Induction of the 47 kDa platelet substrate of protein kinase C during differentiation of HL-60 cells

Immunoblot analysis showed that the 47 kDa platelet substrate of protein kinase C (P47) was expressed at low levels in undifferentiated HL-60 leukaemia cells. Treatment of these cells with dimethyl sulphoxide, 1 alpha,25-dihydroxycholecalciferol or retinoic acid caused progressive increases in P47 content. Retinoic acid (1 microM) elicited the largest response, a 4-fold increase in P47 protein after 7 days that was accompanied by an increase in translatable P47 mRNA. The induction of P47 by retinoic acid preceded cessation of cell proliferation and development of the capacity to reduce Nitro Blue Tetrazolium, indicating that its expression is an early event in the myeloid differentiation of HL-60 cells.

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Protein kinase C-independent stimulation of activator protein-1 and c-Jun N-terminal kinase activity in human endometrial cancer cells by the LHRH agonist triptorelin

Acta Endocrinologica ◽

10.1530/eje.0.1450651 ◽

2001 ◽

pp. 651-658 ◽

Cited By ~ 21

Author(s):

C Grundker ◽

L Schlotawa ◽

V Viereck ◽

G Emons

Keyword(s):

Cell Proliferation ◽

Endometrial Cancer ◽

Protein Kinase C ◽

Protein Kinase ◽

Cancer Cells ◽

Fold Increase ◽

Lhrh Agonist ◽

Activator Protein 1 ◽

Kinase C ◽

Lhrh Receptor

OBJECTIVE: The expression of luteinizing hormone-releasing hormone (LHRH) and its receptor as a part of an autocrine regulatory system of cell proliferation has been demonstrated in a number of human malignant tumours, including cancers of the endometrium. The signalling pathway through which LHRH acts in endometrial cancer is distinct from that in pituitary gonadotrophs. The LHRH receptor interacts with the mitogenic signal transduction of growth factor receptors via activation of a phosphotyrosine phosphatase, resulting in down-regulation of cancer cell proliferation. In addition, LHRH activates nucleus factor kappaB (NFkappaB) and protects the cancer cells from apoptosis. This study was conducted to investigate additional signalling mechanisms of the LHRH receptor cooperating with NFkappaB in endometrial cancer cells. DESIGN: The LHRH agonist triptorelin-induced activator protein-1 (AP-1) activation was analysed using a pAP-1-SEAP reporter gene assay. Expression of c-jun mRNA was quantified using quantitative reverse transcription (RT)-PCR. c-Jun N-terminal kinase (JNK) activity was measured by quantification of phosphorylated c-Jun protein. RESULTS: Treatment of Ishikawa and Hec-1A human endometrial cancer cells with 100 nM triptorelin resulted in a 3.1-fold and 3.5-fold activation of AP-1 respectively (P<0.05). If the cells had been made quiescent, treatment with triptorelin (100 nM) resulted in a 41.7-fold and 48.6-fold increase of AP-1 activation respectively (P<0.001). This effect was completely blocked by simultaneous treatment with pertussis toxin (PTX). A 17.6-fold and 17.3-fold increase of c-jun mRNA expression respectively (P<0.001) was obtained after 20 min of stimulation with triptorelin (100 nM). Treatment with 1 nM triptorelin resulted in a 12.5-fold or an 11.9-fold increase, and treatment with 10 pM triptorelin resulted in a 6.5-fold or a 5.2-fold increase of maximal c-jun mRNA expression respectively (P<0.001). Maximal c-Jun phosphorylation (68.5-fold and 60.2-fold, respectively, P<0.001) was obtained after 90 min incubation with triptorelin (100 nM). CONCLUSIONS: These results suggest that the LHRH agonist triptorelin stimulates the activity of AP-1 in human endometrial cancer cells mediated through PTX-sensitive G-protein alphai. In addition, triptorelin activates JNK, known to activate AP-1. In earlier investigations we have shown that triptorelin does not activate phospholipase and protein kinase C (PKC) in endometrial cancer cells. In addition, it has been demonstrated that triptorelin inhibits growth factor-induced mitogen activated protein kinase (MAPK, ERK) activity. Thus triptorelin-induced activation of the JNK/AP-1 pathway in endometrial cancer cells is independent of the known AP-1 activators, PKC or MAPK (ERK).

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Diacylglycerol generated during sphingomyelin synthesis is involved in protein kinase C activation and cell proliferation in Madin-Darby canine kidney cells

Biochemical Journal ◽

10.1042/bj20021732 ◽

2003 ◽

Vol 373 (3) ◽

pp. 917-924 ◽

Cited By ~ 20

Author(s):

Jorge CERBÓN ◽

Rosa del Carmen LÓPEZ-SÁNCHEZ

Keyword(s):

Cell Proliferation ◽

Protein Kinase C ◽

Protein Kinase ◽

Fold Increase ◽

Kidney Cells ◽

Madin Darby Canine Kidney ◽

Kinase C ◽

Pkc Activation ◽

Darby Canine Kidney ◽

Canine Kidney

We have investigated the effects of inhibiting sphingomyelin (SM) biosynthesis on cellular diacylglycerol (DAG) content and protein kinase C (PKC) activation during growth initiation in Madin–Darby canine kidney cells. We utilized β-chloroalanine (BCA) to inactivate serine C-palmitoyltransferase, the first enzyme in the sphingolipid biosynthesis pathway. This inactivation prevented growth, but did not affect viability. When the inhibitor was replaced with fresh culture medium, the cells continued their proliferation in a normal way. BCA (2 mM) inhibited [32P]Pi, [3H]palmitic acid and [methyl-3H]choline incorporation into SM, but did not influence the synthesis of other major phospholipids. SM synthesis and DAG generation were decreased by 51% and 47.6% respectively. Particulate PKC activity was not observed in cells incubated with BCA, in contrast with a 5-fold increase in control cells. BCA inhibited 75% of the [3H]thymidine incorporation, and the cells were arrested before the S phase of the cell cycle. Moreover, exogenous d-erythrosphingosine restored SM synthesis, DAG generation and cell proliferation. These data indicate that the contribution of DAG generated during SM synthesis plays an important role in PKC activation and cell proliferation.

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