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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have previously shown that the abundance of vitamin D receptors (VDR) in cultured cells is increased by mitogens such as serum and growth factors, whereas activation of protein kinase-C (PK-C) causes inhibition of VDR gene expression. This study examines the effect of the cAMP-activated protein kinase-A (PK-A) second messenger system on VDR abundance and 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] action. Elevation of intracellular cAMP levels in NIH-3T3 mouse fibroblasts by forskolin or (Bu)2cAMP caused a substantial (8- to 12-fold) increase in VDR abundance, as measured by ligand binding and Western blot analysis. The time course of the forskolin effect on VDR expression was complex. An early rise in VDR abundance occurred at 4 h, followed by a decrease and then a broad secondary rise at 18 h. At the mRNA level, forskolin caused a rapid rise in VDR transcripts after 1 h of exposure, a peak at 2 h, followed by a decline and a subsequent increase at 15 h. Activation of PK-C with the phorbol ester phorbol myristate acetate abolished the forskolin-induced increase in VDR protein and mRNA abundance. NIH-3T3 cells were stably transfected with phOC-CAT, a plasmid carrying a human osteocalcin promoter fragment containing the vitamin D response element fused to the reporter gene chloramphenicol acetyl transferase (CAT). 1,25-(OH)2D3 treatment of transfected cells induced a dose-dependent increase in CAT activity. Up- or down-regulation of VDR in these transfected cells by forskolin or phorbol myristate acetate pretreatment, respectively, resulted in corresponding enhancement or attenuation of 1,25-(OH)2D3-inducible CAT activity.(ABSTRACT TRUNCATED AT 250 WORDS)
Mol Endocrinol 1992 Feb
PMID:Cyclic adenosine 3',5'-monophosphate up-regulates 1,25-dihydroxyvitamin D3 receptor gene expression and enhances hormone action. 131 57

Cytochrome P-450scc (P-450scc) catalyzes the cholesterol side-chain cleavage reaction, a rate-limiting enzymatic step for progesterone synthesis in trophoblastic and other steroidogenic cells. Adrenodoxin is the iron/sulfur protein donating electrons to P-450scc during this reaction. We examined the effects of cholera toxin (CT), an activator of adenylate cyclase, and 12-O-tetradecanoylphorbol acetate (TPA), a phorbol ester protein kinase C activator, on the levels of mRNAs encoding P-450scc and adrenodoxin in JEG-3 choriocarcinoma cells. CT induced in a concentration- and time-dependent manner P-450scc and adrenodoxin mRNA levels to 8-fold and 1.5-fold above that of control, respectively. TPA also increased P-450scc and adrenodoxin mRNA levels about 3-fold and 1.5-fold above that of control, respectively. Epidermal growth factor (EGF) was found to weakly induce P-450scc mRNA accumulation with a maximal 20% stimulation above basal levels. The effects of CT and TPA were apparently additive on both mRNAs. The protein synthesis inhibitor cycloheximide diminished basal, CT-, TPA-, and EGF-stimulated P-450scc mRNA accumulation whereas the opposite was observed for the adrenodoxin mRNA. Insulin-like growth factor I (IGF-I) appeared to have no effect on either mRNA. These data indicate that: (1) the accumulation of P-450scc and adrenodoxin mRNAs is mainly controlled by the cyclic adenosine 3',5'-monophosphate (cAMP)-dependent pathway but their stimulation by TPA- and EGF-induced signals may also play a weaker synergistic role; (2) the protein synthesis inhibitor cycloheximide inhibits basal, CT-, TPA- and EGF-stimulated P-450scc mRNA levels while it increases the expression of adrenodoxin mRNA suggesting that in the malignant trophoblasts these two enzyme mRNAs are differentially controlled.
Mol Cell Endocrinol 1992 Apr
PMID:Regulation of the cholesterol side-chain cleavage cytochrome P-450 and adrenodoxin mRNAs in cultured choriocarcinoma cells. 131 54

1. Using internal perfusion and concentration-clamp procedures applied to Helix neurons, the effects of cAMP, Ca2+, and phorbol esters on ouabain-induced depression of acetylcholine Cl-dependent responses were determined. 2. Intracellular cAMP (10(-4) M) depressed those acetylcholine responses which were blocked by ouabain but had no effect on ouabain-insensitive acetylcholine responses. In the presence of elevated intracellular cAMP, ouabain had no further depressant effect on these acetylcholine responses. Both elevated cAMP and ouabain reduced the acetylcholine response without altering the current-voltage curves. 3. An increase in intracellular Ca2+ concentration depressed the amplitude of current induced by application of acetylcholine in neurons with ouabain-sensitive responses and shifted the dose-response relationship to the right. However, elevated Ca2+ did not reduce the maximal response induced by acetylcholine, nor did it prevent the reduction of that response by ouabain. 4. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a potent stimulator of protein kinase C activity, caused depression of both the ouabain-sensitive and the ouabain-insensitive acetylcholine responses. The inhibitory effect of TPA was markedly enhanced after addition of ATP to the intracellular medium and was greatly reduced by cooling to 5 degrees C. The blocking effect of ouabain, however, reexamined in the presence of TPA. 5. These observations are consistent with the hypothesis that the depression of acetylcholine induced Cl--responses in Helix neurons is a result of an increase in intracellular cAMP concentration but is unrelated to activation of protein kinase C or increases in intracellular Ca2+.
Cell Mol Neurobiol 1992 Apr
PMID:The effects of cAMP, Ca2+, and phorbol esters on ouabain-induced depression of acetylcholine responses in Helix neurons. 131 66

The effects of protein kinase C (PKC) activators on gamma-aminobutyric acidA (GABAA) receptor function were studied by two-electrode voltage-clamp in Xenopus oocytes expressing brain mRNA or subunit cDNAs and in isolated mouse brain cerebellar membrane vesicles (microsacs), using 36Cl- uptake. Both oocytes and microsacs showed transient (desensitizing) and sustained (nondesensitizing) GABAA receptor responses. In oocytes expressing brain mRNA, the PKC activator phorbol myristoyl acetate (PMA), but not the inactive analog phorbol 12-monomyristate, inhibited both transient and sustained GABA-gated chloride currents. The inhibition by PMA was concentration dependent, with an EC50 of approximately 5 nM, and resulted in a decrease in the efficacy, but not the potency, of GABA. Additionally, PMA inhibited GABA-gated chloride currents in oocytes expressing alpha 1 beta 1 gamma 2L subunit cDNAs. The effect of PMA on recombinant receptors was significantly antagonized by PKC inhibitory peptide (PKCI). In the microsac preparation, the PKC activators (-)-7-octylindolactam V and PMA inhibited the sustained phase of 36Cl- flux without altering the transient phase. The action of PMA was blocked by kinase inhibitors and by depletion of Mg-ATP and was mimicked by protein phosphatase inhibitors. These results demonstrate that activation of PKC inhibits GABAA receptor function, and the results from the microsac experiments suggest that PKC-dependent phosphorylation preferentially inactivates a nondesensitized form or state of the receptor.
Mol Pharmacol 1992 Jun
PMID:Activation of protein kinase C selectively inhibits the gamma-aminobutyric acidA receptor: role of desensitization. 131 47

The phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) is a potent activator of protein kinase C (PKC) and is known to affect a variety of biochemical processes in human breast cancer cells. In the present study we have employed MCF-7 cells to investigate the effects of TPA on inositol lipid signalling, the putative pathway leading to PKC activation and intracellular Ca2+ mobilization. Phosphoinositide hydrolysis in MCF-7 cells was stimulated by bombesin (BN) as evidenced by increases in both inositol phosphate production and cytidine diphosphate diacylglycerol (CDP-DG) accumulation. Pretreatment of MCF-7 cells with TPA caused attenuation of both these BN-induced responses. This inhibitory action of TPA on inositol phosphate production was mimicked by diacylglycerol analogues and was reversed by staurosporine, H-7 and tamoxifen, all known inhibitors of PKC. Furthermore, putative down-regulation of PKC by prolonged TPA pretreatment also reversed the inhibitory action of TPA and enhanced BN-induced phosphoinositide hydrolysis. TPA also inhibited BN-induced increases in cytosolic Ca2+ concentration ([Ca2+]i) and caused a dose-dependent inhibition of epidermal growth factor (EGF) binding in MCF-7 cells. However, EGF receptor occupancy was unaffected by BN. These data support an inhibitory role for PKC in the regulation of phosphoinositide hydrolysis and [Ca2+]i in breast cancer cells and provide a potential mechanism for feedback regulation of this signalling pathway in these cells.
Mol Cell Endocrinol 1992 Jun
PMID:Evidence for a role for protein kinase C in the modulation of bombesin-activated cellular signalling in human breast cancer cells. 132 70

The regulation of the guinea-pig pancreatic acinar plasma membrane Ca2+ pump by protein kinase A, protein kinase C and calmodulin was investigated. The results were compared with the effects of these regulators on the high affinity Ca(2+)-ATPase found in this membrane preparation. The catalytic subunit of cyclic AMP-dependent protein kinase stimulated Ca2+ transport 2-fold, but had no effect on Ca(2+)-dependent ATPase activity. Purified protein kinase C, the phorbol ester 12-O-tetradecanoyl phorbol-13-acetate and diacylglycerol derivative, 1-stearoyl-2-arachidonoyl-sn-glycerol, failed to stimulate the Ca(2+)-uptake but augmented the Ca(2+)-dependent ATPase activity. Exogenously added calmodulin failed to stimulate either activity. In addition, two antagonists of calmodulin activity, trifluoperazine and compound 48/80 produced a concentration-dependent inhibition of Ca(2+)-transport. These data suggest the presence of endogenous calmodulin within guinea-pig pancreatic acinar plasma membranes. Both calmodulin antagonists failed to influence the Ca(2+)-dependent ATPase activity. The ability of boiled extracts from guinea-pig pancreatic acinar plasma membranes to stimulate the Ca(2+)-ATPase activity in calmodulin-depleted erythrocyte plasma membranes confirmed the presence of endogenous calmodulin. Our results imply a role for calmodulin and cAMP-dependent protein kinase, but not protein kinase C, in the regulation of Ca2+ efflux from pancreatic acinar cells. These results also provide further evidence suggesting that the high affinity Ca(2+)-ATPase does not catalyze the plasma membrane Ca(2+)-transport activity observed in pancreatic acini.
Mol Cell Biochem 1992 Jun 26
PMID:Regulation of calcium transport in pancreatic acinar plasma membranes from guinea pig. 132 90

Protein phosphorylation catalyzed by the cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA) is implicated in regulating zygotic gene activation in the two-cell mouse embryo (Poueymirou and Schultz; Dev Biol 133:588-599, 1989). We now provide evidence that H8, which is a PKA inhibitor, inhibits expression of an hsp70-driven beta-galactosidase reporter gene and that the concentration-dependence of this inhibition is similar to that for inhibiting expression of a stage-specific gene(s) that is a product of zygotic gene activation. We also demonstrate that neither cAMP nor serum can stimulate the expression, as detected by a histochemical assay, of a cAMP response element (CRE)- or serum response element (SRE)-driven beta-galactosidase reporter gene, respectively, in either germinal vesicle-intact oocytes or aphidicolin-arrested one-cell embryos that are chronologically at the tw-cell stage. In contrast, although 12-O-tetradecanoyl phorbol-13-acetate (TPA) does not stimulate expression of a TPA response element (TRE)-driven beta-galactosidase reporter gene in germinal vesicle-intact oocytes, it stimulates such expression in aphidicolin-arrested one-cell embryos. Moreover, TPA can stimulate the expression of either a CRE- or an SRE-driven beta-galactosidase reporter gene in such embryos. Results of these studies further implicate protein phosphorylation in regulating zygotic gene activation, along with its role in modulating enhancer function in the early mouse embryo.
Mol Reprod Dev 1992 Jul
PMID:Zygotic gene activation in the mouse embryo: involvement of cyclic adenosine monophosphate-dependent protein kinase and appearance of an AP-1-like activity. 132 5

Primary fetal human adrenocortical cells of definitive zone origin were transfected by electroporation with pSV3neo, a plasmid coding for SV40 T antigen and neo, which confers resistance to the antibiotic G418. The clones obtained proliferated for 30 to 40 population doublings after isolation when grown under standard medium conditions, and then entered 'crisis'. When early-passage clones were incubated with cyclic AMP (1:1 N6-monobutyryl and 8-bromo analogues), cell rounding was observed, as in primary cultures of human adrenocortical cells. As previously shown in bovine adrenocortical cells, rounding was inhibited with a monoclonal antibody against urokinase plasminogen activator but not with a monoclonal antibody against tissue plasminogen activator. The regulation of the steroidogenic pathway in clones was investigated. The effects of cyclic AMP and activation of protein kinase C were examined in cells maintained in defined medium or in the presence of serum. 17 alpha-Hydroxylase was strongly induced by cyclic AMP, as evidenced by Northern blotting and by the conversion of progesterone or 25-hydroxy-[1,2-3H]cholesterol, this induction being blocked by low concentrations of 12-O-tetradecanoylphorbol-13-acetate (TPA). Cholesterol side-chain cleavage enzyme was strongly induced by cyclic AMP, and clones also showed low activities of 21-hydroxylase and 11 beta-hydroxylase. Under all circumstances levels of 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD), as assessed by Northern blotting or by conversion of 25-hydroxycholesterol, were very low. 3 beta-HSD was not induced by cyclic AMP or TPA alone, but was induced by the combination of the two agents. The regulation of 17 alpha-hydroxylase and 3 beta-HSD resembles that previously described in primary cultures of human fetal adrenocortical cells. Thus, transfection with SV40 T antigen resulted in the production of clones which preserve the unique characteristics of the human adrenal cortex.
J Mol Endocrinol 1992 Aug
PMID:Expression of 17 alpha-hydroxylase and 3 beta-hydroxysteroid dehydrogenase in fetal human adrenocortical cells transfected with SV40 T antigen. 132 52

Although there is increasing evidence of a pathogenic role for eosinophils in the airway epithelium, there is little direct evidence which demonstrates that eosinophils influence epithelial cell activity in humans. We have cultured human nasal epithelial cells in vitro and studied the effect of isolated human eosinophils on the ciliary beat frequency (CBF) and cell membrane integrity of these cells after incubation in the absence or presence of 0.1 microM phorbol 12-myristate 13-acetate (PMA) or 0.1 mg/ml opsonized latex beads and the absence or presence of 10(-5) M nedocromil sodium. CBF was monitored by an analogue contrast-enhancement technique, and cell damage was assessed by release of 51Cr from the cells. Cell cultures were also assessed for the percentage of eosinophil cationic protein (ECP) released into the medium at the end of incubation. Neither 0.1 microM PMA, 0.1 mg/ml opsonized latex beads, 10(-5) M nedocromil sodium, nor eosinophils alone altered the CBF of the epithelial cells. PMA-stimulated eosinophils, however, attenuated the CBF significantly, from 10.2 +/- 0.3 to 8.8 +/- 0.4 Hz (P less than 0.05) after 15 h of incubation. Similarly, opsonized latex bead-stimulated eosinophils led to a significant attenuation of CBF from 9.2 +/- 0.3 to 8.4 +/- 0.3 Hz (P less than 0.05), 6.9 +/- 0.5 Hz (P less than 0.001), and 7.5 +/- 0.3 Hz (P less than 0.001) after 2, 15, and 24 h of incubation, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
Am J Respir Cell Mol Biol 1992 Sep
PMID:The effect of human eosinophils on cultured human nasal epithelial cell activity and the influence of nedocromil sodium in vitro. 132 9

A hormone-inducible transcriptional system has been established, based on the stable transfection of the rat androgen receptor (rAR) and a reporter plasmid containing the mouse mammary tumour virus promoter linked to the chloramphenicol acetyltransferase gene (pMMTV-CAT) into steroid receptor-negative CV-1 cells. First, the rAR was stably introduced into CV-1 cells. Single clones were tested for stable expression of functionally active AR by analysing the effect of dihydrotestosterone on induction of transiently transfected pMMTV-CAT. Stable transfection and the expression of AR was confirmed by steroid-binding assays. In a second step, a clone expressing physiological amounts of AR protein (30 fmol/mg protein) was stably transfected with pMMTV-CAT to yield a permanent cell line that stably expresses functional AR and MMTV-CAT sequences. This cell line provides a powerful tool for the efficient and accurate determination and quantification of the effects of androgens and anti-androgens on reporter gene transcription. This was demonstrated by investigating the action of the three anti-androgens hydroxyflutamide, casodex and cyproterone acetate. The three compounds were shown to reverse the effects of the androgen R1881 on gene expression but were themselves devoid of agonistic activity.
J Steroid Biochem Mol Biol 1992 Sep
PMID:Stable transfection of androgen receptor and MMTV-CAT into mammalian cells: inhibition of cat expression by anti-androgens. 132 16


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