Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The pattern of side-chain conservation at the cytoplasmic side of the third transmembrane domain of rhodopsin family G protein-coupled receptors, Asp/Glu-Arg-Tyr/X-X-X-Ile/Val, defines a structural "arginine cage" domain. Previous computational and mutagenesis studies of the GnRH receptor indicated an important contribution of local interactions to the function of this domain. We have investigated the functional importance of the intrahelical position and orientation of the arginine cage using insertional mutagenesis. Introduction of a single Ala proximal to the conserved Asp-Arg of this domain caused loss of detectable ligand binding. Inserting a second Ala, however, restored high-affinity agonist binding. Further insertion of three or four Ala residues at this site generated receptors that bound agonist with an affinity 3- to 10-fold higher than that of the wild-type receptor. Loss of detectable coupling to inositol phosphate turnover in all these mutant receptors confirms that the structure required in this region for efficient signaling is highly constrained. In contrast, the recovery of agonist binding with the progressive insertion of two to four Ala residues indicates that specific orientations of this segment can stabilize high-affinity receptor conformations that are uncoupled from signal transduction.
Mol Endocrinol 2001 Mar
PMID:Insertional mutagenesis of the arginine cage domain of the gonadotropin-releasing hormone receptor. 1122 40

In immortalized GnRH neurons, cAMP production is elevated by increased extracellular Ca2+ and the Ca2+ channel agonist, BK-8644, and is diminished by low extracellular Ca2+ and treatment with nifedipine, consistent with the expression of adenylyl cyclase type I (AC I). Potassium-induced depolarization of GT1-7 neurons causes a dose-dependent monotonic increase in [Ca2+]i and elicits a bell-shaped cAMP response. The inhibitory phase of the cAMP response is prevented by pertussis toxin (PTX), consistent with the activation of G(i)-related proteins during depolarization. Agonist activation of the endogenous GnRH receptor in GT1-7 neurons also elicits a bell-shaped change in cAMP production. The inhibitory action of high GnRH concentrations is prevented by PTX, indicating coupling of the GnRH receptors to G(i)-related proteins. The stimulation of cAMP production by activation of endogenous LH receptors is enhanced by low (nanomolar) concentrations of GnRH but is abolished by micromolar concentrations of GnRH, again in a PTX-sensitive manner. These findings indicate that GnRH neuronal cAMP production is maintained by Ca2+ entry through voltage-sensitive calcium channels, leading to activation of Ca2+-stimulated AC I. Furthermore, the Ca2+ influx-dependent activation of AC I acts in conjunction with AC-regulatory G proteins to determine basal and agonist-stimulated levels of cAMP production.
Mol Endocrinol 2001 Mar
PMID:Regulation of Ca2+-sensitive adenylyl cyclase in gonadotropin-releasing hormone neurons. 1122 44

We have used spot fluorescence photobleaching recovery methods to measure the lateral diffusion of GnRH receptor (GnRHR) fused at its C terminus to green fluorescent protein (GFP) after binding of either GnRH agonists or antagonist. Before ligand binding, GnRHR-GFP exhibited fast rates of lateral diffusion (D = 18 +/- 2.8 x 10(-10)cm2 x sec(-1)) and high values for fractional fluorescence recovery (%R) after photobleaching (73 +/- 1%). Increasing concentrations of agonists, GnRH or D-Ala6-GnRH, caused a dose-dependent slowing of receptor lateral diffusion as well as a decreased fraction of mobile receptors. Increasing concentrations of the GnRH antagonist Antide slowed the rate of receptor diffusion but had no effect on the fraction of mobile receptors, which remained high. To determine whether the decrease in %R caused by GnRH agonists was due, in part, to increased receptor self-association, we measured the fluorescence resonance energy transfer efficiency between GnRHR-GFP and yellow fluorescent protein-GNRHR: There was no energy transfer between GnRHR on untreated cells. Treatment of cells with GnRH agonists led to a concentration-dependent increase in the energy transfer between GnRH receptors to a maximum value of 16 +/- 1%. There was no significant energy transfer between GnRH receptors on cells treated with Antide, even at a concentration of 100 nM. These data provide direct evidence that, before binding of ligand, GnRHR exists as an isolated receptor and that binding of GnRH agonists, but not antagonist, leads to formation of large complexes that exhibit slow diffusion and contain receptors that are self-associated.
Mol Endocrinol 2001 May
PMID:Binding of agonist but not antagonist leads to fluorescence resonance energy transfer between intrinsically fluorescent gonadotropin-releasing hormone receptors. 1132 52

Multiple transcription start sites were identified in the human gonadotropin releasing hormone receptor (hGnRHR) gene. Recently, an upstream promoter residing at -1727/-1674, in vicinity of a CAP site at -1673, was characterized. In this report, we elucidated the underlying mechanisms for the regulation of this promoter. Functionally, this promoter was constitutively suppressed by a silencer element (-1673/-1351) situated immediately downstream to it. On the other hand, pituitary adenylate cyclase-activating polypeptide (PACAP), via the cAMP pathway, was found to be the extracellular cue to control the upstream promoter. Following PACAP-27, PACAP-38 (30 nM) and forskolin (25 microM) treatment, there were significant increases in the reporter gene activities. By deletion analysis, the region residing at -1727 to -1577, containing the distal promoter and 97 bp of the silencer was subsequently found to be responsible for PACAP/cAMP induction. To localize the PACAP-dependent cis-acting element(s) within the silencer, block replacement scanning mutation was performed and a hGnRHR gene PACAP-responsive element (GPRE) was identified at -1676/-1648. The actions of PACAPs and forskolin on the GPRE were further evidenced by gel mobility shift assays. There was an increase in protein binding onto this element only after peptide treatment. As GnRH receptor number on gonadotrope cell surface is a key factor in regulating gonadotropin release, the present study provides an insight into the interplay between PACAP and GnRH receptors on pituitary gonadotropes to control human reproductive functions.
Mol Cell Endocrinol 2001 May 15
PMID:Interplay of pituitary adenylate cyclase-activating polypeptide with a silencer element to regulate the upstream promoter of the human gonadotropin-releasing hormone receptor gene. 1136 53

Exposure of Atlantic croaker (Micropogonias undulatus) to the polychlorinated biphenyl mixture (Aroclor 1254, PCB; 1 mg/kg body wt/day for 30 days) during the early-recrudescence phase of the gonadal cycle results in the impairment of LH secretion and gonadal growth. In order to determine whether impairment was due to disruption of the stimulatory GnRH neuroendocrine pathway, we compared various parameters of the GnRH-LH system in early recrudescing vs. spermiating (mature) fish. Seabream GnRH (GnRH) content in the preoptic anterior hypothalamic area (POAH) and pituitary, pituitary GnRH receptor concentrations, and basal and GnRH analog (GnRHa)-induced LH secretion were significantly higher in gonadally mature croaker compared to early-recrudescing fish. In a subsequent experiment, the effects of PCB on the same neuroendocrine indices were investigated during the gonadal recrudescence phase of croaker. PCB exposure during the period of testicular maturation prevented the natural increase in GnRH content in the POAH but not in the pituitary. This finding suggests that PCB may impair GnRH synthesis in the POAH. The number of pituitary GnRH receptors also remained significantly lower in the PCB-exposed group, which was likely due to an impairment of GnRH release. The GnRH content in the POAH, number of pituitary GnRH receptors, and LH secretion in the PCB-exposed group were comparable to those in early-recrudescing fish, suggesting an impairment of normal maturation of the GnRH-LH system during the gonadal recrudescence phase. This impairment may be due to a direct action of PCB on GnRH neurons and/or indirectly via interference with other neurotransmitter pathways that modulate GnRH function.
Comp Biochem Physiol B Biochem Mol Biol 2001 Jun
PMID:Alterations in the GnRH-LH system in relation to gonadal stage and Aroclor 1254 exposure in Atlantic croaker. 1139 57

Perciforms have three forms of gonadotropin-releasing hormone (GnRH) in their brain. All three GnRHs are potent secretogogues for luteinizing hormone (LH) from the pituitary. The pivotal role of GnRH-R-GnRH interactions in reproductive homeostasis is well established; however, there is a paucity of information on how a GnRH-R responds to the three endogenous GnRH forms in a perciform species. In this study, a recombinant pituitary GnRH-R from striped bass (stb) was expressed in a mammalian cell line (COS-7) and a fish cell line (CHSE-214). Activation of the signaling pathways was monitored by reporter gene (luciferase) based assays, which were specific for cAMP-PKA or Ca 2+/calmodulin kinase (activated via c-fos promoter) signaling pathways. The stbGnRH-R expressed in two different cell lines triggered different downstream signaling in response to the treatments with chicken (c) GnRH II. Interestingly, when endogenous GnRHs were used in combinations, the luciferase activity was significantly attenuated in transfected CHSE-214 cells.
Comp Biochem Physiol B Biochem Mol Biol 2001 Jun
PMID:Recombinant perciform GnRH-R activates different signaling pathways in fish and mammalian heterologous cell lines. 1139 71

Gonadotropin and GnRH receptors belong to the family of G protein coupled receptors. Gain of function mutations have been described, yielding constitutively active receptors. In the case of the LH receptor these dominant mutations determine familial male limited precocious puberty. Somatic mutations of this receptor may in some cases provoke Leydig-cell adenomas. The constitutive LH receptor is not associated with female precocious puberty. Inactivating mutations are recessive. Alterations in the GnRH receptor determine hypogonadotropic hypogonadism. The clinical diagnosis of this etiology of hypogonadism is extremely difficult, especially in sporadic cases. Mutations of gonadotropin receptors determine primary amenorrhea in girls, whereas in boys they are responsible for Leydig cell aplasia or hypoplasia (LH receptor) or of a variable alteration of spermatogenesis (FSH receptor). Mutations provoking only partial alterations of receptor functions are relatively more frequent, than those inducing complete receptor inactivity. They provide interesting insights into the physiology of GnRH and gonadotropin action.
Mol Cell Endocrinol 2001 Jun 20
PMID:Inherited disorders of GnRH and gonadotropin receptors. 1142 Jan 33

Pulsatile GnRH acts at the GnRH receptor on gonadotropes to stimulate gonadotropin gene expression, hormone synthesis and secretion. The pituitary gonadotropins, LH and FSH, stimulate steroid production and gametogenesis in males and in females. Gonadotropin production thus requires the normal development and function of hypothalamic GnRH-producing neurons and pituitary gonadotrope cells. Genes involved in gonadotrope development and/or gene expression include SF1, DAX1, KAL, GNRHR, PC1, HESX1, LHX3, PROP1, LH beta, and FSH beta. Given the complex control of gonadotropin biosynthesis and secretion, it is not surprising that genetic abnormalities have been identified at several of these steps. Some of the mutations that will be reviewed include: (1) SF1 and DAX1-orphan nuclear receptors that are expressed at multiple levels throughout the reproductive axis; (2) KAL-X-linked Kallmann syndrome, where there is abnormal development of hypothalamic GnRH-producing neurons; (3) PC1-causing abnormal processing of GnRH and GNRHR mutations that impair action at the GnRH receptor; (4) HESX1, LHX3, PROP1-abnormal development/function of the gonadotrope cell lineage; (5) LH beta and FSH beta-mutations in the gonadotropin genes that cause structural abnormalities in the hormones. Although all of these gene defects lead to gonadotropin deficiency, each disorder is associated with unique phenotypic or hormonal features. Characterization of the molecular basis of gonadotropin deficiency is useful for directing therapy and for genetic counseling. Identification of these mutations also provides insight into the pathways that govern reproduction.
Mol Cell Endocrinol 2001 Jun 20
PMID:Inherited disorders of the gonadotropin hormones. 1142 Jan 32

Mammalian gonadotropin-releasing hormone (GnRH) receptors preferentially bind mammalian GnRH, which has Arg in position eight. The Glu(7.32(301)) residue, which determines selectivity of the mouse GnRH receptor for Arg(8)-containing GnRH, is Asp(7.32(302)) in the human GnRH receptor. We have confirmed that Asp(7.32(302)) confers selectivity of the human GnRH receptor for Arg(8) of GnRH and investigated the mechanism of this specificity using site-directed mutagenesis and ligand modification. We find that although Arg(8) and Asp(7.32(302)) are required for high-affinity binding of GnRH, conformationally constrained peptides, with D-amino acid substitutions in position six or with a 6,7 gamma-lactam, bind the human GnRH receptor with high affinity, which is independent of the presence of Asp(7.32(302)) in the receptor or Arg(8) in the ligand. The ability of the ligand constraints to compensate for the absence of both Arg(8) and Asp(7.32(302)) indicates that these residues both have roles in stabilizing a high affinity ligand conformation and that their roles are complementary. This suggests that the Arg(8) and Asp(7.32(302)) side chains interact to induce a high affinity conformation of native GnRH. Thus, Asp(7.32(302)) of the human GnRH receptor determines selectivity for mammalian GnRH by its ability to induce a high affinity conformation of its native ligand. However, this initial interaction seems not to contribute to the final ligand-receptor complex. We propose that Arg(8) interacts transiently with Asp(7.32(302)) to induce a high-affinity ligand conformation of GnRH, which then interacts with a binding pocket that is common for both constrained and unconstrained analogs of GnRH.
Mol Pharmacol 2001 Dec
PMID:Role of aspartate7.32(302) of the human gonadotropin-releasing hormone receptor in stabilizing a high-affinity ligand conformation. 1172 35

The presence of progesterone response element (PRE) in the 5'-flanking region of the human GnRH receptor (GnRHR) suggests the possible regulation of this gene by progesterone (P). In the present study, we examined the effects of P in transcriptional regulation of human GnRHR gene expression at the pituitary and placenta levels since the GnRHR has been detected in both tissues. By the use of transient transfection assays, a differential regulation of human GnRHR promoter activity by P was observed. P treatment resulted in a decrease in promoter activity in the pituitary alphaT3-1 cells, suggesting a P-mediated inhibitory action. Interestingly, P is found to have a stimulatory role at the placental expression of this gene. Addition of RU486 to, or inhibition of endogenous P production by, the placental JEG-3 cells leads to a decrease in promoter activity, which is reversed by the replacement of P. Further studies have identified a putative PRE, namely human GR-PRE (located between -535 and -521, related to translation start site), that may be responsible for the P action since the mutation of these motifs reversed the P-mediated effects. The binding of PR to this element is confirmed by antibody supershift assays. The physiological effects of P are mediated through two PR isoforms, namely PR-A and PR-B. In the present study, overexpression of human PR-A resulted in a decrease in human promoter activity in both pituitary and placental cells. Interestingly, overexpression of PR-B exhibits a cell-dependent transcriptional activity, whereby it functions as a transcription activator in the placenta but as a transcription repressor in the pituitary. In summary, our results demonstrated a differential usage of PR-A and PR-B in transcriptional regulation of human GnRHR gene expression by P at the pituitary and placenta levels.
Mol Endocrinol 2001 Dec
PMID:Differential role of PR-A and -B isoforms in transcription regulation of human GnRH receptor gene. 1173 10


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