title: A Perspective on Studying G-Protein-Coupled Receptor Signaling with Resonance Energy Transfer Biosensors in Living Organisms.
abstract: The last frontier for a complete understanding of G-protein-coupled receptor (GPCR) biology is to be able to assess GPCR activity, interactions, and signaling in vivo, in real time within biologically intact systems. This includes the ability to detect GPCR activity, trafficking, dimerization, protein-protein interactions, second messenger production, and downstream signaling events with high spatial resolution and fast kinetic readouts. Resonance energy transfer (RET)-based biosensors allow for all of these possibilities in vitro and in cell-based assays, but moving RET into intact animals has proven difficult. Here, we provide perspectives on the optimization of biosensor design, of signal detection in living organisms, and the multidisciplinary development of in vitro and cell-based assays that more appropriately reflect the physiologic situation. In short, further development of RET-based probes, optical microscopy techniques, and mouse genome editing hold great potential over the next decade to bring real-time in vivo GPCR imaging to the forefront of pharmacology.
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands (M.J.W.A.-H.); Department of Biochemistry, Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Quebec, Canada (M.B.); Department of Anesthesiology, Washington University, St. Louis, Missouri (M.R.B.); Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, Amsterdam, The Netherlands (J.U., J.G.); Cell Signalling Research Group, School of Biomedical Sciences, Medical School, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom (J.W., S.J.H.); Bio-Imaging-Center/Rudolf-Virchow-Zentrum and Institute of Pharmacology and Toxicology, University of Würzburg, Würzburg, Germany (C.H.); and Institute of Chemistry, University of Tartu, Tartu, Estonia (A.R.).
authors.forename: Jakobus
authors.initials: J
authors.lastname: van Unen
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands (M.J.W.A.-H.); Department of Biochemistry, Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Quebec, Canada (M.B.); Department of Anesthesiology, Washington University, St. Louis, Missouri (M.R.B.); Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, Amsterdam, The Netherlands (J.U., J.G.); Cell Signalling Research Group, School of Biomedical Sciences, Medical School, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom (J.W., S.J.H.); Bio-Imaging-Center/Rudolf-Virchow-Zentrum and Institute of Pharmacology and Toxicology, University of Würzburg, Würzburg, Germany (C.H.); and Institute of Chemistry, University of Tartu, Tartu, Estonia (A.R.).
authors.forename: Jeanette
authors.initials: J
authors.lastname: Woolard
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands (M.J.W.A.-H.); Department of Biochemistry, Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Quebec, Canada (M.B.); Department of Anesthesiology, Washington University, St. Louis, Missouri (M.R.B.); Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, Amsterdam, The Netherlands (J.U., J.G.); Cell Signalling Research Group, School of Biomedical Sciences, Medical School, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom (J.W., S.J.H.); Bio-Imaging-Center/Rudolf-Virchow-Zentrum and Institute of Pharmacology and Toxicology, University of Würzburg, Würzburg, Germany (C.H.); and Institute of Chemistry, University of Tartu, Tartu, Estonia (A.R.).
authors.forename: Ago
authors.initials: A
authors.lastname: Rinken
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands (M.J.W.A.-H.); Department of Biochemistry, Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Quebec, Canada (M.B.); Department of Anesthesiology, Washington University, St. Louis, Missouri (M.R.B.); Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, Amsterdam, The Netherlands (J.U., J.G.); Cell Signalling Research Group, School of Biomedical Sciences, Medical School, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom (J.W., S.J.H.); Bio-Imaging-Center/Rudolf-Virchow-Zentrum and Institute of Pharmacology and Toxicology, University of Würzburg, Würzburg, Germany (C.H.); and Institute of Chemistry, University of Tartu, Tartu, Estonia (A.R.).
authors.forename: Carsten
authors.initials: C
authors.lastname: Hoffmann
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands (M.J.W.A.-H.); Department of Biochemistry, Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Quebec, Canada (M.B.); Department of Anesthesiology, Washington University, St. Louis, Missouri (M.R.B.); Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, Amsterdam, The Netherlands (J.U., J.G.); Cell Signalling Research Group, School of Biomedical Sciences, Medical School, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom (J.W., S.J.H.); Bio-Imaging-Center/Rudolf-Virchow-Zentrum and Institute of Pharmacology and Toxicology, University of Würzburg, Würzburg, Germany (C.H.); and Institute of Chemistry, University of Tartu, Tartu, Estonia (A.R.).
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Hill
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands (M.J.W.A.-H.); Department of Biochemistry, Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Quebec, Canada (M.B.); Department of Anesthesiology, Washington University, St. Louis, Missouri (M.R.B.); Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, Amsterdam, The Netherlands (J.U., J.G.); Cell Signalling Research Group, School of Biomedical Sciences, Medical School, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom (J.W., S.J.H.); Bio-Imaging-Center/Rudolf-Virchow-Zentrum and Institute of Pharmacology and Toxicology, University of Würzburg, Würzburg, Germany (C.H.); and Institute of Chemistry, University of Tartu, Tartu, Estonia (A.R.).
authors.forename: Joachim
authors.initials: J
authors.lastname: Goedhart
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands (M.J.W.A.-H.); Department of Biochemistry, Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Quebec, Canada (M.B.); Department of Anesthesiology, Washington University, St. Louis, Missouri (M.R.B.); Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, Amsterdam, The Netherlands (J.U., J.G.); Cell Signalling Research Group, School of Biomedical Sciences, Medical School, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom (J.W., S.J.H.); Bio-Imaging-Center/Rudolf-Virchow-Zentrum and Institute of Pharmacology and Toxicology, University of Würzburg, Würzburg, Germany (C.H.); and Institute of Chemistry, University of Tartu, Tartu, Estonia (A.R.).
authors.forename: Michael R
authors.initials: MR
authors.lastname: Bruchas
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands (M.J.W.A.-H.); Department of Biochemistry, Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Quebec, Canada (M.B.); Department of Anesthesiology, Washington University, St. Louis, Missouri (M.R.B.); Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, Amsterdam, The Netherlands (J.U., J.G.); Cell Signalling Research Group, School of Biomedical Sciences, Medical School, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom (J.W., S.J.H.); Bio-Imaging-Center/Rudolf-Virchow-Zentrum and Institute of Pharmacology and Toxicology, University of Würzburg, Würzburg, Germany (C.H.); and Institute of Chemistry, University of Tartu, Tartu, Estonia (A.R.).
authors.forename: Michel
authors.initials: M
authors.lastname: Bouvier
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands (M.J.W.A.-H.); Department of Biochemistry, Institute for Research in Immunology and Cancer, Université de Montréal, Montréal, Quebec, Canada (M.B.); Department of Anesthesiology, Washington University, St. Louis, Missouri (M.R.B.); Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, Amsterdam, The Netherlands (J.U., J.G.); Cell Signalling Research Group, School of Biomedical Sciences, Medical School, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom (J.W., S.J.H.); Bio-Imaging-Center/Rudolf-Virchow-Zentrum and Institute of Pharmacology and Toxicology, University of Würzburg, Würzburg, Germany (C.H.); and Institute of Chemistry, University of Tartu, Tartu, Estonia (A.R.) merel.adjobo-hermans@radboudumc.nl.
authors.forename: Merel J W
authors.initials: MJ
authors.lastname: Adjobo-Hermans
mesh_terms: D000818:Animals; D015374:Biosensing Techniques / Q000379:methods* / Q000639:trends; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D043562:Receptors, G-Protein-Coupled