A single phenylalanine residue in β-arrestin2 critically regulates its binding to G protein-coupled receptors.

Journal Article (Journal Article)

Arrestins and their yeast homologs, arrestin-related trafficking adaptors (ARTs), share a stretch of 29 amino acids called the ART motif. However, the functionality of that motif is unknown. We now report that deleting this motif prevents agonist-induced ubiquitination of β-arrestin2 (β-arr2) and blocks its association with activated G protein-coupled receptors (GPCRs). Within the ART motif, we have identified a conserved phenylalanine residue, Phe116, that is critical for the formation of β-arr2-GPCR complexes. β-arr2 Phe116Ala mutant has negligible effect on blunting β2-adrenergic receptor-induced cAMP generation unlike β-arr2, which promotes rapid desensitization. Furthermore, available structures for inactive and inositol hexakisphosphate 6-activated forms of bovine β-arr2 revealed that Phe116 is ensconced in a hydrophobic pocket, whereas the adjacent Phe117 and Phe118 residues are not. Mutagenesis of Phe117 and Phe118, but not Phe116, preserves GPCR interaction of β-arr2. Surprisingly, Phe116 is dispensable for the association of β-arr2 with its non-GPCR partners. β-arr2 Phe116Ala mutant presents a significantly reduced protein half-life compared with β-arr2 and undergoes constitutive Lys-48-linked polyubiquitination, which tags proteins for proteasomal degradation. We also found that Phe116 is critical for agonist-dependent β-arr2 ubiquitination with Lys-63-polyubiquitin linkages that are known mediators of protein scaffolding and signal transduction. Finally, we have shown that β-arr2 Phe116Ala interaction with activated β2-adrenergic receptor can be rescued with an in-frame fusion of ubiquitin. Taken together, we conclude that Phe116 preserves structural stability of β-arr2, regulates the formation of β-arr2-GPCR complexes that inhibit G protein signaling, and promotes subsequent ubiquitin-dependent β-arr2 localization and trafficking.

Full Text

Duke Authors

Cited Authors

  • Jean-Charles, P-Y; Rajiv, V; Sarker, S; Han, S; Bai, Y; Masoudi, A; Shenoy, SK

Published Date

  • May 2022

Published In

Volume / Issue

  • 298 / 5

Start / End Page

  • 101837 -

PubMed ID

  • 35307348

Pubmed Central ID

  • PMC9052155

Electronic International Standard Serial Number (EISSN)

  • 1083-351X

Digital Object Identifier (DOI)

  • 10.1016/j.jbc.2022.101837


  • eng

Conference Location

  • United States