Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P05412 (c-Jun)
 11,453 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The hypothalamic decapeptide gonadotropin-releasing hormone stimulates mobilization of two discrete pools of calcium in clonal (alphaT3-1) and primary pituitary gonadotropes. A multidisciplinary approach was implemented to investigate the effects of discrete calcium fluctuations on the signaling pathways linking the gonadotropin-releasing hormone receptor to activation of mitogen-activated protein kinases and immediate early genes. Blockade of calcium influx through nifedipine-sensitive voltage-gated calcium channels reduced buserelin-induced activation of extracellular signal-regulated kinase (ERK) and c-Fos while activation of c-Jun N-terminal kinase and c-Jun was unaffected. Inhibition of buserelin-stimulated ERK activity by nifedipine was also observed in rat pituitary cells in primary culture. Direct activation of alphaT3-1 cell L-type calcium channels with the agonist Bay-K 8644 resulted in phosphorylation of ERK and induction of c-Fos. However, simple voltage-induced channel activation did not produce a sufficient calcium signal, since depolarization with 35 mM KCl failed to induce activation of ERK. Depletion of intracellular calcium stores with thapsigargin did not affect buserelin-induced ERK activation. An inhibitor of protein kinase C decreased calcium influx through nifedipine-sensitive calcium channels and phosphorylation of ERK induced by buserelin. Pharmacological inhibition of protein kinase C did not block Bay-K 8644-induced ERK activation. These observations suggest that calcium influx through L-type channels is required for GnRH-induced activation of ERK and c-Fos and that the influence of calcium lies downstream of protein kinase C.
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PMID:Calcium influx through L-type channels is required for selective activation of extracellular signal-regulated kinase by gonadotropin-releasing hormone. 1051 57

Receptors coupled to heterotrimeric G proteins are linked to activation of mitogen-activated protein kinases (MAPKs) via receptor- and cell-specific mechanisms. We have demonstrated recently that gonadotropin-releasing hormone (GnRH) receptor occupancy results in activation of extracellular signal-regulated kinase (ERK) through a mechanism requiring calcium influx through L-type calcium channels in alphaT3-1 cells and primary rat gonadotropes. Further studies were undertaken to explore the signaling mechanisms by which the GnRH receptor is coupled to activation of another member of the MAPK family, c-Jun N-terminal kinase (JNK). GnRH induces activation of the JNK cascade in a dose-, time-, and receptor-dependent manner in clonal alphaT3-1 cells and primary rat pituitary gonadotrophs. Coexpression of dominant negative Cdc42 and kinase-defective p21-activated kinase 1 and MAPK kinase 7 with JNK and ERK indicated that specific activation of JNK by GnRH appears to involve these signaling molecules. Unlike ERK activation, GnRH-stimulated JNK activity does not require activation of protein kinase C and is not blocked after chelation of extracellular calcium with EGTA. GnRH-induced JNK activity was reduced after treatment with the intracellular calcium chelator BAPTA-AM (1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester), whereas activation of ERK was not affected. Chelation of intracellular calcium also reduced GnRH-induced activation of JNK in rat pituitary cells in primary culture. GnRH-induced induction and activation of the JNK target c-Jun was inhibited after chelation of intracellular calcium, whereas induction of c-Fos, a known target of ERK, was unaffected. Therefore, although activation of ERK by GnRH requires a specific influx of calcium through L-type calcium channels, JNK activation is independent of extracellular calcium but sensitive to chelation of intracellular calcium. Our results provide novel evidence that GnRH activates two MAPK superfamily members via strikingly divergent signaling pathways with differential sensitivity to activation of protein kinase C and mobilization of discrete pools of calcium.
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PMID:Divergent signaling pathways requiring discrete calcium signals mediate concurrent activation of two mitogen-activated protein kinases by gonadotropin-releasing hormone. 1079 94

Clinical applications of GnRH agonists (GnRHa) are based primarily on the decrease in gonadotropin release after down-regulation of the GnRH receptor (GnRHR) by continuous GnRHa administration. However, the molecular mechanisms underlying the transcriptional regulation of the human GnRHR gene after prolonged GnRH treatment remain poorly understood. In the present study GnRHa-mediated regulation of human GnRHR gene transcription was studied by transiently transfecting the mouse gonadotrope-derived (alphaT3-1) cells with a 2297-bp human GnRHR promoter-luciferase construct (p2300-LucF). A dose- and time-dependent decrease in human GnRHR promoter activity was observed after GnRHa treatment. An average 71% decrease in promoter activity was observed after 24-h treatment with 0.1 microM GnRHa, which was blocked by cotreatment of the GnRH antagonist, antide. This effect was mimicked by phorbol 12-myristate 13-acetate (TPA) administration. In addition, the GnRHa- and TPA-mediated decrease in the human GnRHR promoter activity was reversed by a specific protein kinase C (PKC) inhibitor, GF109203X, or depletion of PKC by TPA pretreatment. These findings indicate that the activation of the PKC pathway is important in regulating the human GnRHR gene expression. By progressive 5'-deletion studies, we have identified a 248-bp DNA fragment (-1018 to -771, relative to the translation start site) at the 5'-flanking region of the human GnRHR gene that is responsible for the GnRHa-mediated down-regulation of human GnRHR promoter activity. Analysis of this sequence reveals the existence of two putative activating protein-1 (AP-1) sites with 87% homology to the consensus sequence (5'-TGA(G/C)T(C/A)A-3'), located at -1000 to -994 (5'-TTAGACA-3', in complementary orientation) and -943 to 937 (5'-TGAATAA-3'). Using competitive gel mobility shift assays, AP-1 binding was observed within this 248-bp region. Site-directed mutation of the putative AP-1-binding site located at -1000 to -994 abolished the GnRHa-induced inhibition. Further competitive GMSA and supershift experiments confirmed the identity of AP-1 binding in this region. By the use of Western blot analysis, a significant increase in c-Jun (100%; P < 0.05) and c-Fos (50%; P < 0.05) protein levels was observed after GnRHa treatment in alphaT3-1 cells. In addition, our data suggested that a change in AP-1 composition, particularly c-Fos, was important in mediating GnRHa-induced inhibition of human GnRHR gene expression. We conclude that activation of the PKC pathway by GnRH is important in controlling human GnRHR gene expression. In addition, the putative AP-1-binding site located at -1000 to -994 of the human GnRHR5'-flanking region has been functionally identified to be involved in mediating this down-regulatory effect.
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PMID:Transcriptional down-regulation of human gonadotropin-releasing hormone (GnRH) receptor gene by GnRH: role of protein kinase C and activating protein 1. 1101 15

GnRH has been showed to regulate hCG expression and secretion from the placenta through a GnRH receptor (GnRHR)-mediated process. Recently, we have reported the isolation of human GnRHR full-length complementary DNA from the human placental cells including choriocarcinoma JEG-3 cells, immortalized extravillous trophoblasts, and primary cultures of trophoblasts. Despite these observations, the molecular mechanism that controls the transcription regulation of the GnRHR gene expression in the placenta remains unknown. Here we described the identification of an upstream placenta-specific promoter located between nucleotide (nt) -1737 and -1346 (relative to the translation start site) for the human GnRHR gene. Using transient transfection studies, this upstream promoter has been shown to determine the placental cell-specific expression of this gene. Primer extension studies further confirmed the utilization of this promoter in JEG-3 cells in vivo. By mutagenesis coupled to functional studies, we have identified four putative transcription factor-binding sites, namely human glucocorticoid receptor (hGR)-Oct-1 (nt -1718 to -1710), hGR-cAMP response element (CRE; nt -1649 to -1641), hGR-GATA (nt -1602 to -1597), and hGR-activating protein-1 (nt -1518 to -1511), that are essential to the expression of this gene. Mutations of these cis-acting motifs reduced the promoter activity. The CRE and GATA motifs were subsequently shown to be placenta specific, as mutations of these motifs caused a dramatic loss in promoter activities in the placental JEG-3 cells, but not in the ovarian carcinoma OVCAR-3, monkey kidney COS-1, and human embryonic kidney 293 cells. Gel mobility assays confirmed the binding of nuclear proteins Oct-1, CRE-binding protein, GATA-2, GATA-3, c-Fos, and c-Jun from JEG-3 cells to these four elements.
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PMID:Functional mapping of a placenta-specific upstream promoter for human gonadotropin-releasing hormone receptor gene. 1125 Sep 31

The initial waves of gene induction caused by GnRH in the LbetaT2 gonadotrope cell line have recently been identified using microarrays. We now investigate the relationship of the concentration of GnRH to the level of biosynthesis induced. Using an optimized custom cDNA microarray, we show that a large number of genes are induced in a concentration-dependent fashion. Detailed time course studies of the induction of six induced transcripts using quantitative real-time PCR suggest that the amplitude, but not the temporal pattern, depends on the concentration of GnRH. The early genes appear to show a delay in gene induction, followed by a linear phase of increase. The relationship of rate of synthesis and GnRH concentration was studied by mathematical modeling of the induction of two genes, gly96 and tis11. In both cases, only the rates of increase, but not the lag times, are influenced by the concentration of GnRH exposure. Western blot analyses for c-Jun and Egr1 show that the levels of nuclear protein for these transcription factors also depend on the concentration of GnRH. These studies indicate that, despite the complex signaling network connecting the receptor to the activated genes, the biosynthetic rate of RNA polymerase at induced genes is correlated with the concentration of GnRH at the GnRH receptor.
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PMID:Coupling of GnRH concentration and the GnRH receptor-activated gene program. 1204 3

Although it is recognized that estrogen is one of the most important regulators of GnRH receptor (GnRHR) gene expression, the mechanism underlying the regulation at the transcriptional level is unknown. In the present study, we demonstrated that 17beta-estradiol (E2) repressed human GnRHR promoter via an activator protein 1-like motif and estrogen receptor-alpha, of which the DNA-binding domain and the ligand-binding domain were indispensable for the repression. Interestingly, the same cis-acting motif was also found to be important for both the basal activity and phorbol 12-myristate 13-acetate responsiveness of the GnRHR promoter. EMSAs indicated that multiple transcription factors including c-Jun and c-Fos bound to the activator protein 1-like site and that their DNA binding activity was not significantly affected by E2 treatment. In addition, we demonstrated that the E2 repression could be antagonized by phorbol 12-myristate 13-acetate, which stimulated c-Jun phosphorylation on serine 63, a process that is a prerequisite for recruitment of the transcriptional coactivator cAMP response element binding protein (CREB)-binding protein (CBP). Concomitantly, we found that overexpression of CBP could reverse the suppression in a dose-dependent manner. Taken together, our data indicate that E2-activated estrogen receptor-alpha represses human GnRHR gene transcription via an indirect mechanism involving CBP and possibly other transcriptional regulators.
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PMID:An activator protein 1-like motif mediates 17beta-estradiol repression of gonadotropin-releasing hormone receptor promoter via an estrogen receptor alpha-dependent mechanism in ovarian and breast cancer cells. 1294 46

Rapid engagement of the extracellular signal-regulated kinase (ERK) cascade via the Gq/11-coupled GnRH receptor (GnRHR) is mediated by transactivation of the epidermal growth factor receptor (EGFR). Here we show that the cross-talk between GnRHR and EGFR in gonadotropic cells is accomplished via gelatinases A and B (matrix metalloproteinases (MMPs) 2 and 9), identifying gelatinases as the first distinct members of the MMP family mediating EGFR transactivation by G protein-coupled receptors. Using a specific MMP2 and MMP9 inhibitor, Ro28-2653, GnRH-dependent EGFR transactivation was abrogated. Proving the specificity of the effect, transient transfection of alphaT3-1 cells with ribozymes directed against MMP2 or MMP9 specifically blocked EGFR tyrosine phosphorylation in response to GnRH stimulation. GnRH challenge of alphaT3-1 cells furthered the release of active MMP2 and MMP9 and increased their gelatinolytic activities within 5 min. Rapid release of activated MMP2 or MMP9 was inhibited by ribozyme-targeted down-regulation of MT1-MMP or MMP2, respectively. We found that GnRH-induced Src, Ras, and ERK activation were also gelatinase-dependent. Thus, gelatinase-induced EGFR transactivation was required to engage the extracellular-signal regulated kinase cascade. Activation of c-Jun N-terminal kinase and p38 MAPK by GnRH was unaffected by EGFR or gelatinase inhibition that, however, suppressed GnRH induction of c-Jun and c-Fos. Our findings suggest a novel role for gelatinases in the endocrine regulation of pituitary gonadotropes.
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PMID:Matrix metalloproteinases 2 and 9 mediate epidermal growth factor receptor transactivation by gonadotropin-releasing hormone. 1296 32

The gonadotropin-releasing hormone receptor (GnRHR) is expressed primarily in the gonadotropes of the anterior pituitary. Pituitary homeobox 1 (Pitx-1) has been shown to activate pituitary-specific gene expression by direct DNA binding and/or protein-protein interaction with other transcription factors. We hypothesized that Pitx-1 might also dictate tissue-specific expression of the mouse GnRHR (mGnRHR) gene in a similar manner. Pitx-1 activated the mGnRHR gene promoter, and transactivation was localized to sequences between -308 and -264. Pitx-1 bound to this region only with low affinity. This region includes an activating protein 1 (AP-1) site, which was previously shown to be important for mGnRHR gene expression. Further characterization indicated that an intact AP-1 site was required for full Pitx-1 responsiveness. Furthermore, Pitx-1 and AP-1 were synergistic in the activation of the mGnRHR gene promoter. A Pitx-1 homeodomain (HD) point mutation, which eliminated DNA binding ability, caused only a partial reduction of transactivation, whereas deletion of the HD completely prevented transactivation. Pitx-1 interacted directly with c-Jun, and the HD was sufficient for this interaction. While the point mutation in the Pitx-1 HD did not affect interaction with c-Jun, deletion of the HD eliminated the interaction. Taken together, our studies indicate that Pitx-1 can direct transactivation of the mGnRHR gene, in part by DNA binding and in part by an action of Pitx-1 as a cofactor for AP-1, augmenting AP-1 activity through a novel protein-protein interaction between c-Jun and the HD of Pitx-1.
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PMID:Essential role of the homeodomain for pituitary homeobox 1 activation of mouse gonadotropin-releasing hormone receptor gene expression through interactions with c-Jun and DNA. 1522 17

Gonadotropin-releasing hormone (GnRH) receptor is present in 80% of ovarian cancer, and numerous studies have provided evidence for a role of GnRH in cell proliferation. In this study, the effect of GnRH on the invasion potential of ovarian cancer cells was investigated. In vitro migration and cell invasion assays with the ovarian cancer cell lines Caov-3 and OVCAR-3 revealed the biphasic nature of GnRH; low concentrations of GnRH agonist (GnRHa) increased the cell motility and invasiveness of these cells, but at increased concentrations, the stimulatory effect was insignificant. Reverse transcription-PCR, Western blot, and gelatin zymography showed that the expression of metastasis-related proteinases, matrix metalloproteinase (MMP)-2 and MMP-9, was up-regulated and activated by GnRHa. Moreover, we observed that GnRHa was able to transactivate the MMP-2 and MMP-9 promoters. The invasive/migratory phenotype activated by GnRHa can be blocked by specific inhibitors or neutralizing antibodies to MMP-2 and MMP-9. Knockdown of the GnRH receptor using small interfering RNA significantly inhibited the GnRH-induced MMP activation, invasion, and migration. In addition, we showed that the c-Jun NH(2)-terminal kinase, but not extracellular signal-regulated kinase 1/2 or p38 mitogen-activated protein kinase, signaling pathway was critical for GnRH-mediated up-regulation of MMP, cell invasion, and motility. These results indicate for the first time an expanded role for GnRH in other aspects of ovarian tumor progression, such as metastasis, via activation of MMP and the subsequent increase in cell migration and invasion.
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PMID:Gonadotropin-releasing hormone promotes ovarian cancer cell invasiveness through c-Jun NH2-terminal kinase-mediated activation of matrix metalloproteinase (MMP)-2 and MMP-9. 1710 27

The GnRH receptor is a G protein-coupled receptor (GPCR), and its ligand GnRH is the central regulator of the reproductive system. GnRH receptors are known to target a wide variety of signal transduction pathways. Several recent studies have shown that activation of GPCRs can impact on beta-catenin signaling. beta-Catenin is the main effecter of the Wnt signaling pathway where it acts with the transcription factors T cell factor/lymphoid enhancer factor to mediate the transcription of Wnt target genes. We show that GnRH treatment promotes the nuclear accumulation of beta-catenin, activation of T cell factor-dependent transcription, and up-regulation of Wnt target genes, c-Jun, Fra-1, and c-Myc. These results are observed in human embryonic kidney 293/GnRH receptor-expressing cells and have been recapitulated in LbetaT2 and alphaT3-1 mouse gonadotrope cells. In addition to these findings, we show that GnRH treatment mediates the inactivation of glycogen synthase kinase-3, a protein serine/threonine kinase that regulates beta-catenin degradation within the Wnt signaling pathway. Our findings extend the number of GPCRs that can target beta-catenin signaling through diverse pathways. Furthermore, this is the first demonstration of the targeting of Wnt/beta-catenin signaling by a peptide hormone GPCR.
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PMID:Nuclear stabilization of beta-catenin and inactivation of glycogen synthase kinase-3beta by gonadotropin-releasing hormone: targeting Wnt signaling in the pituitary gonadotrope. 1771 75


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