation: Institute of Pharmacology and Toxicology, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.|Rudolf Virchow Center, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.
authors.forename: Ulrike
authors.initials: U
authors.lastname: Zabel
authors.affiliation: Institute of Pharmacology and Toxicology, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.|Comprehensive Heart Failure Center, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.
authors.forename: Kristina
authors.initials: K
authors.lastname: Lorenz
authors.affiliation: Institute of Pharmacology and Toxicology, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Nuber
authors.affiliation: Molecular Pharmacology Group, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, UK.
authors.forename: Graeme
authors.initials: G
authors.lastname: Milligan
authors.affiliation: MRC Toxicology Unit, University of Leicester, Hodgkin Building, Lancaster Road, Leicester LE1 9HN, UK.
authors.forename: Andrew B
authors.initials: AB
authors.lastname: Tobin
authors.affiliation: Institute of Pharmacology and Toxicology, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.|Rudolf Virchow Center, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.|Comprehensive Heart Failure Center, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Lohse
authors.affiliation: Institute of Pharmacology and Toxicology, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.|Rudolf Virchow Center, University of Würzburg, Versbacher Str. 9, 97078 Würzburg, Germany.
authors.forename: Carsten
authors.initials: C
authors.lastname: Hoffmann
mesh_terms: D000818:Animals; D019390:Arrestins / Q000737:chemistry / Q000378:metabolism*; D015374:Biosensing Techniques; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D018360:Crystallography, X-Ray; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry / Q000378:metabolism*; D015398:Signal Transduction; D013379:Substrate Specificity; D013997:Time Factors; D000071557:beta-Arrestins
chemical_list: D019390:Arrestins; D043562:Receptors, G-Protein-Coupled; D000071557:beta-Arrestins