Involvement of the Amino Terminus of the B₂ Receptor in Agonist-induced Receptor Dimerization*

Abstract

The mechanisms and the functional importance of G-protein-coupled receptor dimerization are poorly understood. We therefore analyzed dimerization of the bradykinin B₂receptor. The binding of the agonist bradykinin to the B₂receptor endogenously expressed on PC-12 cells led to the formation of receptor dimers, whereas the B₂ antagonist HOE140 did not induce dimerization, suggesting that B₂ receptor dimerization was linked to receptor activation. Addition of a peptide corresponding to the amino terminus of the receptor reduced the amount of detected B₂ receptor dimers, whereas peptides derived from the extracellular loops had no effect. To further analyze the role of the amino terminus of the receptor in receptor dimerization, we created two different rat B₂ receptor variants with truncated amino termini, B₂ ⁵³ and B₂ ⁶⁵, starting at amino acids 53 and 65. In contrast to the wild-type B₂ receptor and to B₂ ⁵³, bradykinin did not induce dimerization of the B₂ ⁶⁵ receptor. Both receptor variants were similar to the wild-type B₂ receptor with respect to agonist binding and signal generation. However, B₂ ⁶⁵ was not phosphorylated, did not desensitize, and was not downregulated upon bradykinin stimulation. Likewise, antibodies directed to the amino terminus of the receptor partially reduced internalization of [³H]bradykinin on PC-12 cells. These findings suggest that the amino terminus of the B₂ receptor is necessary for triggering agonist-induced B₂ receptor dimerization, and receptor dimers are involved in receptor-mediated signal attenuation.