The lutropin/chorionicgonadotropin receptor (LHR) is a member of the rhodopsin-like subfamily of G protein-coupled receptors (GPCRs) that have been shown to mediate the internalization of its five (activation: three; inactivation: two) naturally occurring mutation. Gonadotropin receptors are members of the seven transmembrane (TM) receptor families. Several point mutations in TM II, III, V and VI have been identified in the luteinizing hormone receptor (LHR) gene, leading to constitutive activation and inactivation of the receptor. In eelLHR, we generated 3 types of constitutive activating mutations (M410T, L469R and D590Y) and 2 types of constitutive inactivating mutations (D383N and Y546F) to investigate how they work on hormone-receptor interaction. To assess the functional effects of 5 receptor mutations directly, wild-type (WT) and mutant receptors were transiently expressed in CHO-K1 cells. We evaluated the basal and cAMP stimulation by rec-LH hormone. The activity was shown to be a dose-dependent increase in cAMP production in LHR-WT expressing cells with an EC50 of 24.3 ng/ml and basal cAMP level of 2.6 nM. However, three activation mutants (D590Y, L469R and M410T) was most elevated the basal cAMP response at 12.8, 21.7 and 6.1 nM, respectively. In two inactivation mutants (D383N and Y546F) are very low in the basal cAMP activation. The EC50 was also considerably decreased to 42.3 ng/ml and 1181 ng/ml, respectively.

The lutropin/choriogonadotropin receptor (LHR) is a member of the rhodopsin-like subfamily of G protein coupled receptor (GPCRs), that has been shown to mediate the internalization of its two naturally occurring agonist, lutropin and choriogonadotropin (CG). The clustered agonist-receptor complex is internalized by a dynamin-dependent pathway and traverses the endosomal compartment without agonist dissociation Dissociation of the agonist-receptor complex occurs in the lysosomes, where both the agonist and receptor are degrade. Recently, constitutively activating mutations of the receptor have been identified that are associated with familial male-precocious puberty (FMPP). A FMPP is a form of sexual precocious puberty in boys in which testosterone levels are elevated independent of changes in luteinizing hormone-releasing hormone and serum luteinizing hormone levels, We have now analyzed two naturally occurring, constitutively active mutants of the human LHR. These mutations were introduced into the rat LHR (rLHR) and are designated L435R and D556Y. Cells expressing rLHR-D556Y bind human choriogonadotropin (hCG) with normal affinity, exhibit a 25-fold increase in basal cAMP and respond to hCG with a normal increase in cAMP accumulation. Cells expressing rLHR-L435R also bind hCG with normal affinity, exhibit a 47-fold increase in basal cAMP, and do not respond to hCG with a further increase in cAMP accumulation. This mutation enhances the internalization of the free and agonist-occupied receptors ~2- and ~17- fold, respectively We conclude that the state of activation of the rLHR can modulate its basal and/or agonist-stimulated internalization. Since the internalization of hCG is involved in the termination of hCG actions, we suggest that the lack of responsiveness detected in cells expressing rLHR-L435R is due to the fast rate of internalization of the bound hCG. The finding that membranes expressing rLHR-L435R respond to hCG with an increase in adenylyl cyclase activity supports this suggestion. Autonomous Leydig cell activity in FMPP is caused by a constitutively activating LH/CGR.