Arrestin and Its Splice Variant Arr1–370A(p44)

Schröder, Katrin; Pulvermüller, Alexander; Hofmann, Klaus Peter

doi:10.1074/jbc.m206211200

Cited by 38 publications

(28 citation statements)

References 53 publications

(31 reference statements)

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“…Furthermore, Loop V-VI movement may be coincident with other proposed conformational changes in arrestin (34 -36). For example, the C-terminal tail of arrestin is clearly important for regulating arrestin function since truncation of the C terminus and various C-terminal mutations result in constitutively active arrestin (34,(37)(38)(39)(40). It is possible the C terminus may "hold" arrestin in an inactive conformation, and its mobilization may have to occur to enable arrestin to bind Rho*-P. Such movement has in fact been measured using spin probes in the C terminus of arrestin (12).…”

Section: Discussionmentioning

confidence: 99%

Dynamics of Arrestin-Rhodopsin Interactions

Sommer

Farrens

McDowell

et al. 2007

Journal of Biological Chemistry

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In this study we investigate conformational changes in Loop V-VI of visual arrestin during binding to light-activated, phosphorylated rhodopsin (Rho*-P) using a combination of site-specific cysteine mutagenesis and intramolecular fluorescence quenching. Introduction of cysteines at positions in the N-domain at residues predicted to be in close proximity to Ile-72 in Loop V-VI of arrestin (i.e. Glu-148 and Lys-298) appear to form an intramolecular disulfide bond with I72C, significantly diminishing the binding of arrestin to Rho*-P. Using a fluorescence approach, we show that the steady-state emission from a monobromobimane fluorophore in Loop V-VI is quenched by tryptophan residues placed at 148 or 298. This quenching is relieved upon binding of arrestin to Rho*-P. These results suggest that arrestin Loop V-VI moves during binding to Rho*-P and that conformational flexibility of this loop is essential for arrestin to adopt a high affinity binding state.Rhodopsin activity is controlled by a multistep process whereby activated rhodopsin is phosphorylated on its C terminus by rhodopsin kinase, leading to the binding and quenching of rhodopsin activity by visual arrestin. Visual arrestin belongs to the arrestin superfamily, which includes rod and cone arrestins as well the ␤-arrestins (1). The ␤-arrestins, which are involved in the inactivation of agonist-occupied G-proteincoupled receptors (for review, see Ref. 2), have an additional role not seen in visual arrestins whereby they direct receptor endocytosis by acting as a clathrin or AP-2 adaptor; e.g. Refs. 3 and 4. Visual arrestin is an important model system not only for understanding the visual response but also for understanding the broadreaching mechanism for controlling cellular signal transduction cascades mediated by G protein-coupled receptors.The crystal structures of rod, cone, and ␤-1 arrestin are remarkably similar. All show a structure consisting of two cupshaped domains formed from a seven-stranded ␤ sandwich (5-7). These structures, of the unbound state, have provided an important framework on which to base predictions for the binding interaction between arrestin and light-activated phosphorylated rhodopsin (Rho*-P). 4 Interestingly, in the crystal structures of visual arrestin, Loop V-VI (residues 68 -78 in bovine visual arrestin (7)) is seen in two primary conformers, one in which Loop V-VI is extended away from the main bulk of the protein (Fig. 1, ␣ conformer) and one with the loop folded into the N-terminal domain of the protein (␤ conformer). Several groups have identified that Loop V-VI has an important role in receptor binding. Experimental evidence shows (i) this loop is a vital component of arrestin that directs receptor preference (8), (ii) insertion of a peptide into this loop can either directly block or be used to competitively block binding to Rho*-P (9, 10), (iii) a fluorescent label at residue 72 in this loop apparently buries itself into the rhodopsin interface upon binding (10, 11), and (iv) spin labels in this loop have reduced ...

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Section: Discussionmentioning

confidence: 99%

Dynamics of Arrestin-Rhodopsin Interactions

Sommer

Farrens

McDowell

et al. 2007

Journal of Biological Chemistry

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“…For example, do the dynamics of full-length arrestin binding and release differ from the splice variant p 44 (53)? What are the effects of exogenously added retinal on arrestin binding and release?…”

Section: Discussionmentioning

confidence: 99%

Dynamics of Arrestin-Rhodopsin Interactions

Sommer

Smith

Farrens

2005

Journal of Biological Chemistry

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In this study, we address the mechanism of visual arrestin release from light-activated rhodopsin using fluorescently labeled arrestin mutants. We find that two mutants, I72C and S251C, when labeled with the small, solvent-sensitive fluorophore monobromobimane, exhibit spectral changes only upon binding light-activated, phosphorylated rhodopsin. Our analysis indicates that these changes are probably due to a burying of the probes at these sites in the rhodopsin-arrestin or phospholipid-arrestin interface. Using a fluorescence approach based on this observation, we demonstrate that arrestin and retinal release are linked and are described by similar activation energies. However, at physiological temperatures, we find that arrestin slows the rate of retinal release ϳ2-fold and abolishes the pH dependence of retinal release. Using fluorescence, EPR, and biochemical approaches, we also find intriguing evidence that arrestin binds to a post-Meta II photodecay product, possibly Meta III. We speculate that arrestin regulates levels of free retinal in the rod cell to help limit the formation of damaging oxidative retinal adducts. Such adducts may contribute to diseases like atrophic age-related macular degeneration (AMD). Thus, arrestin may serve to both attenuate rhodopsin signaling and protect the cell from excessive retinal levels under bright light conditions.The visual photoreceptor rhodopsin is perhaps the best model system for understanding the mechanisms used in Gprotein-coupled receptor (GPCR) 1 signaling, as detailed information exists on the structures and dynamic interactions of the protein constituents (1). Visual activation begins with absorption of light by the 11-cis-retinal chromophore in rhodopsin. The photoactivated form of rhodopsin, Rho* or "Meta II," interacts with and activates the G-protein transducin, which exchanges nucleotide and then diffuses to interact with downstream effectors. Signaling by Rho* is terminated by slow thermal decay and the release of retinal. Alternatively, signaling can be quickly terminated by a process that begins with phosphorylation of rhodopsin's C-terminal tail through the action of rhodopsin kinase (2, 3). The phosphorylated Rho* is then bound by arrestin, which stops signaling by physically occluding the G-protein binding site (4, 5).In the present study, we address how these two inactivation mechanisms are related and, specifically, what governs arrestin release from rhodopsin. Arrestin is known to bind to phosphorylated Meta II, a form in which the photolyzed chromophore all-trans-retinal is still attached to the receptor by a deprotonated Schiff base. Arrestin does not bind phosphorylated opsin, but all-trans retinal added exogenously can stimulate arrestin binding to phosphorylated opsin (6, 7). Early studies showed indirectly that retinal release and arrestin release are probably interrelated events (7,8). However, how these processes are linked or whether arrestin binds other photointermediates of rhodopsin (such as the storage form Meta III) is still unknown...

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“…Size Exclusion Chromatography-Size exclusion chromatography is a very useful tool to determine protein-protein interaction (15,39) and was used in this study to characterize direct complex formation between the four different mouse centrins (MmCen1 to MmCen4), transducin (G t ), and its subunits. To determine the binding of the centrins to transducin the molecular weight shift of the complex was used.…”

Section: Methodsmentioning

confidence: 99%