BETA(2)-ADRENERGIC RECEPTOR DESENSITIZATION, INTERNALIZATION, AND PHOSPHORYLATION IN RESPONSE TO FULL AND PARTIAL AGONISTS

Previous studies indicated that partial agonists cause less desensitiz ation of the beta(2)-adrenergic receptor (beta AR) than full agonists; however, the molecular basis for this in intact cells has not been in vestigated, In the present work, we have determined the rates of desen sitization, internalization, and phosphorylation caused by a series of beta AR agonists displaying a 95-fold range of coupling efficiencies, These studies were performed with HEK-293 cells overexpressing the be ta AR with hemagglutinin and 6-histidine epitopes introduced into the N and C termini, respectively, This modified beta AR behaved identical ly to the wild type receptor with regard to agonist K-d, coupling effi ciency, and desensitization. The coupling efficiencies for beta AR ago nist activation of adenylyl cyclase relative to epinephrine (100%) wer e 4.2% for fenoterol, 4.9% for albuterol, 2.5% for dobutamime, and 1.1 % for ephedrine, At concentrations of these agonists yielding >90% rec eptor occupancy, the rate and extent (0-30 min) of agonist-induced des ensitization of beta AR activation of adenylyl cyclase followed the sa me order as coupling efficiency, i.e. epinephrine greater than or equa l to fenoterol > albuterol > dobutamine > ephedrine, The rate of inter nalization of the beta AR with respect to these agonists also followed the same order as the desensitization and exhibited a slight lag, Lik e internalization and desensitization, beta AR phosphorylation exhibit ed a dependence on agonist strength. The two strongest agonists, epine phrine and fenoterol, provoked 11-13-fold increases in the level of be ta AR phosphorylation after just 1 min, whereas the weak agonists dobu tamine and ephedrine caused only 3-4-fold increases, similar to levels induced by cAMP-dependent protein kinase activation with forskolin. W ith longer treatment times, the level of beta AR phosphorylation decli ned with strong agonists, but it progressively increased with the weak er partial agonists, such that after 30 min the -fold elevation with e pinephrine (6.2 +/- 0.82) was not appreciably different from ephedrine (5.0 +/- 0.96) and significantly less than that caused by albuterol ( 10.4 +/- 1.7), In summary, our results demonstrate an excellent propor tionality between the agonist strength and agonist-induced desensitiza tion, internalization, and the rapid initial phase of phosphorylation. The data support the hypothesis that increasing agonist-coupling effi ciency primarily affects desensitization by increasing the rate of bet a ARK phosphorylation of the beta AR.