title: Chaperoning of the A1-adenosine receptor by endogenous adenosine - an extension of the retaliatory metabolite concept.
abstract: Cell-permeable orthosteric ligands can assist folding of G protein-coupled receptors in the endoplasmic reticulum (ER); this pharmacochaperoning translates into increased cell surface levels of receptors. Here we used a folding-defective mutant of human A1-adenosine receptor as a sensor to explore whether endogenously produced adenosine can exert a chaperoning effect. This A1-receptor-Y(288)A was retained in the ER of stably transfected human embryonic kidney 293 cells but rapidly reached the plasma membrane in cells incubated with an A1 antagonist. This was phenocopied by raising intracellular adenosine levels with a combination of inhibitors of adenosine kinase, adenosine deaminase, and the equilibrative nucleoside transporter: mature receptors with complex glycosylation accumulated at the cell surface and bound to an A1-selective antagonist with an affinity indistinguishable from the wild-type A1 receptor. The effect of the inhibitor combination was specific, because it did not result in enhanced surface levels of two folding-defective human V2-vasopressin receptor mutants, which were susceptible to pharmacochaperoning by their cognate antagonist. Raising cellular adenosine levels by subjecting cells to hypoxia (5% O2) reproduced chaperoning by the inhibitor combination and enhanced surface expression of A1-receptor-Y(288)A within 1 hour. These findings were recapitulated for the wild-type A1 receptor. Taken together, our observations document that endogenously formed adenosine can chaperone its cognate A1 receptor. This results in a positive feedback loop that has implications for the retaliatory metabolite concept of adenosine action: if chaperoning by intracellular adenosine results in elevated cell surface levels of A1 receptors, these cells will be more susceptible to extracellular adenosine and thus more likely to cope with metabolic distress.
authors.affiliation: Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria.
authors.forename: Justyna
authors.initials: J
authors.lastname: Kusek
authors.affiliation: Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria.
authors.forename: Qiong
authors.initials: Q
authors.lastname: Yang
authors.affiliation: Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria.
authors.forename: Martin
authors.initials: M
authors.lastname: Witek
authors.affiliation: Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria.
authors.forename: Christian W
authors.initials: CW
authors.lastname: Gruber
authors.affiliation: Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria.
authors.forename: Christian
authors.initials: C
authors.lastname: Nanoff
authors.affiliation: Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria michael.freissmuth@meduniwien.ac.at.
authors.forename: Michael
authors.initials: M
authors.lastname: Freissmuth
mesh_terms: D000241:Adenosine / Q000378:metabolism*; D058916:Adenosine A1 Receptor Antagonists / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D018832:Molecular Chaperones / Q000378:metabolism; D009154:Mutation; D010950:Plasma Cells / Q000378:metabolism; D017510:Protein Folding; D043682:Receptor, Adenosine A1 / Q000235:genetics* / Q000378:metabolism*; D013312:Stress, Physiological / Q000187:drug effects
chemical_list: D058916:Adenosine A1 Receptor Antagonists; D004791:Enzyme Inhibitors; D018832:Molecular Chaperones; D043682:Receptor, Adenosine A1; D000241:Adenosine