Involvement of Tyrosine Residues Located in the Carboxyl Tail of the Human β2-Adrenergic Receptor in Agonist-Induced Down-Regulation of the Receptor

Chronic exposure of various cell types to adrenergic agonists leads to a decrease in cell surface β2-adrenergic receptor (β2AR) number. Sequestration of the receptor away from the cell surface as well as a down-regulation of the total number of cellular receptors are believed to contribute to this agonist-mediated regulation of receptor number. However, the molecular mechanisms underlying these phenomena are not well characterized. Recently, tyrosine residues located in the cytoplasmic tails of several membrane receptors, such as the low density lipoprotein and mannose-6-phosphate receptors, have been suggested as playing an important role in the agonist-induced internalization of these receptors. Accordingly, we assessed the potential role of two tyrosine residues in the carboxyl tail of the human β2AR in agonist-induced sequestration and down-regulation of the receptor. Tyr-350 and Tyr-354 of the human β2AB were replaced with alanine residues by site-directed mutagenesis and both wild-type and mutant β2AR were stably expressed in transformed Chinese hamster fibroblasts. The mutation dramatically decreased the ability of the β2AR to undergo isoproterenol-induced down-regulation. However, the substitution of Tyr-350 and Tyr-354 did not affect agonist-induced sequestration of the receptor. These results suggest that tyrosine residues in the cytoplasmic tail of human β2AR are crucial determinants involved in its down-regulation.