Cloning and characterization of the human GnRH receptor.
A cDNA encoding the human GnRH receptor (GnRHR) has been cloned and functionally expressed in both Xenopus oocytes and COS-1 cells. The 2160 bp cDNA encodes a 328 amino acid protein with a predicted amino acid sequence that is 90% identical to that of the mouse GnRHR (Tsutsumi et al. (1992) Mol. Endocrinol. 6, 1163-1169). Injection of synthetic RNA transcript into oocytes led to the development of a depolarizing response to agonists when assayed by voltage-clamp electrophysiology. Consistent with the expression of a mammalian GnRHR, the response was blocked by GnRH antagonists. Following expression of the human GnRHR in COS-1 cells, agonists and an antagonist displaced [125I]GnRH agonist from membrane isolates with nanomolar range dissociation constants similar to those described for displacement from human pituitary membranes. Transfected COS-1 cells manifested a GnRH-stimulated increase in phosphoinositol turnover, with an EC50 of approximately 3 nM, which was inhibited by GnRH antagonists. Northern blot analysis revealed a single band of approximately 4.7 kb expressed in human pituitary which was not detected in testis. The predicted structure of the human GnRHR is similar to that previously reported for the mouse receptor. Although the mammalian GnRHR is a seven transmembrane domain receptor, it differs from other G-protein coupled receptors in several respects, most notably the lack of a cytoplasmic C-terminal domain. The present study demonstrates that the cDNA isolated encodes the human GnRHR and suggests that several unique features conserved among mammalian GnRHRs may be essential for receptor function and/or regulatory control.
Fishberg Research Center in Neurobiology, Mount Sinai School of Medicine, New York, NY 10029.
L
L
Chi
W
W
Zhou
A
A
Prikhozhan
C
C
Flanagan
J S
JS
Davidson
M
M
Golembo
N
N
Illing
R P
RP
Millar
S C
SC
Sealfon
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry / Q000235:genetics; D005260:Female; D015870:Gene Expression; D006801:Humans; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011966:Receptors, LHRH / Q000737:chemistry / Q000235:genetics*; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D014981:Xenopus
D011966:Receptors, LHRH; D011994:Recombinant Proteins; D004247:DNA