Cell surface protein-protein interaction analysis with combined time-resolved FRET and snap-tag technologies: application to GPCR oligomerization

Damien Maurel (1, 2) , Laetitia Comps-Agrar (1) , Carsten Brock (1) , Marie-Laure Rives (1) , Emmanuel Bourrier (2) , Mohammed Akli Ayoub (1) , Hervé Bazin (2) , Norbert Tinel (2) , Thierry Durroux (1) , Laurent Prézeau (1) , Eric Trinquet (2) , Jean-Philippe Pin (1)

1 IGF - Institut de Génomique Fonctionnelle
2 Cis Bio International

Cell-surface proteins are important in cell-cell communication. They assemble into heterocomplexes that include different receptors and effectors. Elucidation and manipulation of such protein complexes offers new therapeutic possibilities. We describe a methodology combining time-resolved fluorescence resonance energy transfer (FRET) with snap-tag technology to quantitatively analyze protein-protein interactions at the surface of living cells, in a high throughput-compatible format. Using this approach, we examined whether G protein-coupled receptors (GPCRs) are monomers or assemble into dimers or larger oligomers--a matter of intense debate. We obtained evidence for the oligomeric state of both class A and class C GPCRs. We also observed different quaternary structure of GPCRs for the neurotransmitters glutamate and gamma-aminobutyric acid (GABA): whereas metabotropic glutamate receptors assembled into strict dimers, the GABA(B) receptors spontaneously formed dimers of heterodimers, offering a way to modulate G-protein coupling efficacy. This approach will be useful in systematic analysis of cell-surface protein interaction in living cells.

Mots clés: GPCR; Biophysique; FRET; Dimerization; GABA

Domaines: Neurobiologie; Pharmacologie

Damien Maurel, Laetitia Comps-Agrar, Carsten Brock, Marie-Laure Rives, Emmanuel Bourrier, et al.. Cell surface protein-protein interaction analysis with combined time-resolved FRET and snap-tag technologies: application to GPCR oligomerization. Nature Methods, 2008, 5 (6), pp.561. ⟨10.1038/nmeth.1213⟩. ⟨hal-00318856⟩