Agonist-induced translocation of the kinin B(1) receptor to caveolae-related rafts
| Title | Agonist-induced translocation of the kinin B(1) receptor to caveolae-related rafts |
| Publication Type | Journal Article |
| Year of Publication | 2002 |
| Authors | Sabourin, T, Bastien L, Bachvarov DR, Marceau F |
| Journal | Mol Pharmacol |
| Volume | 61 |
| Pagination | 546-53 |
| Date Published | Mar |
| ISBN Number | 0026-895X (Print)0026-895X (Linking) |
| Accession Number | 11854434 |
| Keywords | *beta-Cyclodextrins, Animals, Antigens, Human Platelet/analysis, Bacterial Proteins/metabolism, Biological Transport/drug effects, Bradykinin/metabolism, Caveolae/drug effects/*metabolism, Caveolin 1, Caveolins/metabolism, Cell Fractionation, Cells, Cultured, COS Cells, Cyclodextrins/pharmacology, Endocytosis/physiology, Green Fluorescent Proteins, Humans, Kallidin/*analogs & derivatives/*pharmacology, Luminescent Proteins/metabolism, Membrane Microdomains/drug effects/*metabolism, Rabbits, Receptor, Bradykinin B1, Receptors, Bradykinin/agonists/*metabolism, Subcellular Fractions, Transfection, Tritium |
| Abstract | The kallikrein-kinin system, activated during inflammatory conditions and the regulation of specific cardiovascular and renal functions, includes two G protein-coupled receptors for bradykinin (BK)-related peptides. The B(1) receptor (B(1)R) subtype is not believed to undergo agonist-induced phosphorylation and endocytosis. A conjugate made of the rabbit B(1)R fused with the yellow variant of green fluorescent protein (YFP) was expressed in mammalian cells. In COS-1 or human embryonic kidney (HEK) 293 cells, the construction exhibited a nanomolar affinity for the agonist radioligand [(3)H]Lys-des-Arg(9)-BK or the antagonist ligand [(3)H]Lys-[Leu(8)]des-Arg(9)-BK and a pharmacological profile virtually identical to that of wild-type B(1)R. Lys-des-Arg(9)-BK stimulation of HEK 293 cells stably expressing B(1)R-YFP but not stimulation of untransfected cells released [(3)H]arachidonate in a phospholipase A(2) assay. B(1)R-YFP was visualized as a continuous labeling of the plasma membranes in stably transfected HEK 293 cells (confocal microscopy). Addition of Lys-des-Arg(9)-BK (1-100 nM) rapidly concentrated the receptor-associated fluorescence into multiple aggregates that remained associated with the plasma membrane (no significant internalization) and colocalized with caveolin-1. This reaction was slowly reversible upon agonist washing at 37 degrees C and prevented pretreatment with a B(1)R antagonist. beta-Cyclodextrin treatment, which extracts cholesterol from membranes and disrupts caveolae-related rafts, prevented agonist-induced redistribution of B(1)R-YFP but not the PLA(2) activation mediated by this receptor. The agonist radioligand copurified with caveolin-1 to a greater extent than the tritiated antagonist in buoyant fractions of HEK 293 cells treated with the ligands. Agonist-induced cellular translocation of the kinin B(1)R to caveolae-related rafts without endocytosis is a novel variation on the theme of G protein-coupled receptor adaptation. |
| URL | http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11854434 |