Phorbol ester promotes histone H3-Ser10 phosphorylation at the LDL receptor promoter in a protein kinase C-dependent manner

Tamoxifen inhibits bone resorption by disrupting calmodulin-dependent processes. Since tamoxifen inhibits protein kinase C in other cells, we compared the effects of tamoxifen and the phorbol ester, phorbol myristate acetate, on osteoclast activity. Phorbol esters stimulate bone resorption and calmodulin levels four-fold (k0.5 = 0.10.3 µM). In contrast, tamoxifen inhibited osteoclast activity ~60% with an IC50 of 1.5 µM, had no apparent effect on protein kinase C activity in whole-cell lysates, and reduced protein kinase Cα recovered by immunoprecipitation 75%. Phorbol esters stimulated resorption in a time-dependent manner that was closely correlated with a similar-fold increase in calmodulin. Protein kinase Cα, β, δ, ε, and ζ were all down-regulated in response to phorbol ester treatment. Tamoxifen and trifluoperazine inhibited PMA-dependent increases in bone resorption and calmodulin by 85 ± 10%. Down-regulation of protein kinase C isoforms by phorbol esters suggests that the observed increases in bone resorption and calmodulin levels are most likely due to a mechanism independent of protein kinase C and dependent on calmodulin. In conclusion, the data suggest that protein kinase C negatively regulates calmodulin expression and support the hypothesis that the effects of both phorbol esters and tamoxifen on osteoclast activity is mediated by calmodulin.Key words: osteoclast, calmodulin, tamoxifen, osteoporosis, protein kinase C.

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Post-transcriptional regulation of apolipoprotein E expression in mouse macrophages by phorbol ester

Biochemical Journal ◽

10.1042/bj2920105 ◽

1993 ◽

Vol 292 (1) ◽

pp. 105-111 ◽

Cited By ~ 15

Author(s):

L Dory

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Apolipoprotein E ◽

Phorbol Ester ◽

Mrna Levels ◽

Dependent Manner ◽

Kinase C ◽

Protein Kinase C Activity ◽

Mouse Macrophages ◽

Apoe Expression

Phorbol ester-mediated differentiation of THP-1 cells (a human monocytic cell line) into mature macrophages is associated with a transcriptional induction of apolipoprotein E (apoE) expression [Auwerx, Deeb, Brunzell, Peng and Chait (1988) Biochemistry 27, 2651-2655]. Endotoxin, on the other hand, which may also act through activation of protein kinase C, is a potent inhibitor of apoE expression in mouse macrophages [Werb and Chin (1983) J. Biol. Chem. 258, 10642-10648]. The present experiments examine the effect of phorbol ester, an activator of protein kinase C, on the apoE expression in mouse thioglycollate-elicited peritoneal macrophages. Phorbol ester inhibits apoE expression in a specific, time- and dose-dependent manner. A 75% inhibition in the rate of apoE secretion, but not that of total protein, was observed following a 4.5 h incubation with 160 nM phorbol ester, although nearly full inhibition was obtained with 40 nM. The changes in apoE secretion were paralleled by similar changes in apoE synthesis, indicating synthesis as the primary site of action. The decreased rates of apoE synthesis are shown not to be due to increased apoE degradation. The profound inhibition of apoE synthesis was not accompanied by significant changes in apoE mRNA levels at any concentration of phorbol ester (up to 16 microM), or length of treatment (up to 24 h), suggesting a post-transcriptional locus of regulation of apoE expression. Although the early changes in apoE synthesis correlate with increased microsomal protein kinase C activity, the suppression of apoE expression persists even during conditions of nearly complete (> 95%) loss of protein kinase C activity, suggesting that the direct or indirect effect of protein kinase C on apoE expression is mediated by a stable phosphorylated protein, or that the observed effects are mediated through a protein kinase C species that is not readily downregulated by phorbol esters. The presented studies clearly demonstrate the potential importance of the translational regulation of apoE expression through the protein kinase C signal transduction pathway.

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DIACYLGLYCEROL AND PHORBOL ESTER REDUCE AEQUORIN-INDICATED [Ca++] ELEVATIONS INDUCED BY THROMBIN OR ADP

10.1055/s-0038-1644503 ◽

1987 ◽

Author(s):

J A Ware ◽

M Smith ◽

E W Salzman

Keyword(s):

Protein Kinase C ◽

Platelet Aggregation ◽

Protein Kinase ◽

Phorbol Ester ◽

Time Dependent ◽

Dependent Manner ◽

Kinase C ◽

Protein Kinase C Activators ◽

Protein Kinase C Inhibitor ◽

Ca Homeostasis

Platelet aggregation and secretion induced by phorbol ester (PMA) or diacylglycerol (DAG) are preceded by an increase in [Ca++] that is detected byaequorin, but not by quin2, fura-2, or indo-1, suggesting that these indicatorsreflect different aspects of Ca++ homeostasis, possibly different functional Ca++ pools. Addition of two conventional agonists in subthreold concentrations synergistically enhances the [Ca++] rise and aggregation.However, if PMA or DAG is the first agonist the subsequent quin2-indicated [Ca++] rise after thrombin is reduced.Whether aequorin-indicated [Ca++] is similarly affected is unknown. We studied gel-filtered platelets loaded with aequorin or a fluorophore and added PMA, DAG, thrombin or ADP, alone or in combination. Either PMA or DAG alone caused a concentration-dependent increase in [Ca++] detectable with aequorin but not with the fluorophores; simultaneous addition of thrombin or ADP with DAG or PMA produced a larger [Ca++] rise than either alone. However, addition of DAG or PMA as a first agonist reduced subsequent aequorin-indicated [Ca++] rises following thrombin or ADP in a concentration and time-dependent manner. Inhibition of ADP or thrombin-induced [Ca++] rise was not always accompanied by inhibition of aggregation or secretion. Combination of subthreshold concentrations of ADP and thrombin produced an enhanced [Ca++] rise and aggregation. However, this synergistic effect was inhibited by preincubation with DAG or PMA. Neither this effect nor DAG-induced [Ca++] rise was inhibited by the protein kinase C inhibitor H-7. In genera^ preincubation of platelets with an agonist enhances Ca rise and aggregation in response to a second agonist; in contrasl protein kinase C activators, which themselves elevate [Ca++] as shown by aequorin, inhibit aequorin-indicated Ca rises after ADP or thrombin, and limit synergism between these two agonists.

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Role of phospholipase C and protein kinase C in vasoconstrictor-induced prostaglandin synthesis in cultured rat renal mesangial cells

Biochemical Journal ◽

10.1042/bj2340125 ◽

1986 ◽

Vol 234 (1) ◽

pp. 125-130 ◽

Cited By ~ 49

Author(s):

J Pfeilschifter ◽

A Kurtz ◽

C Bauer

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Phospholipase C ◽

Phorbol Ester ◽

Mesangial Cells ◽

Common Mechanism ◽

Dependent Manner ◽

Pge2 Synthesis ◽

Kinase C ◽

Angiotension Ii

It was the aim of the present study to find out if a common mechanism exists by which the vasoconstrictive hormones angiotension II, noradrenaline and 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine (AGEPC) increase prostaglandin E2 (PGE2) synthesis in cultures of rat renal mesangial cells. Angiotension II, noradrenaline and AGEPC stimulated PGE2 synthesis and uptake of 45Ca2+ in cultured mesangial cells. Both of these effects could be completely suppressed by the calcium channel blocker verapamil. Angiotensin II, noradrenaline and AGEPC caused a rapid breakdown of phosphatidylinositol 4,5-bisphosphate with a concomitant increase of 1,2-diacylglycerol and inositol trisphosphate, indicating an activation of phospholipase C by these hormones. Addition of verapamil had no effect on the hormone-induced stimulation of phospholipase C. The synthetic analogue of diacylglycerol, 1-oleoyl-2-acetylglycerol, and the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA), both of which are known to stimulate protein kinase C, enhanced PGE2 synthesis. Chelation of extracellular calcium with EDTA or addition of verapamil abolished the effect of 1-oleoyl-2-acetylglycerol and phorbol ester on PGE2 synthesis. 1-Oleoyl-2-acetylglycerol and phorbol ester increased the uptake of 45Ca2+ by the cells in a dose-dependent manner and this effect could be blocked by verapamil. The entirety of these data leads us to suggest that vasoconstrictor-evoked synthesis of PGE2 in rat mesangial cells is mediated by the subsequent activation of phospholipase C and protein kinase C. The activation of protein kinase C by diacylglycerol is likely to be involved in the increase of the calcium permeability of the plasma membrane which is a prerequisite for PGE2 synthesis induced by vasoconstrictive hormones.

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Phospholipase D Is Associated in a Phorbol Ester-Dependent Manner with Protein Kinase C-α and with a 220-kDa Protein Which Is Phosphorylated on Serine and Threonine

Biochemical and Biophysical Research Communications ◽

10.1006/bbrc.1998.8990 ◽

1998 ◽

Vol 248 (3) ◽

pp. 533-537 ◽

Cited By ~ 24

Author(s):

Do Sik Min ◽

John H. Exton

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Phospholipase D ◽

Phorbol Ester ◽

Dependent Manner ◽

Kinase C

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Endotoxin-Induced Tissue Factor in Human Monocytes is Dependent upon Protein Kinase C Activation

Thrombosis and Haemostasis ◽

10.1055/s-0038-1649673 ◽

1993 ◽

Vol 70 (05) ◽

pp. 800-806 ◽

Cited By ~ 29

Author(s):

C Ternisien ◽

M Ramani ◽

V Ollivier ◽

F Khechai ◽

T Vu ◽

...

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Tissue Factor ◽

Factor Ix ◽

Transcriptional Level ◽

Mrna Levels ◽

Dependent Manner ◽

Human Monocytes ◽

Kinase C ◽

Pkc Activation

SummaryTissue factor (TF) is a transmembrane receptor which, in association with factors VII and Vila, activates factor IX and X, thereby activating the coagulation protease cascades. In response to bacterial lipopolysaccharide (LPS) monocytes transcribe, synthesize and express TF on their surface. We investigated whether LPS-induced TF in human monocytes is mediated by protein kinase C (PKC) activation. The PKC agonists phorbol 12- myristate 13-acetate (PMA) and phorbol 12, 13 dibutyrate (PdBu) were both potent inducers of TF in human monocytes, whereas 4 alpha-12, 13 didecanoate (4 a-Pdd) had no such effect. Both LPS- and PMA-induced TF activity were inhibited, in a concentration dependent manner, by three different PKC inhibitors: H7, staurosporine and calphostin C. TF antigen determination confirmed that LPS-induced cell-surface TF protein levels decreased in parallel to TF functional activity under staurosporine treatment. Moreover, Northern blot analysis of total RNA from LPS- or PMA-stimulated monocytes showed a concentration-dependent decrease in TF mRNA levels in response to H7 and staurosporine. The decay rate of LPS-induced TF mRNA evaluated after the arrest of transcription by actinomycin D was not affected by the addition of staurosporine, suggesting that its inhibitory effect occurred at a transcriptional level. We conclude that LPS-induced production of TF and its mRNA by human monocytes are dependent on PKC activation.

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