scholarly journals Differential responsiveness of intestinal epithelial cells to 1,25-dihydroxyvitamin D3--role of protein kinase C

2001 ◽  
Vol 169 (1) ◽  
pp. 145-151 ◽  
Author(s):  
HJ Armbrecht ◽  
MA Boltz ◽  
TL Hodam ◽  
VB Kumar
Keyword(s):  
Vitamin D ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Cell Lines ◽  
Phorbol Esters ◽  
Mrna Levels ◽  
Intestinal Epithelial ◽  
Vitamin D Receptors ◽  
Kinase C ◽  
Crypt Cells

Non-transformed rat intestinal epithelial cell (IEC) lines were used to study the action of 1,25-dihydroxyvitamin D(3) (1,25(OH)2D) in the intestine. The capacity of 1,25(OH)2D to increase the expression of the cytochrome P450 component of the vitamin D 24-hydroxylase (CYP24) was determined in IEC-6 and IEC-18 cell lines. In IEC-6 cells, which are derived from crypt cells isolated from the whole small intestine, 1,25(OH)2D markedly increased expression of CYP24 protein and mRNA within 12 h. In contrast, in IEC-18 cells, which are derived from crypt cells from the ileum only, 1,25(OH)2D did not increase expression of CYP24 until 24-48 h. The maximal levels of CYP24 mRNA seen in the IEC-18 cells were only 31% of the maximal levels seen in the IEC-6 cells. In the presence of 1,25(OH)2D, phorbol esters rapidly increased CYP24 mRNA levels in IEC-18 cells from almost undetectable to levels seen in IEC-6 cells. Protein kinase inhibitors abolished the stimulation by 1,25(OH)2D and by phorbol esters in both cell lines. Stimulation of mRNA levels by phorbol esters required new protein synthesis but stimulation by 1,25(OH)2D did not. These studies demonstrated that the rapid action of 1,25(OH)2D in IEC-6 cells is related to the activation of protein kinase C, an event which is missing in the IEC-18 cells. This differential response to 1,25(OH)2D probably takes place at a post-receptor site, since the number of vitamin D receptors in each cell line was found to be similar.

scholarly journals Phosphorylation of p90 and p52 in response to phorbol-esters in Swiss/3T3 cells overexpressing protein kinase C-alpha.

1992 ◽  
Vol 3 (9) ◽  
pp. 1049-1056 ◽  
Author(s):  
H Eldar ◽  
E Livneh
Keyword(s):  
Protein Kinase C ◽  
Growth Factor ◽  
Protein Kinase ◽  
Cell Lines ◽  
Phorbol Ester ◽  
Phorbol Esters ◽  
Tumor Promotion ◽  
3T3 Cells ◽  
Kinase C ◽  
Pkc Alpha

Cell lines stably overexpressing protein kinase C (PKC)-alpha were previously described by us. These cell lines were generated by the introduction of the full length cDNA coding for PKC-alpha into Swiss/3T3 cells. Here we show that activation of PKC-alpha by phorbol-esters induced in these cells specific phosphorylation of two cellular proteins p90 and p52. Phosphorylation of p80 (MARCKS protein), previously identified as a substrate for PKC, was also enhanced. Phosphorylated p90 and p52 proteins were associated with particulate membrane-enriched fractions and were extractable with the use of nonionic detergents. Time course analysis of phorbol-ester induced phosphorylation of p90 and p52 revealed maximal stimulation of phosphorylation after 15-30 min. Phosphamino acid analysis showed that phosphorylation of p90 and p52 occurred mainly on serine residues. Phosphorylation of p52 was also on threonine residues. Whereas, phorbol ester activation induced phosphorylation of both p90 and p52, the mitogens platelet-derived growth factor (PDGF) and fibroblast growth factor (FGF) enhanced phosphorylation of p90, but not p52. Thus, our studies showed the involvement of PKC-alpha in the regulation of p90 and p52 phosphorylation and provided direct evidence for the role of PKC-alpha in cellular signaling by PDGF and FGF. Moreover, the fact that phosphorylation of p52 was specific to phorbol ester activation may suggest its involvement in tumor promotion. Characterization of p90 and p52 will enable us to reveal the phosphorylation cascade activated downstream to PKC-alpha and to determine their role in mitogenic signaling and tumor promotion.

scholarly journals Calcitonin Induces 25-Hydroxyvitamin D31α-Hydroxylase mRNA Expression via Protein Kinase C Pathway in LLC-PK1Cells

1999 ◽  
Vol 10 (12) ◽  
pp. 2474-2479
Author(s):  
NORIKO YOSHIDA ◽  
TADASHI YOSHIDA ◽  
AKIRA NAKAMURA ◽  
TOSHIAKI MONKAWA ◽  
MATSUHIKO HAYASHI ◽  
...  
Keyword(s):  
Vitamin D ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Mrna Expression ◽  
Protein Kinase A ◽  
Mrna Levels ◽  
Hydroxyvitamin D ◽  
Kinase C ◽  
25 Hydroxyvitamin D ◽  
Kinase A

Abstract. The biosynthesis of 1α, 25-dihydroxyvitamin D3from 25-hydroxyvitamin D3is catalyzed by 25-hydroxyvitamin D31α-hydroxylase (CYP27B1) in renal proximal tubules. It was recently demonstrated that LLC-PK1cells express CYP27B1 mRNA, which is regulated by intracellular cAMP but not vitamin D3. To clarify the effect of calcitonin on vitamin D3metabolismin vitro, LLC-PK1cells were incubated with hormonal factors, and expression of CYP27B1 mRNA was measured by quantitative reverse transcription-PCR. Calcitonin at 100 nmol/L significantly increased CYP27B1 mRNA expression by 24 h (271 ± 21% of control). Incubation with calcitonin over a range of 1 μmol/L to 1 pmol/L resulted in a concentration-dependent increase in CYP27B1 mRNA levels. It is known that the calcitonin receptor has dual intracellular signaling pathways, via protein kinases A and C. Both 500 μmol/L 8-bromo-cAMP, a protein kinase A activator, and 100 nmol/L phorbol 12-myristate 13-acetate, a protein kinase C activator, increased CYP27B1 mRNA levels at 24 h (207 ± 54 and 246 ± 58% of control, respectively). However, calcitonin-induced CYP27B1 mRNA expression was only inhibited by the protein kinase C inhibitors staurosporine and calphostin C. The protein kinase A inhibitors Rp-cAMPS at 10 and 100 μmol/L and H-89 at 10 μmol/L had no effect on the action of calcitonin, in spite of cAMP-activation by calcitonin. The present data suggest that calcitonin upregulates CYP27B1 mRNA expression via the protein kinase C pathway in LLC-PK1cells.

scholarly journals Molecular cloning of gene sequences regulated by tumor promoters and mitogens through protein kinase C.

1987 ◽  
Vol 7 (8) ◽  
pp. 2821-2829 ◽  
Author(s):  
M D Johnson ◽  
G M Housey ◽  
P T Kirschmeier ◽  
I B Weinstein
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Actinomycin D ◽  
Phorbol Esters ◽  
Tumor Promoter ◽  
Cdna Clones ◽  
Mrna Levels ◽  
Tumor Promoters ◽  
Kinase C

cDNA clones representing genes whose expression is modulated by treatment with mitogens and tumor promoters were isolated and characterized. TPA-S1 corresponds to an mRNA species whose abundance was increased markedly within 1 h of exposure to the tumor promoter 12-O-tetradecanoyl phorbol-13-acetate (TPA), and TPA-R1 represents an mRNA that was decreased in TPA-treated cells. The induction of TPA-S1 was blocked by actinomycin D but was not affected by cycloheximide, and it was specific for phorbol esters with tumor-promoting activity. The role of protein kinase C in the induction of TPA-S1 is supported by the following lines of evidence. (i) Agents that activated protein kinase C (TPA, platelet-derived growth factor, and diacylglycerol) also increased TPA-S1 mRNA levels. (ii) A potent PKC inhibitor blocked the induction of TPA-S1. (iii) Down-regulation of PKC activity, by treatment of cells with TPA for 24 h, resulted in a loss of responsiveness to TPA-S1 induction by subsequent TPA treatment. DNA sequence analysis of TPA-S1 predicts a cysteine-rich, secreted protein with a molecular weight of 22.6 X 10(3) that exhibits homology with sequences representing a protein with human erythroid-potentiating activity and protease inhibitory activity.

Phorbol esters, but not the hormonal form of vitamin D, induce changes in protein kinase C during differentiation of human histiocytic lymphoma cell line (U-937)

Life Sciences ◽  
1987 ◽  
Vol 40 (21) ◽  
pp. 2111-2117 ◽  
Author(s):  
G. Mezzetti ◽  
G.P. Bagnara ◽  
M.G. Monti ◽  
L. Pernecco Casolo ◽  
L. Bonsi ◽  
...  
Keyword(s):  
Vitamin D ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Cell Line ◽  
Phorbol Esters ◽  
Lymphoma Cell ◽  
Lymphoma Cell Line ◽  
Kinase C ◽  
Histiocytic Lymphoma

Quantitation of protein kinase C by immunoblot?expression in different cell lines and response to phorbol esters

1987 ◽  
Vol 130 (1) ◽  
pp. 111-117 ◽  
Author(s):  
S. Stabel ◽  
A. Rodriguez-Pena ◽  
S. Young ◽  
E. Rozengurt ◽  
P. J. Parker
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Cell Lines ◽  
Phorbol Esters ◽  
Kinase C

scholarly journals Posttranscriptional regulation of GAP-43 gene expression in PC12 cells through protein kinase C-dependent stabilization of the mRNA.

1993 ◽  
Vol 120 (5) ◽  
pp. 1263-1270 ◽  
Author(s):  
N I Perrone-Bizzozero ◽  
V V Cansino ◽  
D T Kohn
Keyword(s):  
Gene Expression ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Pc12 Cells ◽  
Phorbol Ester ◽  
Phorbol Esters ◽  
Calcium Ionophore ◽  
Mrna Levels ◽  
Mrna Stabilization ◽  
Kinase C

We have previously shown that nerve growth factor (NGF) selectively stabilizes the GAP-43 mRNA in PC12 cells. To study the cellular mechanisms for this post-transcriptional control and to determine the contribution of mRNA stability to GAP-43 gene expression, we examined the effects of several agents that affect PC12 cell differentiation on the level of induction and rate of degradation of the GAP-43 mRNA. The NGF-mediated increase in GAP-43 mRNA levels and neurite outgrowth was mimicked by the phorbol ester TPA, but not by dibutyryl cAMP or the calcium ionophore A12783. Downregulation of protein kinase C (PKC) by high doses of phorbol esters or selective PKC inhibitors prevented the induction of this mRNA by NGF, suggesting that NGF and TPA act through a common PKC-dependent pathway. In mRNA decay studies, phorbol esters caused a selective 6-fold increase in the half-life of the GAP-43 mRNA, which accounts for most of the induction of this mRNA by TPA. The phorbol ester-induced stabilization of GAP-43 mRNA was blocked by the protein kinase inhibitor polymyxin B and was partially inhibited by dexamethasone, an agent that blocks GAP-43 expression and neuronal differentiation in PC12 cells. In contrast, the rates of degradation and the levels of the GAP-43 mRNA in control and TPA-treated cells were not affected by cycloheximide treatment. Thus, changes in GAP-43 mRNA turnover do not appear to require continuous protein synthesis. In conclusion, these data suggest that PKC activity regulates the levels of the GAP-43 mRNA in PC12 cells through a novel, translation-independent mRNA stabilization mechanism.

Molecular cloning of gene sequences regulated by tumor promoters and mitogens through protein kinase C

1987 ◽  
Vol 7 (8) ◽  
pp. 2821-2829
Author(s):  
M D Johnson ◽  
G M Housey ◽  
P T Kirschmeier ◽  
I B Weinstein
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Actinomycin D ◽  
Phorbol Esters ◽  
Tumor Promoter ◽  
Cdna Clones ◽  
Mrna Levels ◽  
Tumor Promoters ◽  
Kinase C

cDNA clones representing genes whose expression is modulated by treatment with mitogens and tumor promoters were isolated and characterized. TPA-S1 corresponds to an mRNA species whose abundance was increased markedly within 1 h of exposure to the tumor promoter 12-O-tetradecanoyl phorbol-13-acetate (TPA), and TPA-R1 represents an mRNA that was decreased in TPA-treated cells. The induction of TPA-S1 was blocked by actinomycin D but was not affected by cycloheximide, and it was specific for phorbol esters with tumor-promoting activity. The role of protein kinase C in the induction of TPA-S1 is supported by the following lines of evidence. (i) Agents that activated protein kinase C (TPA, platelet-derived growth factor, and diacylglycerol) also increased TPA-S1 mRNA levels. (ii) A potent PKC inhibitor blocked the induction of TPA-S1. (iii) Down-regulation of PKC activity, by treatment of cells with TPA for 24 h, resulted in a loss of responsiveness to TPA-S1 induction by subsequent TPA treatment. DNA sequence analysis of TPA-S1 predicts a cysteine-rich, secreted protein with a molecular weight of 22.6 X 10(3) that exhibits homology with sequences representing a protein with human erythroid-potentiating activity and protease inhibitory activity.

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

1993 ◽  
Vol 70 (05) ◽  
pp. 800-806 ◽  
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.

scholarly journals Phosphorylation of the human vitamin D receptor by protein kinase C. Biochemical and functional evaluation of the serine 51 recognition site

1993 ◽  
Vol 268 (20) ◽  
pp. 15118-15126 ◽  
Author(s):  
J.C. Hsieh ◽  
P.W. Jurutka ◽  
S. Nakajima ◽  
M.A. Galligan ◽  
C.A. Haussler ◽  
...  
Keyword(s):  
Vitamin D ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Vitamin D Receptor ◽  
Recognition Site ◽  
Functional Evaluation ◽  
Kinase C
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