Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

GHRP6 is a synthetic hexapeptide which stimulates growth hormone (GH) secretion from the pituitary in vivo and in vitro. We have previously shown that in identified somatotrophs, GHRP6 induces a biphasic increase in cytosolic Ca2+ concentration ([Ca2+]i) consisting of an abrupt increase (first phase) followed by a sustained plateau of elevated [Ca2+]i (second phase). The first phase corresponds to mobilization of intracellular Ca2+ pools and the second phase to influx of extracellular Ca2+ ions through voltage-sensitive Ca2+ channels. In these experiments, we investigated the specific role of each of these two phases in the hormone response to GHRP6. We found that inhibition by thapsigargin of the intracellular Ca2+ mobilization phase significantly inhibited the hormone response to the peptide during 30 min incubations. Inhibition of the extracellular Ca2+ influx phase by nifedipine, a blocker of voltage-sensitive Ca2+ channels, resulted in a 53 percent reduction of the secretory response to 10(-5)M GHRP6. Antagonism of PKC by phloretin, a flavonoid which prevents PKC activation, and PKC depletion induced by a 24 h treatment with 10(-6)M PMA, completely inhibited the response to GHRP6. Somatostatin, which also inhibits the second phase of the Ca2+ response, suppressed the secretory response to GHRP6. We conclude that, Ca2+ is the main second messenger and both Ca2+ mobilization and Ca2+ entry play a role in the response to GHRP6. However, experiments with PKC depletion and SRIF suggest that other messengers are implicated in GHRP6 signalling in somatotrophs.
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PMID:GHRP6-stimulated hormone secretion in somatotrophs: involvement of intracellular and extracellular calcium sources. 886 Dec 87

In cultured pituitary cells of tilapia, gonadotropin-releasing hormone (GnRH; 10 nM 4-24 h), elevation of cyclic AMP (by 10 microM forskolin or 0.2 mM 3-isobutyl-1-methylxanthine: IBMX 0.5-36 h) or activation of protein kinase C (PKC; by 12.5 nM tetradecanoyl phorbol-13-acetate: TPA, 0.5-24 h) all increased gonadotropin (GtH) II beta steady state mRNA levels by three to four-fold. The involvement of PKA and PKC in the GnRH stimulatory effect on both GtH release and GtH II beta mRNA levels was corroborated by use of the PKA and PKC inhibitors, H89 and GF109203X, respectively (100 nM) which attenuated the GnRH effect. Incubation with actinomycin D (8 microM, 4-21 h) after preexposure for 24 h to either forskolin (10 microM) or TPA (12.5 nM), revealed that rates of transcript degradation were slower in forskolin-treated cells (T 1/2 = 14.1 h) than in control or TPA-treated cells (T 1/2 = 8.47 or 8.38 h), suggesting a stabilizing effect on the mRNA. Dopamine (DA; 10 microM, 4-36 h) had no apparent effect on steady state mRNA levels of GtH II beta, but reduced GtH release by as much as 75%. Steady state levels of growth hormone (GH) mRNA were not affected by exposure to GnRH (10 nM, 4-24 h), although GH release was more than doubled. Similarly, activation of PKC (by TPA 12.5 nM, 1.5-36 h), which was shown to be essential for the GnRH-stimulatory effect on GH release, did not alter levels of the GH transcript, but increased GH release by more than fivefold. DA (10 microM, 4-24 h) moderately increased GH transcript levels (160%) with similar kinetics but lower potency than direct elevation of cAMP (by 10 microM forskolin or 0.2 mM IBMX, 0.5-36 h) which increased transcript levels by more than fourfold. The involvement of PKA in the DA effect was confirmed when the PKA inhibitor H89 (100 nM, 15 min prior to DA exposure) attenuated the DA effect on GH mRNA levels. Exposure of cells to actinomycin D (8 microM, 2-16 h) after treatment with forskolin (10 microM, 24 h) led to a slower rate of transcript degradation than in control cells (T 1/2 = 6.5 h vs. T 1/2 = 4.36 h), suggesting that cAMP also elicits a stabilizing effect on GH mRNA. Somatostatin (100 nM, 0.5-36 h) had no clear effect on GH transcript levels, but reduced GH release by as much as 90%. These results suggest that activation of either cAMP-PKA or PKC pathways can, possibly by different mechanisms, stimulate mRNA levels of the GtH II beta gene, but that only the cAMP-PKA pathway stimulates GH mRNA levels. It would appear therefore that GnRH, although stimulating GH release, does not regulate GH transcription in this fish.
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PMID:Differential effects of gonadotropin-releasing hormone, dopamine and somatostatin and their second messengers on the mRNA levels of gonadotropin II beta subunit and growth hormone in the teleost fish, tilapia. 889 62

In goldfish, gonadotropin-releasing hormone (GnRH) stimulation of growth hormone (GH) release has been shown to involve extracellular Ca2+ entry through voltage-sensitive Ca2+ channels and the activation of protein kinase C (PKC). In this study, the possible involvement of extracellular Na+ in mediating the GH response to GnRH was examined using dispersed pituitary cells. Perifusion with Na(+)-depleted medium reversibly reduced the acute GH response to 5-min pulses of either 10 nM salmon (s)GnRH or 10 nM chicken (c)GnRH-II. Similarly, replacement of normal medium with Na(+)-depleted medium attenuated the long-term GH release response to sGnRH and cGnRH-II under static incubation conditions. These results suggest that GnRH-induced GH release requires the presence of extracellular Na+. Treatment with 5-min pulses of the Na(+)-channel agonist veratridine (10 microM) increased GH release in an extracellular Ca(2+)-dependent manner, presumably due to activation of voltage-sensitive Ca2+ channels resulting from the depolarizing effect of increased Na+ influx. On the other hand, Na+ entry through tetrodotoxin (TTX)-sensitive, voltage-dependent Na+ channels is not involved in GnRH-induced GH release. Application of 250 nM TTX, which abolished the voltage-sensitive Na+ currents in identified goldfish somatotropes, did not affect the acute GH responses to 5-min pulses of sGnRH and cGnRH-II. The possible participation of Na+/H+ antiport in mediating the extracellular Na(+)-dependent GnRH action on GH release was then examined. In static incubation experiments, sGnRH- and cGnRH-II-induced GH secretion were reduced by inhibitors of the Na+/H+ antiport, amiloride and dimethylamiloride (DMA). Likewise, the GH response to the PKC activator, tetradecanoyl phorbol acetate, was attenuated by treatment with Na(+)-depleted medium, amiloride, and DMA. The inhibitory actions of amiloride and DMA were selective as these drugs did not affect the GH response elicited by the Ca2+ ionophore ionomycin and the voltage-sensitive Ca2+ channel agonist, Bay K 8644. Taken together, these results indicate that extracellular Na+ and the Na+/H+ exchanger are involved in the mediation of GnRH-stimulated GH release in goldfish. Furthermore, this dependence on Na+ and Na+/H+ antiport probably occurs distal to the activation of PKC by GnRH.
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PMID:Extracellular sodium dependence of GnRH-stimulated growth hormone release in goldfish pituitary cells. 908 72

Previous studies have shown that the activation of p44 and p42 mitogen-activated protein (MAP) kinases (ERK1 and ERK2) by growth hormone (GH) and phorbol esters, but not by epidermal growth factor, in 3T3-F442A preadipocytes is dependent on protein kinase C (PKC). In the present study two approaches have been taken to determine the PKC isoform dependence of MAP kinase activation in these cells. By immunoblotting with specific antibodies, the cells were found to express PKC-alpha, -gamma,-delta, -epsilon and -zeta. Treatment of cells with 500 nM PMA for 3 h led to the complete depletion of PKC-delta and the partial depletion of PKC-alpha but did not significantly affect the expression of the other PKC isoforms. In parallel, such treatment severely attenuated the ability of GH to activate MAP kinase. The degree of this attenuation was not increased by more prolonged PMA pretreatment, indicating that PKC-delta and perhaps PKC-alpha are important for MAP kinase activation by GH. These experiments further revealed that additional PKC isoforms were required for the full activation of MAP kinases by acute treatment with PMA. A second approach involved the use of anti-sense oligodeoxynucleotides (ODNs) to deplete the individual PKC isoforms selectively. Each of the ODNs used effectively depleted the relevant isoform to undetectable levels and did not affect the expression of the other PKC isoforms. Pretreatment of cells with PKC-delta anti-sense ODN, but not with anti-sense ODN to the other phorbol ester-sensitive isoforms, severely attenuated the activation of MAP kinases by GH. PKC-delta anti-sense ODN also blocked (by approx. 50%) the activation of MAP kinases by PMA. Furthermore a combination of PKC-delta and -epsilon anti-sense ODNs completely blocked the effect of PMA on MAP kinases. Collectively, these results indicate that the novel PKC-delta and -epsilon isoforms can couple to the MAP kinase pathway in 3T3-F442A cells but that the activation of MAP kinases by GH specifically involves PKC-delta.
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PMID:Growth hormone and phorbol esters require specific protein kinase C isoforms to activate mitogen-activated protein kinases in 3T3-F442A cells. 916 52

We conducted this study to investigate the mechanisms of action of growth hormone-releasing peptide-2 (D-Ala-D-beta Nal-Ala-Trp-D-Phe-Lys-NH2; GHRP-2) in bovine anterior pituitary primary cell culture. Doses of GHRP-2 from 10(-13) to 10(-7) M) increased (P < .05) GH secretion. The GHRP-2 (10(-7) M) and GH-releasing factor (GRF; 10(-7) M) administered together had an additive effect on the release of GH (P < .05). Somatostatin (1 microM) decreased GH secretion in response to GHRP-2 and(or) GRF (P < .05). Secretion of GH in response to GHRP-2 was blocked (P < .01) by a GRF receptor antagonist (.1 microM). Nifedipine (10 microM), a voltage-dependent Ca2+ channel blocker, inhibited (P < .01) GHRP-2-stimulated GH release. The GH release in response to GHRP-2 and 4 beta-phorbol-12-myristate-13-acetate (10(-7) M), a protein kinase C activator, was additive (P < .01). Forskolin (30 microM), a cAMP elevating agent, further stimulated (P < .01) the GH release in response to GHRP-2. Bovine GH concentrations in culture media were assayed by indirect competitive enzyme immunoassay. These results showed that GHRP-2 1) stimulates GH secretion from bovine pituitary cells, 2) may partially act via GRF receptor, 3) has GH secretion activity caused by Ca2+ influx via Ca2+ channels, and 4) may increase GH secretion via protein kinase C and cAMP pathways.
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PMID:Mechanisms of action of growth hormone-releasing peptide-2 in bovine pituitary cells. 933 79

Previous studies have demonstrated that growth hormone (GH) release in goldfish is under the stimulatory control of gonadotropin-releasing hormone (GnRH) and dopamine and the inhibitory control of somatostatin (SRIF). GnRH stimulation is mediated through protein kinase C (PKC)- and calcium-dependent mechanisms, whereas dopamine D1 receptor activation increases GH secretion through cyclic (c) AMP-dependent intracellular signal transduction pathways. In this study, the mechanisms of SRIF inhibition on GH secretion were examined using primary cultures of dispersed goldfish pituitary cells in static incubation. Application of 1 microM SRIF inhibited the GH-release responses to 100 nM salmon GnRH, 100 nM chicken GnRH-II, and 1 microM SKF38393, a D1 agonist. These results indicate that inhibitory action of SRIF on stimulated GH release is direct, at the level of the pituitary cells. Addition of SRIF reduced the GH release responses to two activators of PKC (100 microM dioctanoyl glycerol and 100 nM tetradecanoyl phorbol acetate) and to two ionophores (10 microM A23187 and 10 microM ionomycin). Similarly, SRIF abolished the GH responses to an activator of adenylate cyclase (10 microM forskolin), a membrane-permeant cAMP analog (1 mM 8-bromo-cAMP), and a voltage-sensitive calcium channel agonist (1 microM Bay K 8644). Taken together, these observations indicate that the inhibitory actions of SRIF on D1- and GnRH-stimulated GH release can be exerted at sites distal to cAMP production and PKC activation, respectively. SRIF also exerts its effect at sites distal to calcium mobilization. Since SRIF inhibition was more effective against Bay K 8644-induced response than against ionophore-induced GH response, an inhibitory action at the level of extracellular calcium entry through voltage-sensitive channels is also possible.
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PMID:Somatostatin inhibition of growth hormone release in goldfish: possible targets of intracellular mechanisms of action. 940 21

To investigate whether D(+)-glucose has a stimulatory effect on the expression of the angiotensinogen (Ang) gene in opossum kidney (OK) cells, we used OK cells with a fusion gene containing various lengths of the 5'-flanking regulatory sequence of the rat Ang gene fused with the human growth hormone (hGH) gene as a reporter, stably integrated into their genomes. The level of expression of the fusion gene was quantified by the amount of immunoreactive-human growth hormone (IR-hGH) secreted into the medium. The addition of D(+)-glucose stimulated the expression of pOGH (Ang N-1498/+18) in OK 27 cells in a dose-dependent manner (5 to 25 mM), whereas the addition of D-mannitol, L-glucose and 2-deoxy-D-glucose (25 mM) had no effect. The stimulatory effect of D(+)-glucose (25 mM) was blocked by the presence of staurosporine or H7 (an inhibitor of protein kinase C) or U73122 (an inhibitor of phospholipase C and A2) but not blocked by the presence of Rp-cAMP (an inhibitor of cAMP-dependent protein kinase A). The addition of D(+)-glucose (25 mM) also stimulated the expression of pOGH (Ang N-960/+18) and pOGH (Ang N-688/+18) in OK 960 and OK 688 cells, respectively. It had no stimulatory effect, however, on the expression of pOGH (Ang N-280/+18) and pOGH (Ang N-35/+18) in OK 280 and OK 35 cells, respectively. The addition of D(+)-glucose also had no effect on the expression of pTKGH in OK 13 cells, an OK cell line, into which had been stably integrated a fusion gene, pTKGH containing the promoter/enhancer DNA sequence of the viral thymidine-kinase (TK) gene fused with a human growth hormone gene as a reporter. These studies demonstrate that the stimulatory effect of high D(+)-glucose concentration (25 mM) on the expression of the angiotensinogen-growth hormone fusion genes in OK cells is mediated via the 5'-flanking region of the angiotensinogen gene and the protein kinase C signal transduction pathway. Our data indicate that a high glucose concentration may activate the renin-angiotensin system in the renal proximal tubular cells.
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PMID:Effect of glucose on the expression of the angiotensinogen gene in opossum kidney cells. 946 Oct 91

Surfactant protein (SP)-A gene transcription is stimulated by factors that increase cyclic AMP. In the present study, we observed that three thyroid transcription factor-1 (TTF-1) binding elements (TBEs) located within a 255 base pair region flanking the 5'-end of the baboon SP-A2 (bSP-A2) gene are required for maximal cyclic AMP induction of bSP-A2 promoter activity. We found that TTF-1 DNA binding activity was increased in nuclear extracts of pulmonary type II cells cultured in the presence of cyclic AMP. By contrast, the levels of immunoreactive TTF-1 protein were similar in nuclear extracts of control and cyclic AMP-treated type II cells. The incorporation of [32P]orthophosphate into immunoprecipitated TTF-1 protein also was markedly increased by cyclic AMP treatment. Moreover, exposure of nuclear extracts from cyclic AMP-treated type II cells either to potato acid phosphatase or alkaline phosphatase abolished the cyclic AMP-induced increase in TTF-1 DNA-binding activity. Interestingly, the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), known to activate protein kinase C, also enhanced incorporation of [32P]orthophosphate into TTF-1 protein; however, the DNA binding activity of TTF-1 was decreased in nuclear extracts of TPA-treated type II cells. Expression vectors encoding TTF-1 and the catalytic subunit of protein kinase A (PKA-cat) were cotransfected into A549 lung adenocarcinoma cells together with an SPA:human growth hormone fusion gene (255 base pairs of 5'-flanking DNA from the baboon SP-A2 gene linked to human growth hormone, as reporter) containing TBEs, or with a reporter gene construct containing three tandem TBEs fused upstream of the bSP-A2 gene TATA box and the transcription initiation site. Coexpression of TTF-1 and PKA-cat increased fusion gene expression 3-4-fold as compared with expression of TTF-1 in the absence of PKA-cat. Moreover, the transcriptional activity of TTF-1 was suppressed by cotransfection of a dominant negative form of PKA regulatory subunit RIalpha. We suggest that a PKA-induced increase of TTF-1 phosphorylation and TBE binding activity mediates cyclic AMP-induced expression of the SP-A gene in lung type II cells.
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PMID:Cyclic AMP-responsive expression of the surfactant protein-A gene is mediated by increased DNA binding and transcriptional activity of thyroid transcription factor-1. 946 16

The challenge of 3T3-F442A fibroblasts with growth hormone led to both a decrease in the mobility on SDS/PAGE and activation of the PDE4A cyclic AMP-specific phosphodiesterase isoform PDE4A5. Activation was mediated by a JAK-2-dependent pathway coupled to the activation of phosphatidylinositol 3-kinase and p70S6 kinase. Activation was not dependent on the ability of growth hormone to stimulate ERK2 or protein kinase C or any effect on transcription. Blockade of activation of murine PDE4A5 ablated the ability of growth hormone to decrease intracellular cAMP levels. Antisense depletion of murine PDE4A5 mimicked the ability of rolipram to enhance the growth hormone-stimulated differentiation of 3T3-F442A cells to adipocytes. It is suggested that activation of PDE4A5 by growth hormone serves as a brake on the differentiation processes.
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PMID:Stimulation of p70S6 kinase via a growth hormone-controlled phosphatidylinositol 3-kinase pathway leads to the activation of a PDE4A cyclic AMP-specific phosphodiesterase in 3T3-F442A preadipocytes. 952 Apr 3

We have previously shown that 10-12 kDa N-terminal fragments of rat proopiomelanocortin (POMC) and human POMC1-76 stimulate mitosis and/or differentiation in lactotrophs of early postnatal rat pituitary. A truncated form, POMC1-26, mimics the differentiation-inducing but not the mitogenic action of the former peptides. To further characterize these two biological responses, the present study compared changes in the intracellular free calcium concentration ([Ca2+]i) in response to POMC1-76 and POMC1-26 in isolated pituitary cells from 14-day-old female rats. Calcium (Ca2+) responses were also used as a guide to determine whether the responsive cells belong to the lactosomatotroph lineage. Application of POMC1-76 or POMC1-26 induced a maintained oscillating [Ca2+]i increase in a small population of cells. Increasing doses of the peptides did not affect the magnitude and the frequency of [Ca2+]i oscillations but clearly augmented the number of responding cells. Approximately 2% of the cells responded at 0.1 nM POMC1-76 or 5 nM POMC1-26, and 11-13% of the cells responded at 10 nM and 500 nM of the respective peptides. About one-third of the cells responsive to these peptides also showed a [Ca2+]i increase in response to growth hormone-releasing peptide-6 (GHRP-6) while, in a small number of responsive cells, [Ca2+]i was depressed by dopamine, suggesting that the former cells are somatotrophs and the latter lactotrophs. This interpretation was confirmed by immunocytochemical identification of prolactin and growth hormone (GH) in the cells. Of the cells showing Ca2+ response to POMC1-76, approximately one-third contained GH and another third prolactin. The remainder contained neither GH nor prolactin. Comparable results were obtained with POMC1-26. The rise of [Ca2+]i induced by the N-terminal POMC peptides persisted after depletion of intracellular Ca2+ stores by thapsigargin. Removal of Ca2+ from the extracellular medium or addition of cadmium completely abolished both the POMC1-76- and POMC1-26-induced [Ca2+]i increase. Nifedipine inhibited the Ca2+ response to both peptides, although only in 55% of the responsive cells. Depletion of some isoforms of protein kinase C by preincubation with the phorbol ester PMA for 24 h did not modify the Ca2+ responses. In contrast, blockade of the protein kinase A pathway with Rp-cAMPs partially inhibited the POMC1-76- or POMC1-26-induced [Ca2+]i increase. The present data show that, in immature pituitary cells, POMC1-76 induces an increase in [Ca2+]i through extracellular Ca2+ influx, possibly mediated in part by protein kinase A activation. The active domain of POMC1-76 seems to comprise its N-terminal moiety. The data support the hypothesis that POMC1-76 exerts a specific function in the development of different members of the lactosomatotroph lineage and that the peptide mobilizes different subsets of cells within this lineage, by a mechanism determined by its concentration.
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PMID:Stimulation of Ca2+ entry in lactotrophs and somatotrophs from immature rat pituitary by N-terminal fragments of proopiomelanocortin. 957 10


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