ENHANCED EXPRESSION OF G-PROTEIN-COUPLED RECEPTOR KINASE-2 SELECTIVELY INCREASES THE SENSITIVITY OF A(2A) ADENOSINE RECEPTORS TO AGONIST-INDUCED DESENSITIZATION

1 G protein-coupled receptor kinases (GRKs) are thought to be importan t in mediating the agonist-induced phosphorylation and consequent dese nsitization of G protein-coupled receptor (GPCR) responses. We have pr eviously shown that stable expression of a dominant negative mutant G protein-coupled receptor kinase 2 (GRK2) construct in NG108-15 mouse n euroblastoma x rat glioma cells suppresses the agonist-induced desensi tization of A(2A) and A(2B) adenosine receptor-stimulated adenylyl cyc lase activity (Mundell et al., 1997). To further determine the role of GRK2 in agonist-induced desensitization of these adenosine receptors, we stably overexpressed wild type GRK2 in NG108-15 cells. 2 In homoge nates prepared from cells overexpressing GRK2, the acute stimulation o f adenylyl cyclase by activation of A(2A) and A(2B) adenosine receptor s was markedly reduced, but could be reversed by pretreating the cells with AD (adenosine deaminase), to remove extracellular adenosine from the medium. On the other hand, acute stimulation of adenylyl cyclase by secretin, iloprost, NaF and forskolin was the same in GRK2 overexpr essing cells and plasmid-transfected control cells. 3 Cells overexpres sing GRK2 were more sensitive to adenosine receptor agonist-induced de sensitization than plasmid-transfected control cells. This effect was selective since the agonist sensitivity of desensitization for secreti n and IP-prostanoid receptor-stimulated adenylyl cyclase activity was not affected by GRK2 overexpression. 4 These results further implicate GRKZ as the likely mechanism by which A(2) adenosine receptors underg o short-term desensitization in NG108-15 cells, and indicate that even when overexpressed, GRK2 retains its substrate specificity for native receptors in intact cells. Furthermore, the susceptibility of GPCRs t o desensitization appears to depend on the level of GRK expression, su ch that in cells that express high levels of GRK2, low agonist concent rations may be sufficient to trigger GRK-mediated desensitization.