Interaction with β-arrestin determines the difference in internalization behavior between β1- and β2-adrenergic receptors

Abstract

The β1-adrenergic receptor (β1AR) shows the resistance to agonist-induced internalization. As β-arrestin is important for internalization, we examine the interaction of β-arrestin with β1AR with three different methods: intracellular trafficking of β-arrestin, binding of in vitro translated β-arrestin to intracellular domains of β1- and β2ARs, and inhibition of βAR-stimulated adenylyl cyclase activities by β-arrestin. The green fluorescent protein-tagged β-arrestin 2 translocates to and stays at the plasma membrane by β2AR stimulation. Although green fluorescent protein-tagged β-arrestin 2 also translocates to the plasma membrane, it returns to the cytoplasm 10-30 min after β1AR stimulation. The binding of in vitro translated β-arrestin 1 and β-arrestin 2 to the third intracellular loop and the carboxyl tail of β1AR is lower than that of β2AR. The fusion protein of β-arrestin 1 with glutathione S-transferase inhibits the β1- and β2AR-stimulated adenylyl cyclase activities, although inhibition of the β1AR-stimulated activity requires a higher concentration of the fusion protein than that of the β2AR-stimulated activity. These results suggest that weak interaction of β1AR with β-arrestins explains the resistance to agonist-induced internalization. This is further supported by the finding that β-arrestin can induce internalization of β1AR when β-arrestin 1 does not dissociate from β1AR by fusing to the carboxyl tail of β1AR.