Two mutations of the lutropin/choriogonadotropin receptor that impair signal transduction also interfere with receptor-mediated endocytosis.
The experiments presented herein were designed to probe a potential role for the activation of the LH/CG receptor (LHR) on the receptor-mediated endocytosis of human CG (hCG). Two mutants of the rat LHR (rLHR) that bind the hormone with high affinity but are deficient in signal transduction were prepared by mutating highly conserved residues that have been previously shown to be important in signal transduction in other members of the G protein-coupled receptor family. Mutation of a highly conserved aspartic acid in the second transmembrane domain of the rLHR (designated rLHR-D383N) does not affect hCG binding but impairs signal transduction. When compared with cells expressing an equivalent density of wild type rLHR (rLHR-wt), concentration-response curves for the hCG-stimulated cAMP accumulation in cells expressing rLRH-D383N- are characterized by an 18-fold increase in the EC50 but no change in the maximal response. Cells expressing rLHR-D383N also display a 4- to 5-fold increase in the half-life of internalization of hCG. Mutation of a highly conserved arginine in the second intracellular loop of the rLHR (designated rLHR-R442H) also does not affect hCG binding but impairs signal transduction. When compared with cells expressing an equivalent density of rLHR-wt, concentration-response curves for the hCG-stimulated cAMP accumulation in cells expressing rLHR-R442H are characterized by a 7-fold increase in the EC50 and a 6- to 10-fold decrease in the maximal response. Cells expressing rLHR-R442H also display a 1.5- to 2-fold increase in the half-life of internalization of hCG. These results, together with the finding that an antagonist of hCG is internalized more slowly than hCG, suggest that the activation of the LHR is needed for the efficient endocytosis of the bound hCG.
Dhanwada
K R
KR
Department of Pharmacology, University of Iowa College of Medicine, Iowa City 52242-1109, USA.
Vijapurkar
U
U
Ascoli
M
M
D001120:Arginine; D001216:Asparagine; D001224:Aspartic Acid; D002460:Cell Line; D006063:Chorionic Gonadotropin / Q000378:metabolism / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D004622:Embryo, Mammalian; D004705:Endocytosis; D006639:Histidine; D006801:Humans; D007668:Kidney; D016296:Mutagenesis; D011487:Protein Conformation; D011974:Receptors, LH / Q000737:chemistry / Q000235:genetics* / Q000502:physiology*; D015398:Signal Transduction; D014162:Transfection
D006063:Chorionic Gonadotropin; D011974:Receptors, LH; D001224:Aspartic Acid; D006639:Histidine; D001216:Asparagine; D001120:Arginine; D000242:Cyclic AMP