Monitoring agonist-promoted conformational changes of β-arrestin in living cells by intramolecular BRET

Pascale G Charest, Sonia Terrillon, Michel Bouvier

Research output: Contribution to journalArticle

127 Citations (Scopus)

Abstract

Recruitment of β-arrestin (β-arr) to agonist-stimulated G-protein-coupled receptors (GPCRs) has a crucial role in controlling signalling efficacy and selectivity. When translocated to the receptor, β-arr is believed to undergo important conformational rearrangement necessary for its downstream actions. To probe these changes in living cells, we constructed an intramolecular bioluminescence resonance energy transfer (BRET)-based biosensor, in which β-arr is sandwiched between the Renilla luciferase (Luc) and the yellow fluorescent protein (YFP). We show that the intramolecular BRET between Luc and YFP was significantly increased following GPCR activation, suggesting a conformational rearrangement bringing the amino terminus and carboxyl terminus of β-arr in closer proximity. Kinetic analysis showed that this conformational change follows the initial β-arr/receptor engagement. In addition to providing new insights into the agonist-induced conformational rearrangements of β-arr in living cells, the double-brilliance β-arr offers a universal biosensor for GPCR activation, allowing the study of native receptors in large-scale screening analysis.

Original languageEnglish (US)
Pages (from-to)334-340
Number of pages7
JournalEMBO Reports
Volume6
Issue number4
DOIs
StatePublished - Apr 2005
Externally publishedYes

Fingerprint

Bioluminescence
Arrestin
Energy Transfer
G-Protein-Coupled Receptors
Energy transfer
Cells
Biosensing Techniques
Biosensors
Monitoring
Renilla Luciferases
Chemical activation
Luciferases
Screening
Proteins
Kinetics

Keywords

  • Arrestin
  • Biosensor
  • BRET
  • GPCR

ASJC Scopus subject areas

  • Genetics
  • Cell Biology

Cite this

Monitoring agonist-promoted conformational changes of β-arrestin in living cells by intramolecular BRET. / Charest, Pascale G; Terrillon, Sonia; Bouvier, Michel.

In: EMBO Reports, Vol. 6, No. 4, 04.2005, p. 334-340.

Research output: Contribution to journalArticle

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