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

We have previously shown that the stimulatory effect of TRH on alpha-MSH secretion from the frog pars intermedia is associated with Ca2+ influx through voltage-dependent Ca2+ channels, activation of a phospholipase C and mobilization of intracellular Ca2+ stores. The aim of the present study was to investigate the contribution of protein kinase C (PKC), adenylyl cyclase (AC), Ca2+/calmodulin-dependent protein kinase II (CAM KII), phospholipase A2, and protein tyrosine kinase (PTK) in TRH-induced alpha-MSH release. Incubation of frog neurointermediate lobes (NILs) with phorbol 12-myristate-13-acetate (24 h), which causes desensitization of PKC, or with the PKC inhibitor NPC-15437, reduced by approximately 50% of the effect of TRH on alpha-MSH release. In most melanotrope cells, TRH induces a sustained and biphasic increase in cytosolic Ca2+ concentration ([Ca2+]i). Preincubation with phorbol 12-myristate-13-acetate or NPC-15437 suppressed the plateau phase of the Ca2+ response. Incubation of NILs with TRH (10(-6) M; 20 min) had no effect on cAMP production. In addition, the AC inhibitor SQ 22,536 did not affect the secretory response of NILs to TRH. These data indicate that the phospholipase C/PKC pathway, but not the AC/protein kinase A pathway, is involved in TRH-induced alpha-MSH release. The calmodulin inhibitor W-7 and the CAM KII inhibitor KN-93 did not significantly reduce the response to TRH. Similarly, the phospholipase A2 inhibitors quinacrine and 7-7'-DEA did not impair the effect of TRH on alpha-MSH secretion. The PTK inhibitors ST638 and Tyr-A23 had no effect on TRH-induced [Ca2+]i increase but inhibited in a dose-dependent manner TRH-evoked alpha-MSH release (ED50 = 1.22x10(-5) M and ED50 = 1.47x10(-5) M, respectively). Taken together, these data indicate that, in frog melanotrope cells, PKC and PTK are involved in TRH-induced alpha-MSH secretion. Activation of PKC is responsible for the sustained phase of the increase in [Ca2+]i, whereas activation of PTK does not affect Ca2+ mobilization.
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PMID:Involvement of protein kinase C and protein tyrosine kinase in thyrotropin-releasing hormone-induced stimulation of alpha-melanocyte-stimulating hormone secretion in frog melanotrope cells. 1038 23

The hypothalamic hormone, TRH, stimulates PRL secretion and gene transcription. We have examined the possibility that the mitogen-activated protein kinase (MAPK) may play a role in mediating TRH effects on the PRL gene. TRH was found to stimulate sustained activation of MAPK in PRL-producing, GH3 cells, consistent with a possible role in transcriptional regulation. A kinase-defective, interfering MAPK kinase (MAPKK) mutant reduced TRH induction of the PRL promoter. Treatment with the MAPKK inhibitor, PD98059, blocked TRH-induced activation of MAPK and also reduced TRH induction of a PRL-luciferase reporter gene, confirming that MAPK activation is necessary for TRH effects on PRL gene expression. Previous studies have demonstrated that the PRL promoter contains binding sites for members of the Ets family of transcription factors, which are important for mediating MAPK responsiveness of the PRL promoter. Mutation of specific Ets sites within the PRL promoter reduced responsiveness to both TRH and MAPK. The finding that DNA elements required for MAPK responsiveness of the PRL gene colocalize with DNA elements required for TRH responsiveness further supports a role for MAPK in mediating TRH effects on the PRL gene. We also explored the signaling mechanisms that link the TRH receptor to MAPK induction. Occupancy of the TRH receptor results in activation of protein kinase C (PKC) as well as increases in the concentration of Ca2+ due to release from intracellular stores and entry of Ca2+ through Ca2+ channels. A PKC inhibitor, GF109203X, and an L-type Ca2+ channel blocker, nimodipine, both partially reduced TRH-induced MAPK activation and PRL promoter activity. The effects of the two inhibitors were additive. These studies are consistent with a signaling pathway involving PKC- and Ca2+-dependent activation of MAPK, which leads to phosphorylation of an Ets transcription factor and activation of the PRL promoter.
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PMID:A role for the mitogen-activated protein kinase in mediating the ability of thyrotropin-releasing hormone to stimulate the prolactin promoter. 1040 61

We studied rapid desensitization of the thyrotropin-releasing hormone receptor (TRH-R) or the m1-muscarinic receptor (m1-R) to a short challenge of threshold TRH concentration and persistent desensitization due to constitutive activity of a mutant TRH-R. Xenopus oocytes expressing TRH-Rs and/or m1-Rs were challenged for 15 s with threshold concentrations of TRH ([TRH]) and then immediately with supraoptimal [TRH] or acetylcholine ([ACh]). The threshold challenge caused desensitization of 50 - 57% of responses to subsequent supraoptimal stimulation with TRH or ACh. The homologous desensitization was reversible within 60 s after removal of the agonist. The protein kinase C (PKC) inhibitor, chelerythrine, inhibited the control responses by 30 - 40%, without affecting the desensitized responses. Chelerythrine or the phosphatase inhibitor, okadaic acid, had little effect on the kinetics of resensitization, indicating limited involvement of PKC. In oocytes coexpressing wild type TRH-Rs or m1-Rs with a constitutively active TRH-R mutant (C335Stop TRH-R), a persistent desensitization (33 - 57%) of the responses to TRH or ACh was observed. Additionally, there was a complete loss of the rapid desensitization induced by threshold [TRH]. Chlorodiazepoxide (CDE), a competitive binding antagonist of TRH-Rs and an inverse agonist of C335Stop TRH-Rs, abolished the persistent desensitization induced by C335Stop TRH-Rs and enabled the rapid desensitization, conferring the wild type phenotype on C335Stop TRH-Rs. Chelerythrine had qualitatively the same effect as CDE. In conclusion, unlike the rapid desensitization, the persistent desensitization caused by the constitutively active C335Stop TRH-Rs is largely mediated by PKC. It abrogates, however, the rapid desensitization, suggesting a common mechanistic step(s).
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PMID:Rapid desensitization of the TRH receptor and persistent desensitization of its constitutively active mutant. 1080 68

TRH has been found to stimulate tyrosine phosphorylation of the epidermal growth factor (EGF) receptor. A specific EGF receptor kinase inhibitor, tyrphostin AG1478, substantially reduced TRH-stimulated tyrosine phosphorylation of the EGF receptor. TRH-induced EGF receptor phosphorylation was found to lead to the recruitment of the adapter proteins Grb2 and Shc. TRH treatment also led to phosphorylation of the related receptor tyrosine kinase, HER2. HER2 activation likely contributes to downstream signaling events and enhances EGF receptor action. TRH-induced tyrosine phosphorylation of the EGF receptor was reduced by incubation with a protein kinase C (PKC) kinase inhibitor, GF109203X. EGF receptor phosphorylation was required for full TRH-induced activation of mitogen-activated protein kinase (MAPK) and stimulation of specific transcriptional responses.
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PMID:Thyrotropin-releasing hormone stimulates phosphorylation of the epidermal growth factor receptor in GH3 pituitary cells. 1097 12

To determine whether the interaction of the TRH receptor with beta-arrestin is necessary for TRH activation of MAPK, cells expressing either intact or truncated, internalization-defective TRH receptors were transfected with a beta-arrestin-green fluorescent protein conjugate. In cells expressing the wild-type pituitary TRH receptor, TRH caused translocation of the beta-arrestin-green fluorescent protein conjugate from the cytosol to the plasma membrane within 30 sec. After 5 min, the beta-arrestin-green fluorescent protein conjugate was visible in vesicles, where it colocalized with rhodamine-labeled TRH. In hypertonic sucrose, the beta-arrestin-green fluorescent protein conjugate translocated to the plasma membrane after TRH addition but did not internalize. In cells expressing the truncated TRH receptor, TRH did not cause translocation of the beta-arrestin-green fluorescent protein conjugate. TRH activated MAPK strongly in cells expressing intact or truncated TRH receptors, indicating that the receptor does not need to bind beta-arrestin or internalize. MAPK activation by TRH, epidermal growth factor, and phorbol ester was strongly inhibited by hypertonic sucrose and concanavalin A, which block movement of proteins into coated pits and coated pit assembly. Hypertonic sucrose did not affect MAPK activation in cells overexpressing MAPK kinase 1. Dominant negative dynamin, which blocks conversion of coated pits to vesicles, also reduced receptor internalization and TRH activation of MAPK. TRH activation of MAPK required PKC but was insensitive to pertussis toxin and did not require ras, epidermal growth factor receptor kinase, or PI3K. These results show that the TRH receptor itself does not need to bind beta-arrestin or undergo sequestration to activate MAPK but that the endocytic pathway must be intact.
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PMID:Activation of MAPK by TRH requires clathrin-dependent endocytosis and PKC but not receptor interaction with beta-arrestin or receptor endocytosis. 1151 3

In mammals, growth hormone (GH) is under a dual hypothalamic control exerted by growth hormone-releasing hormone (GHRH) and somatostatin (SRIH). We investigated GH release in a pleuronectiform teleost, the turbot (Psetta maxima), using a serum-free primary culture of dispersed pituitary cells. Cells released GH for up to 12 days in culture, indicating that turbot somatotropes do not require releasing hormone for their regulation. SRIH dose-dependently inhibited GH release up to a maximal inhibitory effect of 95%. None of the potential stimulators tested induced any change in basal GH release. Also, neither forskolin, an activator of adenylate cyclase, nor phorbol ester (TPA), an activator of protein kinase C, were able to modify GH release, suggesting that spontaneous basal release already represents the maximal secretory capacity of turbot somatotropes. In contrast, forskolin and TPA were able to increase GH release in the presence of SRIH. In this condition (coincubation with SRIH), pituitary adenylate cyclase-activating polypeptide (PACAP) stimulated GH release, whereas none of the other neuropeptides tested (GHRHs; sea bream or salmon or chicken II GnRHs; TRH; CRH) had any significant effect. These data indicate that inhibitory control by SRIH may be the basic control of GH production in teleosts and lower vertebrates, while PACAP may represent the ancestral growth hormone-releasing factor in teleosts, a role taken over in higher vertebrates by GHRH.
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PMID:Pituitary growth hormone secretion in the turbot, a phylogenetically recent teleost, is regulated by a species-specific pattern of neuropeptides. 1175 94

Although it is well known that plasma concentration of prolactin (PRL) increases during aging in rats, how the anterior pituitary (AP) aging per se affects PRL secretion remains obscure. The objectives of this study were to determine if changes in the pituitary PRL responsiveness to acetylcholine (ACh; a paracrine factor in the AP), as compared with that to other PRL stimulators or inhibitors, contribute to the known age-related increase in PRL secretion, and if protein kinase C (PKC) is involved. We also determined if replenishment with aging-declined hormones such as estrogen/thyroid hormone influences the aging-caused effects on pituitary PRL responses. AP cells were prepared from old (23-24-month-old) as well as young (2-3-month-old) ovariectomized rats. Cells were pretreated for 5 days with diluent or 17beta-estradiol (E(2); 0.6 nM) in combination with or without triiodothyronine (T(3); 10 nM). Then, cells were incubated for 20 min with thyrotropin-releasing hormone (TRH; 100 nM), angiotensin II (AII; 0.2-20 nM), vasoactive intestinal peptide (VIP; 10(-9)-10(-5) M), dopamine (DA; 10(-9)-10(-5) M), or ACh (10(-7)-10(-3) M). Cells were also challenged with ACh, TRH, or phorbol 12-myristate 13-acetate (PMA; 10(-6) M) following PKC depletion by prolonged PMA (10(-6) M for 24 h) pretreatment. We found that estrogen priming of AP cells could reverse the aging-caused effects on pituitary PRL responses to AII and DA. In hormone-replenished cells aging enhanced the stimulation of PRL secretion by TRH and PMA, but not by AII and VIP. Aging also reduced the responsiveness of cells to ACh and DA in suppressing basal PRL secretion, and attenuated ACh inhibition of TRH-induced PRL secretion. Furthermore, ACh suppressed TRH-induced PRL secretion mainly via the PMA-sensitive PKC in the old AP cells, but via additional mechanisms in young AP cells. On the contrary, basal PRL secretion was PKC (PMA-sensitive)-independent in the old AP cells, but dependent in the young AP cells. Taken together, these results suggest differential roles of PMA-sensitive PKC in regulating basal and ACh-regulated PRL responses in old versus young AP cells. The persistent aging-induced differences in AP cell responsiveness to ACh, DA, TRH, and PMA following hormone (E(2)/T(3)) replenishment suggest an intrinsic pituitary change that may contribute, in part, to the elevated in vivo PRL secretion observed in aged rats.
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PMID:Differential involvement of protein kinase C in basal versus acetylcholine-regulated prolactin secretion in rat anterior pituitary cells during aging. 1211 96

TRH is a neuropeptide that activates phospholipase C and, when acting on secretory cells, usually induces a biphasic response consisting of a transitory increase in secretion (due to IP(3) mobilization of Ca(2+) from intracellular stores), followed by a sustained plateau phase of stimulated secretion (by protein kinase C-dependent influx of extracellular Ca(2+) through voltage-operated Ca(2+) channels). The melanotrope cell of the amphibian Xenopus laevis displays a unique secretory response to TRH, namely a broad transient but no sustained second phase, consistent with the observation that TRH induces a single Ca(2+) transient rather than the classic biphasic increase in [Ca(2+)](i). The purpose of the present study was to determine the signal transduction mechanism utilized by TRH in generating this Ca(2+) signaling response. Our hypothesis was that the transient reflects the operation of only one of the two signaling arms of the lipase (i.e., either IP(3)-induced mobilization of internal Ca(2+) or PKC-dependent influx of external Ca(2+)). Using video-imaging microscopy it is shown that the TRH-induced Ca(2+) transient is dramatically attenuated under Ca(2+)-free conditions and that thapsigargin has no noticeable effect on the TRH-induced transient. These observations indicate that an IP(3)-dependent mechanism plays no important role in the action of TRH. PKC also does not seem to be involved because an activator of PKC did not induce a Ca(2+) transient and an inhibitor of PKC did not affect the TRH response. Experiments with a bis-oxonol membrane potential probe showed that the TRH response also does not underlie a PKC-independent mechanism that would induce membrane depolarization. We conclude that the action of TRH on the Xenopus melanotrope does not rely on the classical phospholipase C-dependent mechanism.
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PMID:TRH signal transduction in melanotrope cells of Xenopus laevis. 1216 Dec 5

Epidermal growth factor (EGF) and TRH both produce enhanced prolactin (PRL) gene transcription and PRL secretion in GH4 rat pituitary tumor cell lines. These agents also activate protein kinase C (PKC) in these cells. Previous studies have implicated the PKCepsilon isozyme in mediating TRH-induced PRL secretion. However, indirect studies using phorbol ester down-regulation to investigate the role of PKC in EGF- and TRH-induced PRL gene transcription have been inconclusive. In the present study, we examined the role of multiple PKC isozymes on EGF- and TRH-induced activation of the PRL promoter by utilizing general and selective PKC inhibitors and by expression of genes for wild-type and kinase-negative forms of the PKC isozymes. Multiple nonselective PKC inhibitors, including staurosporine, bisindolylmaleimide I, and Calphostin C, inhibited both EGF and TRH induced rat PRL promoter activity. TRH effects were more sensitive to Calphostin C, a competitive inhibitor of diacylglycerol, whereas Go 6976, a selective inhibitor of Ca(2+)-dependent PKCs, produced a modest inhibition of EGF but no inhibition of TRH effects. Rottlerin, a specific inhibitor of the novel nPKCdelta isozyme, significantly blocked both EGF and TRH effects. Overexpression of genes encoding PKCs alpha, betaI, betaII, delta, gamma, and lambda failed to enhance either EGF or TRH responses, whereas overexpression of nPKCeta enhanced the EGF response. Neither stable nor transient overexpression of nPKCepsilon produced enhancement of EGF- or TRH-induced PRL promoter activity, suggesting that different processes regulate PRL transcription and hormone secretion. Expression of a kinase inactive nPKCdelta construct produced modest inhibition of EGF-mediated rPRL promoter activity. Taken together, these data provide evidence for a role of multiple PKC isozymes in mediating both EGF and TRH stimulated PRL gene transcription. Both EGF and TRH responses appear to require the novel isozyme, nPKCdelta, whereas nPKCeta may also be able to transmit the EGF response. Inhibitor data suggest that the EGF response may also involve Ca(2+)-dependent isozymes, whereas the TRH response appears to be more dependent on diacylglycerol.
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PMID:Role of specific protein kinase C isozymes in mediating epidermal growth factor, thyrotropin-releasing hormone, and phorbol ester regulation of the rat prolactin promoter in GH4/GH4C1 pituitary cells. 1245 4

Abnormal GH responses to GnRH test, observed in about 15% of patients with acromegaly, have been reported exclusively in patients bearing tumors without gsp mutation. The absence of responsiveness to GnRH in gsp+ tumors was not predicted on the basis of the mechanism of GnRH action that mainly involves the activation of calcium and protein kinase C dependent pathways. The aim of the present study was to investigate in detail the transduction of GnRH signaling in these tumors. GH-secreting adenomas removed from patients in vivo responsive to GnRH test were studied. Tumor DNA was screened for Gsalpha and GnRH receptor gene sequences. Intracellular calcium ([Ca2+]i) and cAMP levels were measured in dispersed cells and adenylyl cyclase (AC) activity in membrane preparations. DNA analysis showed wild sequence of both Gsalpha and GnRH receptor genes. GnRH caused a significant increase in intracellular Ca2+ that was associated with a significant stimulation of cAMP accumulation. In these cells neither TRH nor GHRP-6 were effective in causing significant modifications of cAMP levels, despite their ability to increase [Ca2+]i. Finally, GnRH was able to directly stimulate AC from 11.1 +/- 3.3 pmol/mg prot/min to 26.9 +/- 5.4 (p<0.005). We report that GnRH was effective in increasing both [Ca2+]i and AC in GH-secreting adenomas removed from responsive patients. The ability of GnRH to signal through Gsalpha protein may account for the lack of GH responses to GnRH observed in acromegalic patients with tumors carrying gsp mutation.
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PMID:Gonadotropin-releasing hormone initiates multiple signaling pathways in human GH-secreting adenomas. 1523 51


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