Addition of catfish gonadotropin-releasing hormone (GnRH) receptor intracellular carboxyl-terminal tail to rat GnRH receptor alters receptor expression and regulation

Mammalian GnRH receptor (GnRHR) is unique among G protein-coupled seven-transmembrane segment receptors due to the absence of an intracellular C-terminal tail frequently important for internalization and/or desensitization of other G protein-coupled receptors. The recent cloning of nonmammalian (i.e. catfish, goldfish, frog, and chicken) GnRHRs shows that these contain an intracellular C terminus. Addition of the 51-amino acid intracellular C terminus from catfish GnRHR (cfGnRHR) to rat GnRHR (rGnRHR) did not affect rGnRHR binding affinity but elevated receptor expression by about 5-fold. Truncation of the added C terminus impaired the elevated receptor-binding sites by 3- to 8-fold, depending on the truncation site. In addition, introducing the C terminus to rGnRHR altered the pattern of receptor regulation from biphasic down-regulation and recovery to monophasic down-regulation, The extent of downregulation was also enhanced. The alteration in receptor regulation due to the addition of a C terminus was reversed by truncation of the added C terminus. Furthermore, addition of the cfGnRHR C terminus to rGnRHR significantly augmented the inositol phospholipid (IP) response of transfected cells to Buserelin, but this did not result from the elevation of receptor-binding sites. Addition of the C terminus did not affect Buserelin-stimulated cAMP and PRL release. GH(3) cells transfected with wild-type cfGnRHR did not show measurable Buserelin binding or significant stimulation of IP, cAMP, or PRL in response to Buserelin (10(-13)-10(-9) M). GH(3) cells transfected with C terminus-truncated cfGnRHR showed no IP response to Buserelin (10(-13)-10(-7) M). These results suggest that addition of the cfGnRHR intracellular C terminus to rGnRHR has a significant impact on rGnRHR expression and regulation and efficiency of differential receptor coupling to G proteins.