scholarly journals Epidermal-growth-factor-induced formation of inositol phosphates in human A431 cells. Differences from the effect of bradykinin

1988 ◽  
Vol 252 (3) ◽  
pp. 857-863 ◽  
Author(s):  
B C Tilly ◽  
P A van Paridon ◽  
I Verlaan ◽  
S W de Laat ◽  
W H Moolenaar
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Cell System ◽  
Ionophore A23187 ◽  
A431 Cells ◽  
Inositol Phosphate Metabolism ◽  
Absolute Requirement ◽  
Epidermal Growth

In human A431 epidermoid carcinoma cells, epidermal growth factor (EGF) rapidly stimulates the breakdown of inositol phospholipids and raises cytoplasmic free [Ca2+]. In this paper, we investigate the action of EGF on inositol phosphate metabolism, and we compare it with the previously described effects of bradykinin on the same cell system [Tilly, van Paridon, Verlaan, Wirtz, de Laat & Moolenaar (1987) Biochem. J. 244, 129-135]. In cells prelabelled with [3H]inositol, EGF slowly but persistently (for at least 30 min) stimulates the formation of [3H]inositol phosphates, whereas bradykinin causes an immediate but transient release of inositol phosphates, which lasts for only a few minutes. The EGF effect is additive to bradykinin stimulation and does not require extracellular Ca2+. In contrast, inositol phosphate formation induced by Ca2+-ionophore A23187 has an absolute requirement for external Ca2+. Treatment of the cells with 12-O-tetradecanoylphorbol 13-acetate completely abolishes the response to EGF and to sub-optimal doses of bradykinin, suggesting a negative-feedback function of protein kinase C. Pretreatment of the cells with pertussis toxin has no effect on inositol phosphate formation induced by either EGF or bradykinin. Unlike bradykinin, EGF stimulates very little accumulation of inositol 1,4,5-trisphosphate, with only a small and rather variable release of Ca2+ from intracellular stores. EGF rapidly but transiently increases inositol 1,3,4-trisphosphate and 1,3,4,5-tetrakisphosphate, but the effects are much smaller than those of bradykinin. In addition, EGF increases both inositol mono- and bis-phosphate. At 10 min after EGF addition, inositol monophosphate, unlike the other inositol phosphates, is still increasing. It is concluded that the EGF-dependent pattern of stimulation is different from that observed in bradykinin-stimulated A431 cells, suggesting that there are separate mechanisms of inositol-lipid hydrolysis involved.

scholarly journals c-fos sequence necessary for basal expression and induction by epidermal growth factor, 12-O-tetradecanoyl phorbol-13-acetate and the calcium ionophore.

1987 ◽  
Vol 7 (10) ◽  
pp. 3490-3502 ◽  
Author(s):  
T M Fisch ◽  
R Prywes ◽  
R G Roeder
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
General Expression ◽  
Base Pair ◽  
Calcium Ionophore ◽  
Ionophore A23187 ◽  
A431 Cells ◽  
Mobility Shift ◽  
Sequence Element ◽  
Epidermal Growth

We have investigated the sequence requirements for induction of the human c-fos gene by epidermal growth factor (EGF), 12-O-tetradecanoyl-13-acetate (TPA), and the calcium ionophore A23187 by transfecting c-fos promoter mutants into HeLa and A431 cells. Induction by both EGF and TPA in HeLa cells required the presence of the c-fos enhancer located at -317 to -298 relative to the mRNA cap site. A23187, however, did not induce expression of the transfected gene, even though it strongly induced expression of the endogenous gene, suggesting that it has different requirements for induction than do EGF and TPA. We have also investigated the role of promoter sequences downstream of the enhancer in general expression and induction of c-fos. A sequence between -97 and -76, which includes an 8-base-pair perfect direct repeat, was needed for efficient general expression but not for induction of the gene. A factor in nuclear extracts that bound specifically to this sequence was detected by a gel mobility shift assay. A 7-base-pair sequence, located between -63 and -57 relative to the mRNA cap site and previously shown to be important for general expression of mouse c-fos, was also important for general expression of the human gene. In addition, this element was important for inducibility by EGF and TPA, since induction was significantly reduced when internal deletion mutants that retained the enhancer but lacked the -63 to -57 sequence element were analyzed in transfecting assays.

Accumulation of inositol phosphates in low-passage human meningioma cells following treatment with epidermal growth factor

1994 ◽  
Vol 80 (5) ◽  
pp. 890-896 ◽  
Author(s):  
Tomoki Todo ◽  
Rudolf Fahlbusch
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Human Meningioma ◽  
Phosphate Accumulation ◽  
Phospholipid Hydrolysis ◽  
Inositol Phosphate Accumulation ◽  
Epidermal Growth ◽  
Inositol Phospholipid

✓ In order to elucidate some of the signal transduction processes in human meningioma cells, the authors studied the effect of epidermal growth factor (EGF) and bromocriptine on inositol phospholipid hydrolysis, using low-passage human meningioma cells in culture. Epidermal growth factor is a well-studied mitogenic factor for meningioma cells, whereas bromocriptine is known to have an inhibitory effect on meningioma cell proliferation. The addition of EGF to meningioma cells caused stimulation of inositol phosphate accumulation in a dose-dependent manner at 60 minutes posttreatment, with the maximum effect (120% to 167% of control) achieved at a concentration of 10 ng/ml. Extraction of separate inositol phosphates revealed that inositol monophosphate (IP1) and inositol bisphosphate (IP2), but not inositol trisphosphate (IP3), accounted for the increase at 60 minutes. Kinetic analysis of EGF-stimulated inositol phospholipid hydrolysis showed that a sharp and transient increase in IP3 from 5 to 12 minutes post-EGF and a transient but more gradual increase in IP2 from 2 to 12 minutes post-EGF were followed by a gradual and steady increase in IP1, which was significantly greater than control after 5 minutes. On the other hand, long-term studies showed a down-regulation of inositol phosphate accumulation (a 64% decrease vs. control) after 7 days of treatment with EGF (10 ng/ml). Bromocriptine (5 µM) exhibited no significant effect on inositol phosphate accumulation at 60 minutes in four of five meningiomas studied. However, of two meningiomas studied with bromocriptine in combination with EGF, both showed a significant additive increase in inositol phosphate accumulation compared to those treated with EGF alone. The results suggest a close involvement of inositol phospholipid turnover in human meningioma cells in response to mitogenic stimulation by EGF.

The phosphatidylinositol signalling system in elongating bovine blastocysts; formation of phosphoinositides, inositol phosphates and stimulation by growth factors

2002 ◽  
Vol 14 (8) ◽  
pp. 515 ◽  
Author(s):  
A. C. Hynes ◽  
J. M. Sreenan ◽  
M. T. Kane
Keyword(s):  
Signal Transduction ◽  
Growth Factors ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Signal Transduction System ◽  
Signalling System ◽  
Sodium Dependent ◽  
Epidermal Growth

The uptake of myo-inositol and its incorporation into the phosphoinositides and inositol phosphates of the phosphatidylinositol (PtdIns) signal transduction system by in vivo elongating cattle blastocysts was investigated using [3H]myo-inositol. Uptake was examined in 13-, 14- and 16-day-old blastocysts and was largely sodium-dependent throughout (P<0.001), indicating the presence of a sodium-dependent inositol transporter. Incorporation of inositol into the three phosphoinositides, PtdIns, PtdInsP and PtdInsP2, and the inositol phosphates of the phosphatidylinositol signal transduction system was examined at Days 14 and 16; incorporation into the three phosphoinositides and into the inositol phosphate species, InsP1, InsP2, InsP3 (including the second messenger, Ins(1,4,5)P3) and InsP4 was detected in both blastocyst stages. The effects of the peptide growth factor, epidermal growth factor (EGF), and the lipid growth factors, lysophosphatidic acid (LPA) and platelet activating factor (PAF), on the activity of the phosphatidylinositol signalling system in 14- and 16-day-old blastocysts were examined. All growth factors significantly stimulated phosphatidylinositol signalling activity. Epidermal growth factor was stimulatory (P<0.001) only in 16-day-old blastocysts, whereas LPA and PAF were active in both 14- (P<0.005 for LPA and P<0.001 for PAF) and 16-day-old blastocysts (P<0.001 for LPA and PAF). These results indicate that the phosphatidylinositol signalling system is present in cattle blastocysts at the elongation stage and is responsive to stimulation by growth factors.

c-fos sequence necessary for basal expression and induction by epidermal growth factor, 12-O-tetradecanoyl phorbol-13-acetate and the calcium ionophore

1987 ◽  
Vol 7 (10) ◽  
pp. 3490-3502
Author(s):  
T M Fisch ◽  
R Prywes ◽  
R G Roeder
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
General Expression ◽  
Base Pair ◽  
Calcium Ionophore ◽  
Ionophore A23187 ◽  
A431 Cells ◽  
Mobility Shift ◽  
Sequence Element ◽  
Epidermal Growth

We have investigated the sequence requirements for induction of the human c-fos gene by epidermal growth factor (EGF), 12-O-tetradecanoyl-13-acetate (TPA), and the calcium ionophore A23187 by transfecting c-fos promoter mutants into HeLa and A431 cells. Induction by both EGF and TPA in HeLa cells required the presence of the c-fos enhancer located at -317 to -298 relative to the mRNA cap site. A23187, however, did not induce expression of the transfected gene, even though it strongly induced expression of the endogenous gene, suggesting that it has different requirements for induction than do EGF and TPA. We have also investigated the role of promoter sequences downstream of the enhancer in general expression and induction of c-fos. A sequence between -97 and -76, which includes an 8-base-pair perfect direct repeat, was needed for efficient general expression but not for induction of the gene. A factor in nuclear extracts that bound specifically to this sequence was detected by a gel mobility shift assay. A 7-base-pair sequence, located between -63 and -57 relative to the mRNA cap site and previously shown to be important for general expression of mouse c-fos, was also important for general expression of the human gene. In addition, this element was important for inducibility by EGF and TPA, since induction was significantly reduced when internal deletion mutants that retained the enhancer but lacked the -63 to -57 sequence element were analyzed in transfecting assays.

Membrane vesicles of A431 cells contain one class of epidermal growth factor binding sites

1990 ◽  
Vol 1052 (3) ◽  
pp. 453-460 ◽  
Author(s):  
Jos A.M. Berkers ◽  
Paul M.P. van Bergen en Henegouwen ◽  
Arie J. Verkleij ◽  
Johannes Boonstra
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Binding Sites ◽  
Membrane Vesicles ◽  
A431 Cells ◽  
Factor Binding ◽  
Epidermal Growth
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