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Pamela L Mellon - One of the best experts on this subject based on the ideXlab platform.

  • homeodomain proteins six3 and six6 regulate Gonadotrope specific genes during pituitary development
    Molecular Endocrinology, 2015
    Co-Authors: Hanne M Hoffmann, Jason D Meadows, Susan L Mayo, Crystal Trang, Sunamita S Leming, Chiara Maruggi, Shannon W Davis, Rachel Larder, Pamela L Mellon
    Abstract:

    Sine oculis-related homeobox 3 (SIX3) and SIX6, 2 closely related homeodomain transcription factors, are involved in development of the mammalian neuroendocrine system and mutations of Six6 adversely affect fertility in mice. We show that both small interfering RNA knockdown in Gonadotrope Cell Lines and knockout of Six6 in both embryonic and adult male mice (Six6 knockout) support roles for SIX3 and SIX6 in transcriptional regulation in Gonadotrope gene expression and that SIX3 and SIX6 can functionally compensate for each other. Six3 and Six6 expression patterns in Gonadotrope Cell Lines reflect the timing of the expression of pituitary markers they regulate. Six3 is expressed in an immature Gonadotrope Cell Line and represses transcription of the early Lineage-specific pituitary genes, GnRH receptor (GnRHR) and the common α-subunit (Cga), whereas Six6 is expressed in a mature Gonadotrope Cell Line and represses the specific β-subunits of LH and FSH (LHb and FSHb) that are expressed later in development. We show that SIX6 repression requires interaction with transducin-like enhancer of split corepressor proteins and competition for DNA-binding sites with the transcriptional activator pituitary homeobox 1. Our studies also suggest that estradiol and circadian rhythm regulate pituitary expression of Six6 and Six3 in adult females but not in males. In summary, SIX3 and SIX6 play distinct but compensatory roles in regulating transcription of Gonadotrope-specific genes as Gonadotrope Cells differentiate.

  • activin and glucocorticoids synergistically activate follicle stimulating hormone β subunit gene expression in the immortalized lβt2 Gonadotrope Cell Line
    Endocrinology, 2007
    Co-Authors: Shauna M Mcgillivray, Djurdjica Coss, Varykina G Thackray, Pamela L Mellon
    Abstract:

    FSH is produced by the pituitary Gonadotrope to regulate gametogenesis. Production of the β-subunit of FSH is the rate-limiting step in FSH synthesis, and a number of peptide and steroid hormones within the reproductive axis have been found to regulate transcription of the FSH β-subunit gene. Although both activin and glucocorticoids are notable regulators of FSHβ by themselves, we find that cotreatment results in a synergistic interaction on the mouse FSHβ promoter at the level of the Gonadotrope using transient transfection of a reporter gene into the LβT2 immortalized Gonadotrope-derived Cell Line. This synergistic interaction is specific to FSHβ, because only additive effects of these two hormones are observed on LH β-subunit, GnRH receptor, and mouse mammary tumor virus gene expression. Components of both activin and glucocorticoid signaling are found to be necessary for synergy, and there are specific cis elements on the mouse FSHβ promoter that contribute to the synergistic response as well. We als...

  • activin regulation of the follicle stimulating hormone β subunit gene involves smads and the tale homeodomain proteins pbx1 and prep1
    Molecular Endocrinology, 2004
    Co-Authors: Janice S Bailey, Djurdjica Coss, Naama Raveharel, Shauna M Mcgillivray, Pamela L Mellon
    Abstract:

    FSH is critical for normal reproductive function in both males and females. Activin, a member of the TGFβ family of growth factors, is an important regulator of FSH expression, but little is known about the molecular mechanisms through which it acts. We used transient transfections into the immortalized Gonadotrope Cell Line LβT2 to identify three regions (at −973/−962, −167, and −134) of the ovine FSH β-subunit gene that are required for full activin response. All three regions contain homology to consensus binding sites for Smad proteins, the intraCellular mediators of TGFβ family signaling. Mutation of the distal site reduces activin responsiveness, whereas mutation of either proximal site profoundly disrupts activin regulation of the FSHβ gene. These sites specifically bind LβT2 nuclear proteins in EMSAs, and the −973/−962 site binds Smad4 protein. Interestingly, the protein complex binding to the −134 site contains Smad4 in association with the homeodomain proteins Pbx1 and Prep1. Using glutathione S...

  • gnrh activates erk1 2 leading to the induction of c fos and lhβ protein expression in lβt2 Cells
    Molecular Endocrinology, 2002
    Co-Authors: Fujun Liu, Pamela L Mellon, Darrell A Austin, Jerrold M Olefsky, Nicholas J G Webster
    Abstract:

    GnRH acts on pituitary Gonadotropes to stimulate the synthesis and release of LH and FSH. However, the signaling pathways downstream of the GnRH receptor that mediate these effects are not fully understood. In this paper, we demonstrate that GnRH activates ERK, c-Jun N-terminal kinase, and p38MAPK in the LbetaT2 Gonadotrope Cell Line. Phosphorylation of both ERK and p38MAPK are stimulated rapidly, 30- to 50-fold in 5 min, but activation of c-Jun N-terminal kinase has slower kinetics, reaching only 10-fold after 30 min. Activation of ERK by GnRH is blocked by inhibition of MAPK kinase (MEK) and partially blocked by inhibition of PKC and calcium, but not PI3K or p38MAPK signaling. We demonstrate that phosphorylated ERK accumulates in the nucleus in a PKC-dependent manner. We also show that GnRH induces c-fos and LHbeta subunit protein expression in LbetaT2 Cells via MEK. Experiments with EGTA or calcium channel antagonists indicated that calcium influx is important for the induction of both genes by GnRH. In conclusion, these results show that GnRH activates all three MAPK subfamilies in LbetaT2 Cells and induces c-fos and LHbeta protein expression through calcium and MEK-dependent mechanisms. These results also demonstrate that the nuclear translocation of ERK by GnRH requires PKC signaling.

  • steroid and pulsatile gonadotropin releasing hormone gnrh regulation of luteinizing hormone and gnrh receptor in a novel Gonadotrope Cell Line
    Molecular Endocrinology, 1996
    Co-Authors: Judith L Turgeon, Yuka Kimura, D W Waring, Pamela L Mellon
    Abstract:

    Properties of a pituitary Gonadotrope include the capacity to regulate gonadotropin synthesis and secretion in response to a GnRH signal. Progress in identifying the steps involved in these processes has been impeded by the lack of a homogeneous in vitro model of Gonadotropes. This paper presents functional characterization of a L beta T2 Gonadotrope Cell Line generated by tumorigenesis in transgenic mice carrying the rat LH beta-subunit regulatory region linked to the SV40 T-antigen oncogene. This Cell Line expresses LH beta, alpha-subunit, and GnRH-receptor (GnRH-R) mRNAs (though not FSH beta), responds to glucocorticoid treatment with a reversible dampening of proliferation, and responds to pulsatile, concentration-dependent GnRH administration with LH secretion. L beta T2 Cells presented with four GnRH pulses (10 nM, 90-min interpulse interval) on each of 4 days respond with incremental increases in LH secretion on successive days. This increase was greatest (15-fold) in the presence of estradiol and dexamethasone. Part of the enhanced responsiveness is apparently due to an increase in GnRH-R; pulsatile GnRH treatment alone as well as steroid treatment alone led to an increase in GnRH-R mRNA levels. When secretion was stimulated on day 4 with 54 mM [K+] pulses, bypassing the GnRH-R, the LH-secretory response indicated that the GnRH pulse history as well as estradiol and dexamethasone have actions on L beta T2-secretory capacity distinct from changes in the GnRH-R. This increase can be explained in part by the marked up-regulation of LH beta, but not alpha-subunit, mRNA observed in GnRH-pulsed Cells. In summary, L beta T2 clonal Gonadotropes exhibit functional characteristics consistent with those of normal pituitary Gonadotropes such as LH secretion via a regulated pathway and changes in GnRH-R and LH beta gene expression in response to signaling by GnRH and steroid hormones and therefore should be a useful tool for dissecting the Cellular and molecular events involved in these fundamental Gonadotrope properties.

Robert P Millar - One of the best experts on this subject based on the ideXlab platform.

  • Gonadotropin-Inhibitory Hormone Inhibits GnRH- Induced Gonadotropin Subunit Gene Transcriptions by Inhibiting AC/cAMP/PKA-Dependent ERK Pathway
    2016
    Co-Authors: In Lt Cells, Robert P Millar, You Lee Son, Takayoshi Ubuka, Haruhiko Kanasaki, Kazuyoshi Tsutsui
    Abstract:

    A neuropeptide that directly inhibits gonadotropin secretion from the pituitary was discovered in quail and named gonadotropin-inhibitory hormone (GnIH). The presence and functional roles of GnIHorthologs, RF-amide-relatedpeptides (RFRP), thatpossess a commonC-terminal LPXRF-amide (X L orQ)motif have also been demonstrated inmammals. GnIH orthologs inhibit gonadotropin synthesis and release by acting on pituitary Gonadotropes andGnRHneurons in the hypothalamus via its receptor (GnIH receptor). It is becoming increasingly clear that GnIH is an important hypo-thalamic neuropeptide controlling reproduction, but the detailed signaling pathway mediating the inhibitory effect of GnIH on target Cells is still unknown. In the present study, we investigated the pathway of GnIH Cell signaling and its possible interaction with GnRH signaling using amouse Gonadotrope Cell Line, LT2. First, we demonstrated the expression ofGnIH receptormRNA in LT2 Cells by RT-PCR. We then examined the inhibitory effects of mouse GnIH orthologs [mouse RFRP (mRFRP)] on GnRH-induced Cell signaling events. We showed that mRFRP effectively inhibited GnRH-induced cAMP signaling by using a cAMP-sensitive reporter system and measuring cAMP levels, indicating thatmRFRP function as an inhibitor of adenylate cyclase.We further showed that mRFRP inhibited GnRH-stimulated ERK phosphorylation, and this effect was mediated by th

  • gonadotropin inhibitory hormone gnih gnih receptor and Cell signaling
    General and Comparative Endocrinology, 2013
    Co-Authors: Takayoshi Ubuka, Robert P Millar, You Lee Son, George E Bentley, Kazuyoshi Tsutsui
    Abstract:

    Gonadotropin-inhibitory hormone (GnIH) is an inhibitor of gonadotropin synthesis and release, which was originally identified in the hypothalamus of the Japanese quail (Coturnix japonica). The GnIH precursor polypeptide encodes one GnIH and two GnIH related peptides (GnIH-RP-1 and GnIH-RP-2) in birds that share the same C-terminal LPXRFamide (X=L or Q) motif. The receptor for GnIH is thought to be the G protein-coupled receptor 147 (GPR147) which has been shown to couple predominantly through the Gαi protein to inhibit cAMP production. The crude membrane fraction of COS-7 Cells transfected with GPR147 cDNA specifically bound GnIH and GnIH-RPs in a concentration-dependent manner. Scatchard plot analysis of the binding showed that GPR147 possessed a single class of high-affinity binding sites. GnIH neurons project to the median eminence to control anterior pituitary function and GPR147 is expressed in the Gonadotropes. GnIH neurons also project to gonadotropin-releasing hormone (GnRH)-I and GnRH-II neurons, and GnRH-I and GnRH-II neurons express GPR147. Thus, GnIH may inhibit gonadotropin synthesis and release by decreasing the activity of GnRH-I neurons as well as directly inhibiting the effects of GnRH on Gonadotropes. GnIH may also partially inhibit reproductive behaviors by inhibiting GnRH-II neurons. GnIH and GPR147 are also expressed in the gonads, possibly acting in an autocrine/paracrine manner. The Cell signaling process of GPR147 was extensively studied using LβT2 Cells, a mouse Gonadotrope Cell Line. In this Cell Line, mouse GnIH inhibits GnRH-induced gonadotropin subunit, LHβ, FSHβ, and common α, gene transcriptions by inhibiting adenylate cyclase/cAMP/PKA dependent ERK pathway. This review summarizes the functions of GnIH, GnIH receptor and its Cell signaling processes in birds and discusses related findings in mammals.

  • reciprocal cross talk between gonadotropin releasing hormone gnrh and prostaglandin receptors regulates gnrh receptor expression and differential gonadotropin secretion
    Molecular Endocrinology, 2007
    Co-Authors: Zvi Naor, Michal Naidich, Adam J Pawson, Henry N Jabbour, Kevin Morgan, Sharon Battersby, M R Millar, Pamela Brown, Robert P Millar
    Abstract:

    The asynchronous secretion of Gonadotrope LH and FSH under the control of GnRH is crucial for ovarian cyclicity but the underlying mechanism is not fully resolved. Because prostaglandins (PG) are autocrine regulators in many tissues, we determined whether they have this role in Gonadotropes. We first demonstrated that GnRH stimulates PG synthesis by induction of cyclooxygenase-2, via the protein kinase C/c-Src/phosphatidylinositol 3′-kinase/MAPK pathway in the LβT2 Gonadotrope Cell Line. We then demonstrated that PGF2α and PGI2, but not PGE2 inhibited GnRH receptor expression by inhibition of phosphoinositide turnover. PGF2α, but not PGI2 or PGE2, reduced GnRH-induction of LHβ gene expression, but not the α-gonadotropin subunit or the FSHβ subunit genes. The prostanoid receptors EP1, EP2, FP, and IP were expressed in rat Gonadotropes. Incubations of rat pituitaries with PGF2α, but not PGI2 or PGE2, inhibited GnRH-induced LH secretion, whereas the cyclooxygenase inhibitor, indomethacin, stimulated GnRH-ind...

  • cloning and functional expression of a mouse gonadotropin releasing hormone receptor
    Molecular Endocrinology, 1992
    Co-Authors: Manami Tsutsumi, Pamela L Mellon, Wei Zhou, Robert P Millar, Colleen A Flanagan, James L Roberts, Kewen Dong, Boaz Gillo, Stuart C Sealfon
    Abstract:

    GnRH plays a pivotal role in the reproductive system, and GnRH analogs have wide therapeutic applications ranging from the treatment of prostatic cancer to infertility. Determination of the predicted structure of the GnRH receptor (GnRHR) would illuminate the mechanisms of receptor activation and regulation and allow directed design of improved GnRH analogs. We report the cloning of a cDNA representing the mouse GnRHR and confirm its identity using Xenopus oocyte expression. Injection of sense RNA transcript leads to the expression of a functional, high affinity GnRHR. Expression of the GnRHR using Gonadotrope Cell Line RNA, however, is blocked by an antisense oligonucleotide. In situ hybridization in the rat anterior pituitary reveals a characteristic GnRHR distribution. The nucleotide sequence encodes a 327-amino acid protein which has the seven putative transmembrane domains characteristic of G protein-coupled receptors, but which lacks a typical intraCellular C-terminus. The unusual structure and nove...

Mark A Lawson - One of the best experts on this subject based on the ideXlab platform.

  • gnrh regulates gonadotropin gene expression through nadph dual oxidase derived reactive oxygen species
    Endocrinology, 2015
    Co-Authors: Taeshin Kim, Mark A Lawson
    Abstract:

    The appropriate control of synthesis and secretion of the gonadotropin hormones LH and FSH by pituitary Gonadotropes is essential for the regulation of reproduction. The hypothalamic neuropeptide GnRH is the central regulator of both processes, coordinating secretion with transcription and translation of the gonadotropin hormone subunit genes. The MAPK family of second messengers is strongly induced in Gonadotropes upon GnRH stimulation, and multiple pathways activate these kinases. IntraCellular reactive oxygen species participate in signaling cascades that target MAPKs, but also participate in signaling events indicative of Cell stress. The NADPH oxidase (NOX)/dual oxidase (DUOX) family is a major enzymatic source of intraCellular reactive oxygen, and we show that GnRH stimulation of mouse primary pituitary Cells and the LβT2 Gonadotrope Cell Line elevates intraCellular reactive oxygen via NOX/DUOX activity. Mouse pituitary and LβT2 Cells abundantly express NOX/DUOX and cofactor mRNAs. Pharmacological i...

  • gnrh induces the unfolded protein response in the lβt2 pituitary Gonadotrope Cell Line
    Molecular Endocrinology, 2009
    Co-Authors: Sharon J Santos, Mark A Lawson
    Abstract:

    The neuropeptide GNRH 1 stimulates the secretion of the reproductive hormone LH in pituitary Gonadotropes. Other secretory Cell types depend on the unfolded protein response (UPR) pathway to regulate protein synthesis and protect against endoplasmic reticulum (ER) stress in response to differentiation or secretory stimuli. This study investigated the role of the UPR in GNRH action within the LbetaT2 Gonadotrope model. Cells were treated with GNRH, and the activation of UPR signaling components and general translational status was examined. The ER-resident stress sensors, Atf6, Eif2ak3, and Ern1, are all present, and GNRH stimulation results in the phosphorylation of eukaryotic translation initiation factor 2A kinase 3 and its downstream effector, eukaryotic translation initiation factor 2A. Additionally, activation of the UPR was confirmed both in LbetaT2 as well as mouse primary pituitary Cells through identifying GNRH-induced splicing of Xbp1 mRNA, a transcription factor activated by splicing by the ER stress sensor, ER to nucleus signaling 1. Ribosome profiling revealed that GNRH stimulation caused a transient attenuation in translation, a hallmark of the UPR, remodeling ribosomes from actively translating polysomes to translationally inefficient ribonucleoprotein complexes and monosomes. The transient attenuation of specific mRNAs was also observed. Overall, the results show that GNRH activates components of the UPR pathway, and this pathway may play an important physiological role in adapting the ER of Gonadotropes to the burden of their secretory demand.

  • androgen responsiveness of the pituitary Gonadotrope Cell Line lbetat2
    Journal of Endocrinology, 2001
    Co-Authors: Mark A Lawson, Christine A Glidewellkenney, F J Lopez
    Abstract:

    Androgens have a profound effect on the hypothalamic-pituitary axis by reducing the synthesis and release of the pituitary gonadotropin LH. The effect on LH is partly a consequence of a direct, steroid-dependent action on pituitary function. Although androgen action has been well studied in vivo, in vitro Cell models of androgen action on pituitary Gonadotropes have been scarce. Recently, an LH-expressing Cell Line, LbetaT2, was generated by tumorigenesis targeted to the LH-producing Cells of the mouse pituitary. The purpose of these studies was to determine the presence of androgen receptor (AR) and establish its function in this Cell Line. RT-PCR analysis indicated that the LbetaT2 Cell Line expresses AR mRNA. Transient transfection assays, using the mouse mammary tumor virus (MMTV) promoter, showed that a functional AR is also present. Testosterone (TEST), dihydrotestosterone (DHT), 7alpha-methyl-19-nortestosterone (MENT), and fluoxymesterone (FLUOXY) increased reporter gene activity in the rank order of potencies MENT>DHT> TEST>FLUOXY. Additionally, activation of MMTV promoter activity by DHT in LbetaT2 Cells was diminished by the AR antagonists casodex and 2-hydroxy-flutamide, indicating that the effects of DHT are mediated through AR. In summary, these studies showed that the LbetaT2 Cell Line is a useful model for the evaluation and molecular characterization of androgen action in pituitary Gonadotropes.

Yoel Sadovsky - One of the best experts on this subject based on the ideXlab platform.

  • activation of luteinizing hormone β gene by gonadotropin releasing hormone requires the synergy of early growth response 1 and steroidogenic factor 1
    Journal of Biological Chemistry, 1999
    Co-Authors: C Dorn, Qinglin Ou, John Svaren, Peter A Crawford, Yoel Sadovsky
    Abstract:

    Abstract We have previously shown that early growth response (Egr) 1-deficient mice exhibit female infertility, reflecting a luteinizing hormone (LH) β deficiency. Egr-1 activates the LHβ gene in vitro through synergy with steroidogenic factor-1 (SF-1), a protein required for Gonadotrope function. To test if this synergy is essential for gonadotropin-releasing hormone (GnRH) stimulation of LHβ, we examined the activity of the LHβ promoter in the Gonadotrope Cell Line LβT2. GnRH markedly stimulated the LHβ promoter (15-fold). Mutation of either Egr-1 or SF-1 elements within the LHβ promoter attenuated this stimulation, whereas mutation of both promoter elements abrogated GnRH induction of the LHβ promoter. Furthermore, GnRH stimulated Egr-1 but not SF-1 expression in LβT2 Cells. Importantly, overexpression of Egr-1 alone was sufficient to enhance LHβ expression. Although other Egr proteins are expressed in LβT2 Cells and are capable of interacting with SF-1, GnRH stimulation of Egr-1 was the most robust. We also found that the nuclear receptor DAX-1, a repressor of SF-1 activity, reduced Egr-1–SF-1 synergy and diminished GnRH stimulation of the LHβ promoter. We conclude that the synergy between Egr-1 and SF-1 is essential for GnRH stimulation of the LHβ gene and plays a central role in the dynamic regulation of LHβ expression.

  • activation of luteinizing hormone β gene by gonadotropin releasing hormone requires the synergy of early growth response 1 and steroidogenic factor 1
    Journal of Biological Chemistry, 1999
    Co-Authors: C Dorn, John Svaren, Peter A Crawford, Yoel Sadovsky
    Abstract:

    We have previously shown that early growth response (Egr) 1-deficient mice exhibit female infertility, reflecting a luteinizing hormone (LH) beta deficiency. Egr-1 activates the LHbeta gene in vitro through synergy with steroidogenic factor-1 (SF-1), a protein required for Gonadotrope function. To test if this synergy is essential for gonadotropin-releasing hormone (GnRH) stimulation of LHbeta, we examined the activity of the LHbeta promoter in the Gonadotrope Cell Line LbetaT2. GnRH markedly stimulated the LHbeta promoter (15-fold). Mutation of either Egr-1 or SF-1 elements within the LHbeta promoter attenuated this stimulation, whereas mutation of both promoter elements abrogated GnRH induction of the LHbeta promoter. Furthermore, GnRH stimulated Egr-1 but not SF-1 expression in LbetaT2 Cells. Importantly, overexpression of Egr-1 alone was sufficient to enhance LHbeta expression. Although other Egr proteins are expressed in LbetaT2 Cells and are capable of interacting with SF-1, GnRH stimulation of Egr-1 was the most robust. We also found that the nuclear receptor DAX-1, a repressor of SF-1 activity, reduced Egr-1-SF-1 synergy and diminished GnRH stimulation of the LHbeta promoter. We conclude that the synergy between Egr-1 and SF-1 is essential for GnRH stimulation of the LHbeta gene and plays a central role in the dynamic regulation of LHbeta expression.

C Dorn - One of the best experts on this subject based on the ideXlab platform.

  • activation of luteinizing hormone β gene by gonadotropin releasing hormone requires the synergy of early growth response 1 and steroidogenic factor 1
    Journal of Biological Chemistry, 1999
    Co-Authors: C Dorn, Qinglin Ou, John Svaren, Peter A Crawford, Yoel Sadovsky
    Abstract:

    Abstract We have previously shown that early growth response (Egr) 1-deficient mice exhibit female infertility, reflecting a luteinizing hormone (LH) β deficiency. Egr-1 activates the LHβ gene in vitro through synergy with steroidogenic factor-1 (SF-1), a protein required for Gonadotrope function. To test if this synergy is essential for gonadotropin-releasing hormone (GnRH) stimulation of LHβ, we examined the activity of the LHβ promoter in the Gonadotrope Cell Line LβT2. GnRH markedly stimulated the LHβ promoter (15-fold). Mutation of either Egr-1 or SF-1 elements within the LHβ promoter attenuated this stimulation, whereas mutation of both promoter elements abrogated GnRH induction of the LHβ promoter. Furthermore, GnRH stimulated Egr-1 but not SF-1 expression in LβT2 Cells. Importantly, overexpression of Egr-1 alone was sufficient to enhance LHβ expression. Although other Egr proteins are expressed in LβT2 Cells and are capable of interacting with SF-1, GnRH stimulation of Egr-1 was the most robust. We also found that the nuclear receptor DAX-1, a repressor of SF-1 activity, reduced Egr-1–SF-1 synergy and diminished GnRH stimulation of the LHβ promoter. We conclude that the synergy between Egr-1 and SF-1 is essential for GnRH stimulation of the LHβ gene and plays a central role in the dynamic regulation of LHβ expression.

  • activation of luteinizing hormone β gene by gonadotropin releasing hormone requires the synergy of early growth response 1 and steroidogenic factor 1
    Journal of Biological Chemistry, 1999
    Co-Authors: C Dorn, John Svaren, Peter A Crawford, Yoel Sadovsky
    Abstract:

    We have previously shown that early growth response (Egr) 1-deficient mice exhibit female infertility, reflecting a luteinizing hormone (LH) beta deficiency. Egr-1 activates the LHbeta gene in vitro through synergy with steroidogenic factor-1 (SF-1), a protein required for Gonadotrope function. To test if this synergy is essential for gonadotropin-releasing hormone (GnRH) stimulation of LHbeta, we examined the activity of the LHbeta promoter in the Gonadotrope Cell Line LbetaT2. GnRH markedly stimulated the LHbeta promoter (15-fold). Mutation of either Egr-1 or SF-1 elements within the LHbeta promoter attenuated this stimulation, whereas mutation of both promoter elements abrogated GnRH induction of the LHbeta promoter. Furthermore, GnRH stimulated Egr-1 but not SF-1 expression in LbetaT2 Cells. Importantly, overexpression of Egr-1 alone was sufficient to enhance LHbeta expression. Although other Egr proteins are expressed in LbetaT2 Cells and are capable of interacting with SF-1, GnRH stimulation of Egr-1 was the most robust. We also found that the nuclear receptor DAX-1, a repressor of SF-1 activity, reduced Egr-1-SF-1 synergy and diminished GnRH stimulation of the LHbeta promoter. We conclude that the synergy between Egr-1 and SF-1 is essential for GnRH stimulation of the LHbeta gene and plays a central role in the dynamic regulation of LHbeta expression.