title: Quaternary structure of a G-protein-coupled receptor heterotetramer in complex with Gi and Gs.
abstract: BACKGROUND
  G-protein-coupled receptors (GPCRs), in the form of monomers or homodimers that bind heterotrimeric G proteins, are fundamental in the transfer of extracellular stimuli to intracellular signaling pathways. Different GPCRs may also interact to form heteromers that are novel signaling units. Despite the exponential growth in the number of solved GPCR crystal structures, the structural properties of heteromers remain unknown.
  
  
  RESULTS
  We used single-particle tracking experiments in cells expressing functional adenosine A1-A2A receptors fused to fluorescent proteins to show the loss of Brownian movement of the A1 receptor in the presence of the A2A receptor, and a preponderance of cell surface 2:2 receptor heteromers (dimer of dimers). Using computer modeling, aided by bioluminescence resonance energy transfer assays to monitor receptor homomerization and heteromerization and G-protein coupling, we predict the interacting interfaces and propose a quaternary structure of the GPCR tetramer in complex with two G proteins.
  
  
  CONCLUSIONS
  The combination of results points to a molecular architecture formed by a rhombus-shaped heterotetramer, which is bound to two different interacting heterotrimeric G proteins (Gi and Gs). These novel results constitute an important advance in understanding the molecular intricacies involved in GPCR function.
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain.
authors.forename: Gemma
authors.initials: G
authors.lastname: Navarro
authors.affiliation: Laboratori de Medicina Computacional, Unitat de Bioestadística, Facultat de Medicina, Universitat Autònoma de Barcelona, 08193, Bellaterra, Spain.
authors.forename: Arnau
authors.initials: A
authors.lastname: Cordomí
authors.affiliation: Max Planck Institute for Intelligent Systems, Heisenbergstrasse 3, 70569, Stuttgart, Germany.|German Cancer Research Center, Bioquant, Im Neuenheimer Feld 267, 69120, Heidelberg, Germany.
authors.forename: Monika
authors.initials: M
authors.lastname: Zelman-Femiak
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain.
authors.forename: Marc
authors.initials: M
authors.lastname: Brugarolas
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain.
authors.forename: Estefania
authors.initials: E
authors.lastname: Moreno
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain.
authors.forename: David
authors.initials: D
authors.lastname: Aguinaga
authors.affiliation: Laboratori de Medicina Computacional, Unitat de Bioestadística, Facultat de Medicina, Universitat Autònoma de Barcelona, 08193, Bellaterra, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Perez-Benito
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain.
authors.forename: Antoni
authors.initials: A
authors.lastname: Cortés
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain.
authors.forename: Vicent
authors.initials: V
authors.lastname: Casadó
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain.
authors.forename: Josefa
authors.initials: J
authors.lastname: Mallol
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain.
authors.forename: Enric I
authors.initials: EI
authors.lastname: Canela
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain.
authors.forename: Carme
authors.initials: C
authors.lastname: Lluís
authors.affiliation: Laboratori de Medicina Computacional, Unitat de Bioestadística, Facultat de Medicina, Universitat Autònoma de Barcelona, 08193, Bellaterra, Spain.
authors.forename: Leonardo
authors.initials: L
authors.lastname: Pardo
authors.affiliation: Max Planck Institute for Intelligent Systems, Heisenbergstrasse 3, 70569, Stuttgart, Germany.|German Cancer Research Center, Bioquant, Im Neuenheimer Feld 267, 69120, Heidelberg, Germany.|Interfaculty Institute of Biochemistry, Hoppe-Seyler-Strasse 4, 72076, Tübingen, Germany.
authors.forename: Ana J
authors.initials: AJ
authors.lastname: García-Sáez
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain. p.mccormick@uea.ac.uk.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain. p.mccormick@uea.ac.uk.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain. p.mccormick@uea.ac.uk.|School of Pharmacy, University of East Anglia, Norwich, NR4 7TJ, UK. p.mccormick@uea.ac.uk.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: McCormick
authors.affiliation: Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain. rfranco123@gmail.com.|Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain. rfranco123@gmail.com.|Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, 08028, Spain. rfranco123@gmail.com.
authors.forename: Rafael
authors.initials: R
authors.lastname: Franco
mesh_terms: D000818:Animals; D057809:HEK293 Cells; D020962:Heterotrimeric GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D018047:Receptors, Purinergic P1 / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D018047:Receptors, Purinergic P1; D020962:Heterotrimeric GTP-Binding Proteins
keywords: BRET; GPCR; Heterotetramer; Heterotrimeric G protein; Molecular modeling; Single-particle tracking