Arrestin Interactions with G Protein-coupled Receptors

DIRECT BINDING STUDIES OF WILD TYPE AND MUTANT ARRESTINS WITH RHODOPSIN, β2-ADRENERGIC, AND m2 MUSCARINIC CHOLINERGIC RECEPTORS (*)

  1. Jeffrey L. Benovic(3)(§)
  1. From the (1)From theDepartment of Pharmacology, Northwestern University Medical School, Chicago, Illinois 60611, the
  2. (2)Department of Medicine, University of Wisconsin, Madison, Wisconsin 53792, and the
  3. (3)Department of Pharmacology, Jefferson Cancer Institute, Thomas Jefferson University, Philadelphia, Pennsylvania 19107
  1. § Established Investigator of the American Heart Association. To whom correspondence should be addressed to: Thomas Jefferson University, 233 South 10th St., Philadelphia, PA 19107. Tel.: 215-955-4607; Fax: 215-923-1098.

Abstract

Arrestins play an important role in quenching signal transduction initiated by G protein-coupled receptors. To explore the specificity of arrestin-receptor interaction, we have characterized the ability of various wild-type arrestins to bind to rhodopsin, the β2-adrenergic receptor (β2AR), and the m2 muscarinic cholinergic receptor (m2 mAChR). Visual arrestin was found to be the most selective arrestin since it discriminated best between the three different receptors tested (highest binding to rhodopsin) as well as between the phosphorylation and activation state of the receptor (>10-fold higher binding to the phosphorylated light-activated form of rhodopsin compared to any other form of rhodopsin). While β-arrestin and arrestin 3 were also found to preferentially bind to the phosphorylated activated form of a given receptor, they only modestly discriminated among the three receptors tested. To explore the structural characteristics important in arrestin function, we constructed a series of truncated and chimeric arrestins. Analysis of the binding characteristics of the various mutant arrestins suggests a common molecular mechanism involved in determining receptor binding selectivity. Structural elements that contribute to arrestin binding include: 1) a C-terminal acidic region that serves a regulatory role in controlling arrestin binding selectivity toward the phosphorylated and activated form of a receptor, without directly participating in receptor interaction; 2) a basic N-terminal domain that directly participates in receptor interaction and appears to serve a regulatory role via intramolecular interaction with the C-terminal acidic region; and 3) two centrally localized domains that are directly involved in determining receptor binding specificity and selectivity. A comparative structure-function model of all arrestins and a kinetic model of β-arrestin and arrestin 3 interaction with receptors are proposed.

Footnotes

  • * This research was supported in part by Grants GM44944 (to J. L. B.), GM47417 (to J. L. B.), GM47120 (to J. J. O.), and HL31601 (to M. M. H.) from the National Institutes of Health, a grant-in-aid from the American Heart Association (to M. M. H.), and a grant from the National Kidney Foundation of Wisconsin (to J. J. O.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

  • 1 The abbreviations used are:

    β2AR
    β2-adrenergic receptor
    arrestin
    a generic term that includes all arrestins
    arr
    visual arrestin
    βarr
    β-arrestin
    arr3
    arrestin 3
    βARK
    β-adrenergic receptor kinase
    G protein
    guanine nucleotide binding protein
    m2 mAChR
    m2 muscarinic cholinergic receptor
    mAChRGraphic
    carbachol-activated mAChR
    P-mAChR
    phosphorylated mAChR
    P-mAChRGraphic
    phosphorylated carbachol-activated mAChR
    β2ARGraphic
    isoproterenol-activated β2AR
    P-β2AR
    phosphorylated β2AR
    P-β2ARGraphic
    isoproterenol-activated phosphorylated β2AR
    Rh
    dark rhodopsin
    RhGraphic
    light-activated rhodopsin
    P-Rh
    phosphorylated rhodopsin
    P-RhGraphic
    phosphorylated light-activated rhodopsin
    S
    short
    L
    long.

  • Received July 8, 1994.
  • Revision received November 14, 1994.
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