title: Agonist-induced internalization and recycling of the glucagon-like peptide-1 receptor in transfected fibroblasts and in insulinomas.
abstract: Glucagon-like peptide-1 (GLP-1) is the most potent stimulator of glucose-induced insulin secretion and its pancreatic beta-cell receptor is a member of a new subfamily of G-protein-coupled receptors which includes the receptors for vasoactive intestinal polypeptide, secretin and glucagon. Here we studied agonist-induced GLP-1 receptor internalization in receptor-transfected Chinese hamster lung fibroblasts using three different approaches. First, iodinated GLP-1 bound at 4 degrees C to transfected cells was internalized with a t 1/2 of 2-3 min following warming up of the cells to 37 degrees C. Secondly, exposure to GLP-1 induced a shift in the distribution of the receptors from plasma membrane-enriched to endosomes-enriched membrane fractions, as assessed by Western blot detection of the receptors using specific antibodies. Thirdly, continuous exposure of GLP-1 receptor-expressing cells to iodinated GLP-1 led to a linear accumulation of peptide degradation products in the medium following a lag time of 20-30 min, indicating a continuous cycling of the receptor between the plasma membrane and endosomal compartments. Potassium depletion and hypertonicity inhibited transferrin endocytosis, a process known to occur via coated pit formation, as well as GLP-1 receptor endocytosis. In contrast to GLP-1, the antagonist exendin-(9-39) did not lead to receptor endocytosis. Surface re-expression following one round of GLP-1 receptor endocytosis occurred with a half-time of about 15 min. The difference in internalization and surface re-expression rates led to a progressive redistribution of the receptor in intracellular compartments upon continuous exposure to GLP-1. Finally, endogenous GLP-1 receptors expressed by insulinoma cells were also found to be internalized upon agonist binding. Together our data demonstrate that the GLP-1 receptor is internalized upon agonist binding by a route similar to that taken by single transmembrane segment receptors. The characterization of the pathway and kinetics of GLP-1-induced receptor endocytosis will be helpful towards understanding the role of internalization and recycling in the control of signal transduction by this receptor.
authors.affiliation: Department of Pharmacology and Toxicology, University of Lausanne, Switzerland.
authors.forename: C
authors.initials: C
authors.lastname: Widmann
authors.forename: W
authors.initials: W
authors.lastname: Dolci
authors.forename: B
authors.initials: B
authors.lastname: Thorens
mesh_terms: D000818:Animals; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D002999:Clone Cells; D003034:Coated Pits, Cell-Membrane / Q000378:metabolism; D006224:Cricetinae; D004705:Endocytosis; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D000067757:Glucagon-Like Peptide-1 Receptor; D006801:Humans; D007340:Insulinoma / Q000378:metabolism; D007457:Iodine Radioisotopes; D007700:Kinetics; D008024:Ligands; D008954:Models, Biological; D010446:Peptide Fragments / Q000494:pharmacology; D018027:Receptors, Glucagon / Q000819:agonists* / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: C000593833:GLP1R protein, human; D000067757:Glucagon-Like Peptide-1 Receptor; D007457:Iodine Radioisotopes; D008024:Ligands; D010446:Peptide Fragments; D018027:Receptors, Glucagon; C083773:exendin (9-39)