Low affinity of beta1-adrenergic receptor for beta-arrestins explains the resistance to agonist-induced internalization.
It has been reported that beta-arrestin is essential for the internalization of many G protein-coupled receptors. Since beta1-adrenergic receptor (beta1AR) shows the resistance to agonist-induced internalization, we examine the interaction of beta-arrestin with beta1AR with three different approaches: translocation of beta-arrestin to the plasma membrane, direct binding of in vitro translated beta-arrestin to intracellular domains of beta1- and beta2ARs, inhibition of beta1- and beta2AR-stimulated adenylyl cyclase activities by beta-arrestin. The enhanced green fluorescent protein (EGFP)-tagged beta-arrestin 2 (beta-arrestin 2-GFP) translocates to and stays at the plasma membrane by beta2AR stimulation. Beta-arrestin 2-GFP also translocates to the plasma membrane upon beta1AR stimulation. However, it returns to the cytoplasm 10 - 30 min after agonist stimulation. The amount of beta-arrestin bound to the third intracellular loop and the carboxyl tail of beta1AR is lower than that of beta2AR. The fusion protein of beta-arrestin 1 with glutathione-S-transferase inhibits the beta1- and beta2AR-stimulated adenylyl cyclase activities. However, inhibition of the beta1AR-stimulated activity requires a higher amount of the fusion protein than that of the beta2AR-stimulated activity. These results suggest that affinity of beta1AR for beta-arrestins is lower than that of beta2AR, and explains the resistance to agonist-induced internalization. This conclusion is further supported by the finding that beta-arrestin can induce internalization of beta1AR when beta-arrestin 1 fused to the carboxyl tail of beta1AR.
Shiina
T
T
Laboratory of Pharmacology and Toxicology, Graduate School of Pharmaceutical Sciences, University of Tokyo, Japan.
Nagao
T
T
Kurose
H
H
D000067956:Adenylyl Cyclase Inhibitors; D000262:Adenylyl Cyclases / Q000378:metabolism; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D019390:Arrestins / Q000378:metabolism*; D001665:Binding Sites / Q000187:drug effects; D005982:Glutathione Transferase / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D007545:Isoproterenol / Q000494:pharmacology; D008164:Luminescent Proteins / Q000378:metabolism; D018613:Microscopy, Confocal; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D018342:Receptors, Adrenergic, beta-1 / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D000071559:beta-Arrestin 1; D000071560:beta-Arrestin 2; D000071557:beta-Arrestins
C000605253:ARRB1 protein, human; C000605254:ARRB2 protein, human; D000067956:Adenylyl Cyclase Inhibitors; D000318:Adrenergic beta-Agonists; C000605256:Arrb1 protein, rat; C000605257:Arrb2 protein, rat; D019390:Arrestins; D008164:Luminescent Proteins; D018342:Receptors, Adrenergic, beta-1; D011993:Recombinant Fusion Proteins; D000071559:beta-Arrestin 1; D000071560:beta-Arrestin 2; D000071557:beta-Arrestins; D049452:Green Fluorescent Proteins; D005982:Glutathione Transferase; D000262:Adenylyl Cyclases; D007545:Isoproterenol