title: Agonist-induced internalisation of the glucagon-like peptide-1 receptor is mediated by the Gαq pathway.
abstract: The glucagon-like peptide-1 receptor (GLP-1R) is a G-protein-coupled receptor (GPCR) and an important target in the treatment of type 2 diabetes mellitus (T2DM). Upon stimulation with agonist, the GLP-1R signals through both Gαs and Gαq coupled pathways to stimulate insulin secretion. The agonist-induced GLP-1R internalisation has recently been shown to be important for insulin secretion. However, the molecular mechanisms underlying GLP-1R internalisation remain unknown. The aim of this study was to determine the role of GLP-1R downstream signalling pathways in its internalisation. Agonist-induced human GLP-1R (hGLP-1R) internalisation and activity were examined using a number of techniques including immunoblotting, ELISA, immunofluorescence and luciferase assays to determine cAMP production, intracellular Ca(2+) accumulation and ERK phosphorylation. Agonist-induced hGLP-1R internalisation is dependent on caveolin-1 and dynamin. Inhibition of the Gαq pathway but not the Gαs pathway affected hGLP-1R internalisation. Consistent with this, hGLP-1R mutant T149M and small-molecule agonists (compound 2 and compound B), which activate only the Gαs pathway, failed to induce internalisation of the receptor. Chemical inhibitors of the Gαq pathway, PKC and ERK phosphorylation significantly reduced agonist-induced hGLP-1R internalisation. These inhibitors also suppressed agonist-induced ERK1/2 phosphorylation demonstrating that the phosphorylated ERK acts downstream of the Gαq pathway in the hGLP-1R internalisation. In summary, agonist-induced hGLP-1R internalisation is mediated by the Gαq pathway. The internalised hGLP-1R stimulates insulin secretion from pancreatic β-cells, indicating the importance of GLP-1 internalisation for insulin secretion.
authors.affiliation: Institute of Life Science 1, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK.
authors.forename: Aiysha
authors.initials: A
authors.lastname: Thompson
authors.affiliation: Institute of Life Science 1, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK. Electronic address: k.venkateswarlu@swansea.ac.uk.
authors.forename: Venkateswarlu
authors.initials: V
authors.lastname: Kanamarlapudi
mesh_terms: D004305:Dose-Response Relationship, Drug; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000502:physiology*; D052216:Glucagon-Like Peptide 1 / Q000494:pharmacology*; D000067757:Glucagon-Like Peptide-1 Receptor; D057809:HEK293 Cells; D006801:Humans; D018027:Receptors, Glucagon / Q000819:agonists* / Q000378:metabolism*
chemical_list: C000593833:GLP1R protein, human; D000067757:Glucagon-Like Peptide-1 Receptor; D018027:Receptors, Glucagon; D052216:Glucagon-Like Peptide 1; D043802:GTP-Binding Protein alpha Subunits, Gq-G11
keywords: 2-Aminoethoxydiphenyl borate (2-APB); BAPTA-AM; Dynasore; Endocytosis; G protein-coupled receptor (GPCR); GLP-1; GLP-1 receptor (GLP-1R); Glucagon like peptide-1 (GLP-1); Go6976; Gα(q) pathway.; PBP10; PD98059; Ro318820; U73122; chlorpromazine; compound 2; compound B; filipin; genistein; monodansylcadaverine (MDC); tunicamycin