DEPENDENCE OF AGONIST ACTIVATION ON AN AROMATIC MOIETY IN THE DPLIY MOTIF OF THE GONADOTROPIN-RELEASING-HORMONE RECEPTOR

In the GnRH receptor, the NPX(2-3)Y motif that is present in the seven th transmembrane helix of most G protein-coupled receptors is unusual in containing Asp instead of Asn but retains the highly conserved Tyr residue, The importance of this aromatic residue in the DPLIY sequence of the GnRH receptor function was analyzed by replacing Tyr(322) with Ala or Phe residues, The Y(322)A mutant receptor expressed in COS-7 c ells had high agonist binding affinity, but its ability to interact wi th G protein(s) and to activate inositol phosphate production in respo nse to GnRH was abolished. Although functionally inactive, the Y(322)A , mutant receptor was internalized at about 50% of the rate of the wil d type receptor in agonist-treated cells, When Tyr(322) was replaced w ith Phe to preserve its aromatic nature, the (YF)-F-322 mutant recepto r displayed normal G protein activation and inositol phosphate respons es to GnRH and was internalized in the same manner as the wild type re ceptor, These findings demonstrate that the aromatic moiety of the Tyr (322) component of the DPLIY motif in the GnRH receptor is a critical determinant of agonist-induced receptor activation and signal transduc tion.