Effect of different G protein-coupled receptor kinases on phosphorylation and desensitization of the alpha sub(1B)-adrenergic receptor

The alpha sub(1B)-adrenergic receptor ( alpha sub(1B)AR), its truncated mutant T368, different G protein-coupled receptor kinases (GRK) and arrestin proteins were transiently expressed in COS-7 or HEK293 cells alone and/or in various combinations. Coexpression of beta -adrenergic receptor kinase ( beta ARK) 1 (GRK2) or 2 (GRK3) could increase epinephrine-induced phosphorylation of the wild type alpha sub(1B)AR above basal as compared to that of the receptor expressed alone. On the other hand, overexpression of the dominant negative beta ARK (K220R) mutant impaired agonist-induced phosphorylation of the receptor. Overexpression of GRK6 could also increase epinephrine-induced phosphorylation of the receptor, whereas GRK5 enhanced basal but not agonist-induced phosphorylation of the alpha sub(1B)AR. Increasing coexpression of beta ARK1 or beta ARK2 resulted in the progressive attenuation of the alpha sub(1B)AR-mediated response on polyphosphoinositide (PI) hydrolysis. However, coexpression of beta ARK1 or 2 at low levels did not significantly impair the PI response mediated by the truncated alpha sub(1B)AR mutant T368, lacking the C terminus, which is involved in agonist-induced desensitization and phosphorylation of the receptor. Similar attenuation of the receptor-mediated PI response was also observed for the wild type alpha sub(1B)AR, but not for its truncated mutant, when the receptor was coexpressed with beta -arrestin 1 or beta -arrestin 2. Despite their pronounced effect on phosphorylation of the alpha sub(1B)AR, overexpression of GRK5 or GRK6 did not affect the receptor-mediated response. In conclusion, our results provide the first evidence that beta ARK1 and 2 as well as arrestin proteins might be involved in agonist-induced regulation of the alpha sub(1B)AR. They also identify the alpha sub(1B)AR as a potential phosphorylation substrate of GRK5 and GRK6. However, the physiological implications of GRK5- and GRK6-mediated phosphorylation of the alpha sub(1B)AR remain to be elucidated.