Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.24 (mitogen-activated protein kinase)
 95,810 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This study investigated whether estradiol (E2) or testosterone (T) activate extracellular signal-regulated kinase (ERK) and calcium/calmodulin-dependent kinase II (Ca/CaMK II), as indicated by enzyme phosphorylation in rat pituitaries. In vivo studies used adult female rats given E2, T, or empty silastic capsules (vehicle controls). Twenty-four hours later, the rats were given a single pulse of GnRH (300 ng) or BSA-saline (to controls) and killed 5 min later. GnRH stimulated a two- to three-fold rise in activated Ca/CaMK II, and E2 and T had no effect on Ca/CaMK II activation. In contrast, both GnRH and T stimulated threefold increases in ERK activity, with additive effects seen following the combination of GnRH+T. E2 had no effect on ERK activity. In alpha T3 clonal gonadotrope cells, dihydrotestosterone did not activate ERK alone but enhanced and prolonged the ERK responses to GnRH, demonstrating direct effects on the gonadotrope. Thus, the ERK response to GnRH plus androgen was enhanced in both rat pituitary and alpha T3 cells. In vitro studies with cultured rat pituitary cells examined the effect of GnRH+/-T in the presence of the mitogen-activated protein (MAP) kinase kinase inhibitor, PD-098059 (PD). Results showed that PD suppressed ERK activational and FSH beta transcriptional responses to T. These findings suggest that one site of T regulation of FSH beta transcription is through the selective stimulation of the ERK pathway.
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PMID:Testosterone stimulates follicle-stimulating hormone beta transcription via activation of extracellular signal-regulated kinase: evidence in rat pituitary cells. 1550 29

Bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9) are oocyte-specific growth factors that appear to play key roles in granulosa cell development and fertility in most mammalian species. We have evaluated the role(s) of these paracrine factors in the development and function of both the cumulus cells and oocytes by assessing cumulus expansion, oocyte maturation, fertilization, and preimplantation embryogenesis in Gdf9+/-Bmp15-/- [hereafter, double mutant (DM)] mice. We found that cumulus expansion, as well as the expression of hyaluronon synthase 2 (Has2) mRNA was impaired in DM oocyte-cumulus cell complexes. This aberrant cumulus expansion was not remedied by coculture with normal wild-type (WT) oocytes, indicating that the development and/or differentiation of cumulus cells in the DM, up to the stage of the preovulatory luteinizing hormone (LH) surge, is impaired. In addition, DM oocytes failed to enable FSH to induce cumulus expansion in WT oocytectomized (OOX) cumulus. Moreover, LH-induced oocyte meiotic resumption was significantly delayed in vivo, and this delayed resumption of meiosis was correlated with the reduced activation of mitogen-activated protein kinase (MAPK) in the cumulus cells, thus suggesting that GDF9 and BMP15 also regulate the function of cumulus cells after the preovulatory LH surge. Although spontaneous in vitro oocyte maturation occurred normally, oocyte fertilization and preimplantation embryogenesis were significantly altered in the DM, suggesting that the full complement of both GDF9 and BMP15 are essential for the development and function of oocytes. Because receptors for GDF9 and BMP15 have not yet been identified in mouse oocytes, the effects of the mutations in the Bmp15 and Gdf9 genes on oocyte development and functions must be produced indirectly by first affecting the granulosa cells and then the oocyte. Therefore, this study provides further evidence for the existence and functioning of an oocyte-granulosa cell regulatory loop.
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PMID:Synergistic roles of BMP15 and GDF9 in the development and function of the oocyte-cumulus cell complex in mice: genetic evidence for an oocyte-granulosa cell regulatory loop. 1553 64

Both the viability of hen prehierarchal follicles and subsequent differentiation associated with the selection of a single follicle per day into the preovulatory hierarchy depend on circulating FSH and the expression of FSH receptor (FSH-R) in granulosa cells. The present study addresses mechanisms that mediate both basal expression plus selective up-regulation of FSH-R mRNA in granulosa cells from prehierarchal follicles. Results demonstrate that FSH-R mRNA is both expressed and functional in granulosa cells collected from growing prehierarchal follicles as small as those of 1-2 mm in diameter, as indicated by rapid induction of steroidogenic acute regulatory (StAR) protein expression by FSH in vitro. Real-time polymerase chain reaction determined that relative FSH-R expression within the granulosa layer from individual prehierarchal follicles of 6-8 mm in diameter was similar among the 8-13 follicles within this cohort, with the notable exception that the granulosa layer from a single follicle (presumably the selected follicle) showed elevated expression. Levels of FSH-R mRNA expression were enhanced by both recombinant human (rh) transforming growth factor (TGF) beta1 and, to a lesser extent, rh-activin A after 20 h of culture. This stimulatory effect was effectively blocked by mitogen-activated protein (MAP) kinase signaling induced by TGF alpha treatment. Finally, inhibition of MAP kinase signaling, using the selective inhibitor U0126, promoted FSH-R expression and further enhanced TGF beta1-induced FSH-R expression in vitro. Collectively, results suggest that premature granulosa cell differentiation normally is suppressed by tonic MAP kinase signaling. At the time of follicle selection, a release from inhibitory MAP kinase signaling is proposed to occur, which enables the full potentiation of FSH-R expression mediated by intrafollicular factors.
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PMID:Regulation of follicle-stimulating hormone-receptor messenger RNA in hen granulosa cells relative to follicle selection. 1553 65

Follicular development is characterized by both proliferation and differentiation of granulosa cells (GCs) under the control of FSH. However, the cellular mechanism by FSH is not known. Using cultured GCs, we examined whether FSH activated ERK1/2 was involved in the regulation of the proliferation related gene proliferating cell nuclear antigen (PCNA) and steroidogenesis. GCs were obtained from the ovaries of DES treated immature rats and cultured in serum free medium. The results showed that FSH activated ERK1/2 in a time dependent manner, with a peak at 20 min. Such activation was PKA dependent as was inhibited by specific inhibitors. FSH induced PCNA expression in a time dependent manner, with a maximum stimulation at 2 h. Similarly, StAR and steroid levels increased as FSH treatment time extended, with a maximum progesterone and StAR production at 48 h. ERK1/2 inactivation by UO126 inhibited the stimulatory effects of FSH on both PCNA and StAR expression and steroid synthesis in the GCs (p less than 0.01). Immunocytochemical studies further revealed that ERK1/2 inhibition led to a reduction of mitochondrial StAR in the GCs by FSH. These observations suggested that the stimulation of FSH on PCNA expression and steroidogenesis in GCs was mediated at least partially by ERK1/2.
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PMID:Role of ERK1/2 in FSH induced PCNA expression and steroidogenesis in granulosa cells. 1556 28

The current study was done to elucidate the mechanism of the FSH stimulation of IGF-binding protein 3 (IGFBP-3) expression and map the FSH response element on the pig IGFBP-3 promoter. Forskolin induced IGFBP-3 reporter activity in transiently transfected granulosa cells. The protein kinase A (PKA) inhibitor [N-[2-(p-bromocinnamyl)amino)ethyl]-5-isoquinolinesulfonamide, 2HCl] (and cotransfection with a PKA inhibitor expression vector), the phosphatidylinositol-3 kinase inhibitor [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one], and the ERK inhibitor [1,4-diamino-2,3-dicyano-1,4-bis(2-aminophenylthio)butadiene], all blocked FSH stimulation. Use of serial deletion constructs and site-directed mutagenesis show that a TATA box-binding protein site is required for FSH stimulation and that a specific protein 1 (Sp1) site is required for basal transcription. Gel shift assays of nuclear protein with a -61/-25 probe detected four protein-DNA complexes, with bands I and II having significantly higher intensities in FSH-treated cells than in controls. Mutation of the Sp1 site prevented formation of bands I and II whereas mutation of the TATA box-binding protein site prevented formation of band IV. Use of specific antibodies showed that Sp1 participates in formation of band I, Sp3 band II, and p300 in both I and II. Band III was nonspecifically competed out. We conclude that FSH stimulation of IGFBP-3 transcription is mediated by cAMP via the PKA pathway and requires the P1-3 kinase and likely the MAPK pathways.
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PMID:Follicle-stimulating hormone induction of ovarian insulin-like growth factor-binding protein-3 transcription requires a TATA box-binding protein and the protein kinase A and phosphatidylinositol-3 kinase pathways. 1571 91

Atrial natriuretic peptide (ANP) as well as its receptors is found in mammalian ovary and follicular cells and its function in oocyte meiotic maturation has also been reported in Xenopus, hamster and rat. But the results are controversial and the physiological mechanism of ANP on oocyte maturation is not clear, especially the relationship between gonadotrophin and ANP as well as the signal transduction, and these need further study. The present study conducted experiments to examine these questions by using drug treatment and Western blot analysis and focused on pig oocyte meiotic maturation and cumulus expansion in vitro. The results revealed that ANP could inhibited FSH-induced pig oocyte maturation and cumulus expansion and prevent the full phosphorylation of mitogen-activated protein kinase in both oocytes and cumulus cells, and that these inhibitory effects could be mimicked by 8-Br-cyclic guanosine 5'-monophosphate (8-Br-cGMP), but blocked by a protein kinase G (PKG) inhibitor KT5823. Zaprinast, a cGMP-specific phosphodiesterase inhibitor, could enhance the inhibitory effect of ANP on oocyte maturation. A specific analogue of ANP, C-ANP-(4-23), which binds to the natriuretic peptide receptor-C (NPRC), had no effect in either FSH-induced or spontaneous oocyte maturation. Treatment with forskolin, a stimulator of adenylate cyclase, had a biphasic effect; 44 h treatment induced cumulus expansion but inhibited oocyte maturation while 2 h treatment induced maturation of cumulus-enclosed oocytes (CEOs). Both ANP and C-ANP-(4-23) could inhibit the effect of forskolin on CEO maturation, and these inhibitory effects of ANP/C-ANP-(4-23) could be blocked by preincubation with pertussis toxin (PT), consistent with mediation by a Gi protein(s) in the cumulus cells. All these results suggest that ANP is a multifunctional regulator of FSH and forskolin on pig CEO maturation by two signalling mechanisms: one is via a cGMP/PKG pathway, the other is via NPRC receptors in cumulus cells and the activation of the PT-sensitive Gi protein(s).
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PMID:Atrial natriuretic peptide inhibits the actions of FSH and forskolin in meiotic maturation of pig oocytes via different signalling pathways. 1582 Nov 10

MAPK plays an important role during meiotic maturation in mammalian oocytes, whereas the necessity of MAPK during meiotic resumption in porcine oocytes is still controversial. Here, by applying the method of ultracentrifugation to move the opaque lipid droplets to the edge of the oocyte, therefore allowing clear visualization of porcine germinal vesicles, oocytes just before germinal vesicle breakdown (GVBD) and those that had just undergone GVBD were selected for the assay of MAPK activation. Our results showed that phosphorylation of MAPK in oocytes occurred after GVBD in all three different culture models: spontaneous maturation model, inhibition-induction maturation model, and normal maturation model. Moreover, we found that activation of MAPK in cumulus cells but not in oocytes was essential for GVBD in cumulus-enclosed oocytes. Then the cross-talk between cAMP and MAPK in cumulus cells was investigated by using cell-type-specific phosphodiesterase (PDE) isoenzyme inhibitors. Our results showed that PDE3 subtype existed in oocytes, whereas PDE4 subtype existed in cumulus cells. PDE3 inhibitor prevented meiotic resumption of oocytes, whereas PDE4 inhibitor enhanced the ability of FSH or forskolin to activate MAPK in cumulus cells. We propose that increased cAMP resulting from inhibition of PDE3 in oocytes blocks GVBD, whereas increased cAMP resulting from inhibition of PDE4 activates MAPK pathway in cumulus cells, which is essential for GVBD induction.
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PMID:Cyclic adenosine 3',5'-monophosphate-dependent activation of mitogen-activated protein kinase in cumulus cells is essential for germinal vesicle breakdown of porcine cumulus-enclosed oocytes. 1600 24

The AMP-activated protein kinase (AMPK) is a major regulator of energy metabolism involved in fatty acid and cholesterol synthesis. In the ovary, cholesterol plays a key role in steroid production. We report the presence of AMPK in rat ovaries, and we have investigated its role in granulosa cells. We show using RT-PCR and Western blot that the mRNAs for the alpha1/2 and beta1/2 subunits and the proteins are found in the ovaries. Immunohistochemistry localized the alpha1 AMPK subunit in granulosa cells, corpus luteum, and oocyte and less abundantly in theca cells. Treatment with 1 mm 5-amino-imidazole-4-carboxyamide-1-beta-D-ribofuranoside (AICAR), an activator of AMPK, increased dose-dependent and time-dependent phosphorylation of AMPKalpha1 on Thr172 in primary granulosa cells. Simultaneously, phosphorylation of acetyl-coenzyme A carboxylase at Ser79 was also increased. AICAR treatment for 48 h halved progesterone secretion, 3beta-HSD protein and mRNA levels, and phosphorylation of both basal MAPK ERK1/2 and p38 and in response to IGF-I and/or FSH in granulosa cells. AICAR treatment (1 mM) had no detectable effect on basal and FSH- and/or IGF-I-induced estradiol production and on granulosa cell proliferation or viability. Adenovirus-mediated expression of dominant negative AMPK totally abolished the effects of AICAR on progesterone secretion, 3beta-HSD protein production, and MAPK ERK1/2 and p38 phosphorylation. Moreover, we showed using specific in- hibitors of ERK1/2 and p38 MAPK that the MAPK ERK1/2 and not p38 is involved in progesterone secretion and 3beta-HSD expression, strongly suggesting that the activation of AMPK in response to AICAR reduces progesterone production through the MAPK ERK1/2 signaling pathway in rat granulosa cells.
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PMID:Adenosine 5'-monophosphate-activated protein kinase regulates progesterone secretion in rat granulosa cells. 1602 Apr 77

FSH, a key regulator of gonadal function, contains a beta-subunit (FSHbeta) that is transcriptionally induced by activin, a member of the TGFbeta-superfamily. This study used 4.7 kb of the ovine FSHbeta-promoter linked to luciferase (oFSHbetaLuc) plus a well-characterized activin-responsive construct, p3TPLuc, to investigate the hypothesis that Smad3, TGFbeta-activated kinase 1 (TAK1), or both cause activin-mediated induction of FSH. Overexpression of either Smad3 or TAK1 induced oFSHbetaLuc in gonadotrope-derived LbetaT2 cells as much as activin itself. Induction of p3TPLuc by activin is known to require Smad3 activation in many cell types, and this was true in LbetaT2 cells, where 10-fold induction by activin (2-8 h after activin treatment) was blocked more than 90% by two dominant negative (DN) inhibitors of Smad3 [DN-Smad3 (3SA) and DN-Smad3 (D407E)]. By contrast, 6.5-fold induction of oFSHbetaLuc by activin (10-24 h after activin treatment) was not blocked by either DN-Smad inhibitor, suggesting that activation of Smad3 did not trigger induction of oFSHbetaLuc. By contrast, inhibition of TAK1 by a DN-TAK1 construct led to a 50% decrease in activin-mediated induction of oFSHbetaLuc, and a specific inhibitor of TAK1 (5Z-7-Oxozeanol) blocked induction by 100%, indicating that TAK1 is necessary for activin induction of oFSHbetaLuc. Finally, inhibiting p38-MAPK (often activated by TAK1) blocked induction of oFSHbetaLuc by 60%. In conclusion, the data presented here indicate that activation of TAK1 (and probably p38-MAPK), but not Smad3, is necessary for triggering induction of oFSHbeta by activin.
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PMID:Transforming growth factor beta-activated kinase 1 is a key mediator of ovine follicle-stimulating hormone beta-subunit expression. 1608 41

The pattern of GnRH release is associated with differential synthesis and release of LH and FSH. Using a perifusion system, we previously reported that stimulation of the LbetaT2 cell line with varying GnRH pulse frequencies resulted in differential stimulation of LHbeta and FSHbeta gene transcription, analogous to previous observations in primary gonadotropes. In the present study, we investigated the patterns of MAPK activation by GnRH and the role of MAPK in mediating the frequency-dependent effects. In static culture, ERK activation in LbetaT2 cells stimulated with continuous GnRH (10 nM) was maximal by 10 min and persisted for up to 6 h, with a return to basal levels by 20 h. In contrast, stimulation with continuous GnRH (10 nM) in perifused cells resulted in a more sustained activation of ERK. To investigate the effects of GnRH pulse frequency on ERK activation, perifused LbetaT2 cells were stimulated with pulsatile GnRH at a frequency of one pulse every 30 min or one pulse every 2 h for 20 h (10 nM, 5 min/pulse). After the final GnRH pulse, cells were lysed at frequent intervals and levels of ERK phosphorylation were measured. Under high-frequency conditions, ERK activation was maximal 10 min after the GnRH pulse and returned to baseline levels by 20 min. In contrast, under lower GnRH pulse frequency conditions, ERK activation occurred more rapidly and activation was more sustained, with a slower rate of ERK dephosphorylation. These changes resulted in different levels of nuclear phosphorylated ERK. Blockade of ERK activation abolished GnRH-dependent activation of LHbeta and FSHbeta transcription at both high and low pulse frequencies. These results demonstrate that in perifused LbetaT2 cells, distinct patterns of ERK activation/inactivation are regulated by GnRH pulse frequency, and the difference in ERK activation may be important for GnRH pulse frequency-dependent differential stimulation of LHbeta and FSHbeta gene expression.
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PMID:Gonadotropin-releasing hormone pulse frequency-dependent activation of extracellular signal-regulated kinase pathways in perifused LbetaT2 cells. 1614 98


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