Molecular Basis for Subtype-specific Desensitization of Inhibitory Adenosine Receptors

The differing effects of short-term agonist exposure on the two inhibitory adenosine receptor (AR) subtypes have been examined using Chinese hamster ovary cells stably expressing the hemagglutinin epitope-tagged human A1AR and rat A3AR. Under conditions in which exposure of transfected cells to 5 microM (-)-(R)-N6-(phenylisopropyl)adenosine resulted in the functional desensitization and phosphorylation of the A3AR, neither property was exhibited by the A1AR. However, a stably expressed chimeric A1-A3AR, termed A1CT3AR, in which the C-terminal domain of the A1AR distal to its predicted palmitoylation site was replaced by the corresponding region of the A3AR, was able to undergo functional desensitization and agonist-stimulated phosphorylation in a manner similar to that exhibited by the A3AR. Moreover, purified G-protein-coupled receptor kinases 2, 3, and 5 were each capable of enhancing the agonist-dependent phosphorylation of the A3AR and A1CT3AR in vitro. Taken together, these data demonstrate that the C-terminal domain of the A3AR distal to its predicted palmitoylation site is responsible for this receptor's ability to undergo a rapid agonist-dependent desensitization and are consistent with a model in which phosphorylation of the A3AR within this domain by one or more G-protein-coupled receptor kinases initiates the desensitization process.