title: Internalized Receptor for Glucose-dependent Insulinotropic Peptide stimulates adenylyl cyclase on early endosomes.
abstract: Until very recently, G-protein dependent signal of GPCRs was thought to originate exclusively from the plasma membrane and internalized GPCRs were considered silent. Here, we demonstrated that, once internalized and located in the membrane of early endosomes, glucose-dependent Insulinotropic receptor (GIPR) continues to trigger production of cAMP and PKA activation. Direct evidence is based on identification of the active form of Gαs in early endosomes containing GIPR using a genetically encoded GFP tagged nanobody, and on detection of a distinct FRET signal accounting for cAMP production at the surface of endosomes containing GIP, compared to endosomes without GIP. Furthermore, decrease of the sustained phase of cAMP production and PKA activation kinetics as well as reversibility of cAMP production and PKA activity following GIP washout in cells treated with a pharmacological inhibitor of GIPR internalization, and continuous increase of cAMP level over time in the presence of dominant-negative Rab7, which causes accumulation of early endosomes in cells, were noticed. Hence the GIPR joins the few GPCRs which signal through G-proteins both at plasma membrane and on endosomes.
authors.affiliation: Laboratoire de Physique et Chimie des Nano-objets (LPCNO), team RTTC, Université de Toulouse, CNRS, INSA, INSERM, Université Paul Sabatier, Toulouse, France.
authors.forename: Sadek
authors.initials: S
authors.lastname: Ismail
authors.affiliation: Laboratoire de Physique et Chimie des Nano-objets (LPCNO), team RTTC, Université de Toulouse, CNRS, INSA, INSERM, Université Paul Sabatier, Toulouse, France.
authors.forename: Marie-Julie
authors.initials: MJ
authors.lastname: Gherardi
authors.affiliation: German Center for Cardiovascular Research, University Medical Center Hamburg-Eppendorf and Institute of Experimental Cardiovascular Research, Hamburg, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Froese
authors.affiliation: Cellular Imaging Facility Rangueil, INSERM U1048/I2MC, Toulouse, France.
authors.forename: Madjid
authors.initials: M
authors.lastname: Zanoun
authors.affiliation: Laboratoire de Physique et Chimie des Nano-objets (LPCNO), team RTTC, Université de Toulouse, CNRS, INSA, INSERM, Université Paul Sabatier, Toulouse, France.
authors.forename: Véronique
authors.initials: V
authors.lastname: Gigoux
authors.affiliation: Laboratoire de Physique et Chimie des Nano-objets (LPCNO), team RTTC, Université de Toulouse, CNRS, INSA, INSERM, Université Paul Sabatier, Toulouse, France.
authors.forename: Pascal
authors.initials: P
authors.lastname: Clerc
authors.affiliation: INSERM, UMR1048, University of Toulouse 3, Institute of metabolic and cardiovascular diseases, Toulouse, France.
authors.forename: Frederique
authors.initials: F
authors.lastname: Gaits-Iacovoni
authors.affiliation: Structural Biology Brussels, Vrije Universiteit Brussel, Brussel, Belgium; Structural Biology Research Center, VIB, Brussel, Belgium.
authors.forename: Jan
authors.initials: J
authors.lastname: Steyaert
authors.affiliation: German Center for Cardiovascular Research, University Medical Center Hamburg-Eppendorf and Institute of Experimental Cardiovascular Research, Hamburg, Germany.
authors.forename: Viacheslav O
authors.initials: VO
authors.lastname: Nikolaev
authors.affiliation: Laboratoire de Physique et Chimie des Nano-objets (LPCNO), team RTTC, Université de Toulouse, CNRS, INSA, INSERM, Université Paul Sabatier, Toulouse, France. Electronic address: Daniel.Fourmy@inserm.fr.
authors.forename: Daniel
authors.initials: D
authors.lastname: Fourmy
mesh_terms: D000262:Adenylyl Cyclases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D060807:Bioluminescence Resonance Energy Transfer Techniques; D002864:Chromogranins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000242:Cyclic AMP / Q000819:agonists / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004705:Endocytosis; D011992:Endosomes / Q000201:enzymology / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005749:Gastric Inhibitory Polypeptide / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport; D011964:Receptors, Gastrointestinal Hormone / Q000819:agonists / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015290:Second Messenger Systems; D061905:Single-Domain Antibodies / Q000235:genetics / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D000090802:rab7 GTP-Binding Proteins
chemical_list: D002864:Chromogranins; D005456:Fluorescent Dyes; D008164:Luminescent Proteins; D010446:Peptide Fragments; D011964:Receptors, Gastrointestinal Hormone; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D061905:Single-Domain Antibodies; C413663:fluorescent protein 583; C549099:gastric inhibitory polypeptide (1-30); D000090802:rab7 GTP-Binding Proteins; C000716463:rab7 GTP-binding proteins, human; D049452:Green Fluorescent Proteins; D005749:Gastric Inhibitory Polypeptide; C041086:gastric inhibitory polypeptide receptor; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; C406732:GNAS protein, human; D019205:GTP-Binding Protein alpha Subunits, Gs; D020691:rab GTP-Binding Proteins; D000262:Adenylyl Cyclases
keywords: BRET; Early endosomes; FRET; Glucose-dependent-insulinotropic receptor; Internalization; cAMP