scholarly journals Participation of both Gab1 and Gab2 in the activation of the ERK/MAPK pathway by epidermal growth factor

2005 ◽  
Vol 391 (1) ◽  
pp. 143-151 ◽  
Author(s):  
Songshu Meng ◽  
Zhengming Chen ◽  
Teresita Munoz-Antonia ◽  
Jie Wu
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Mapk Pathway ◽  
Inhibitory Effect ◽  
Similar Amount ◽  
Small Interfering Rnas ◽  
Erk Activation ◽  
Activation Pathway ◽  
Docking Proteins ◽  
Epidermal Growth

Three members of Gab family docking proteins, Gab1, Gab2 and Gab3, have been identified in humans. Previous studies have found that the hepatocyte growth factor preferentially utilizes Gab1 for signalling, whereas Bcr-Abl selectively signals through Gab2. Gab1–SHP2 interaction has been shown to mediate ERK (extracellular-signal-regulated kinase) activation by EGF (epidermal growth factor). However, it was unclear whether EGF selectively utilizes Gab1 for signalling to ERK and whether Gab2 is dispensable in cells where Gab1 and Gab2 are co-expressed. Using T47D and MCF-7 human breast carcinoma cells that express endogenous Gab1 and Gab2, we examined the role of these docking proteins in EGF-induced ERK activation. It was found that EGF induced a similar amount of SHP2–Gab1 and SHP2–Gab2 complexes. Expression of either SHP2-binding defective Gab1 or Gab2 mutant blocked EGF-induced ERK activation. Down-regulation of either Gab1 or Gab2 by siRNAs (small interfering RNAs) effectively inhibited the EGF-stimulated ERK activation pathway and cell migration. Interestingly, the inhibitory effect of Gab1 siRNA could be rescued not only by expression of an exogenous mouse Gab1 but also by an exogenous human Gab2 and vice versa, but not by IRS1 (insulin receptor substrate 1). These results reveal that Gab2 plays a pivotal role in the EGF-induced ERK activation pathway and that it can complement the function of Gab1 in the EGF signalling pathway. Furthermore, Gab1 and Gab2 are critical signalling threshold proteins for ERK activation by EGF.

scholarly journals Differential Activation of Epidermal Growth Factor (EGF) Receptor Downstream Signaling Pathways by Betacellulin and EGF

Endocrinology ◽  
2004 ◽  
Vol 145 (9) ◽  
pp. 4232-4243 ◽  
Author(s):  
Tsugumichi Saito ◽  
Shuichi Okada ◽  
Kihachi Ohshima ◽  
Eijiro Yamada ◽  
Minoru Sato ◽  
...  
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Signaling Pathways ◽  
Tyrosine Phosphorylation ◽  
Egf Receptor ◽  
Chinese Hamster Ovary Cells ◽  
Inhibitory Effect ◽  
Erk Activation ◽  
Downstream Signaling ◽  
Epidermal Growth

Abstract To determine the downstream signaling pathways regulated by betacellulin (BTC) in comparison with epidermal growth factor (EGF), we used Chinese hamster ovary cells overexpressing the human EGF receptor (ErbB1/EGFR). The overall time-dependent activation of EGFR autophosphorylation was identical in cells treated with 1 nm BTC or 1.5 nm EGF. Analysis of site-specific EGFR phosphorylation demonstrated that the BTC and EGF tyrosine phosphorylation of Y1086 was not significantly different. In contrast, the autophosphorylation of Y1173 was markedly reduced in BTC-stimulated cells, compared with EGF stimulation that directly correlated with a reduced BTC stimulation of Shc tyrosine phosphorylation, Ras, and Raf-1 activation. On the other hand, Y1068 phosphorylation was significantly increased after BTC stimulation, compared with EGF in parallel with a greater extent of Erk phosphorylation. Expression of a dominant interfering MEK kinase 1 (MEKK1) and Y1068F EGFR more efficiently blocked the enhanced Erk activation by BTC, compared with EGF. Interestingly BTC had a greater inhibitory effect on apoptosis, compared with EGF, and expression of Y1068F EGFR abolished this enhanced inhibitory effect. Together, these data indicated that although BTC and EGF share overlapping signaling properties, the ability of BTC to enhance Erk activation occurs independent of Ras. The increased BTC activation results from a greater extent of Y1068 EGFR tyrosine phosphorylation and subsequent increased recruitment of the Grb2-MEKK1 complex to the plasma membrane, compared with EGF stimulation. The increased Erk activation by BTC associated with antiapoptotic function.

The calcium signal for BALB/MK keratinocyte terminal differentiation counteracts epidermal growth factor (EGF) very early in the EGF-induced proliferative pathway

1988 ◽  
Vol 8 (2) ◽  
pp. 557-563
Author(s):  
P P Di Fiore ◽  
J Falco ◽  
I Borrello ◽  
B Weissman ◽  
S A Aaronson
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Dna Synthesis ◽  
Fold Increase ◽  
Inhibitory Effect ◽  
Lymphoid Cells ◽  
Epidermal Keratinocytes ◽  
Calcium Signal ◽  
High Calcium ◽  
Epidermal Growth

BALB/MK mouse epidermal keratinocytes require epidermal growth factor (EGF) for proliferation and terminally differentiate in response to high calcium concentrations. We show that EGF is an extremely potent mitogen, causing BALB/MK cultures to enter the cell cycle in a synchronous manner associated with a greater than 100-fold increase in DNA synthesis. Analysis of the expression of proto-oncogenes which have been reported to be activated during the cascade of events following growth factor stimulation of fibroblasts or lymphoid cells revealed a very rapid but transient 100-fold increase in c-fos RNA but little or no effect on the other proto-oncogenes analyzed. Exposure of EGF-synchronized BALB/MK cells to high levels of calcium was associated with a striking decrease in the early burst of c-fos RNA as well as the subsequent peak of cell DNA synthesis. Since the inhibitory effect of high calcium on c-fos RNA expression was measurable within 30 min, our studies imply that the EGF proliferative and calcium differentiation signals must interact very early in the pathway of EGF-induced proliferation. Our results also establish that c-fos RNA modulation is an important early marker of cell proliferation in epithelial as well as mesenchymal cells.

scholarly journals Epidermal growth factor inhibits growth of A431 human epidermoid carcinoma in serum-free cell culture.

1982 ◽  
Vol 93 (1) ◽  
pp. 1-4 ◽  
Author(s):  
D W Barnes
Keyword(s):  
Growth Factor ◽  
Cell Growth ◽  
Epidermal Growth Factor ◽  
Culture Medium ◽  
Cell Types ◽  
Inhibitory Effect ◽  
Free Cell ◽  
Bovine Insulin ◽  
A431 Cell ◽  
Epidermal Growth

A medium consisting of a rich basal nutrient mixture supplemented with bovine insulin (10 micrograms/ml), human transferrin (10 micrograms/ml), human cold-insoluble globulin (5 micrograms/ml), and ethanolamine (0.5 mM) supported the growth of the A431 human epidermoid cell line in the absence of serum with a generation time equal to that of cells in serum-containing medium. Addition of epidermal growth factor (EGF) to this culture medium at concentration mitogenic for other cell types resulted in a marked inhibition of A431 cell growth. Inhibitory effects of EGF were observed at 1 ng/ml and near-maximal effects were observed at 10 ng/ml. The inhibitory effect of EGF could be reversed by the omission of EGF in subsequent medium changes and could be prevented by the addition of anti-EGF antibody to the culture medium. Inhibition of A431 cell growth by EGF also could be demonstrated in serum-containing medium.

scholarly journals The calcium signal for BALB/MK keratinocyte terminal differentiation counteracts epidermal growth factor (EGF) very early in the EGF-induced proliferative pathway.

1988 ◽  
Vol 8 (2) ◽  
pp. 557-563 ◽  
Author(s):  
P P Di Fiore ◽  
J Falco ◽  
I Borrello ◽  
B Weissman ◽  
S A Aaronson
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Dna Synthesis ◽  
Fold Increase ◽  
Inhibitory Effect ◽  
Lymphoid Cells ◽  
Epidermal Keratinocytes ◽  
Calcium Signal ◽  
High Calcium ◽  
Epidermal Growth

BALB/MK mouse epidermal keratinocytes require epidermal growth factor (EGF) for proliferation and terminally differentiate in response to high calcium concentrations. We show that EGF is an extremely potent mitogen, causing BALB/MK cultures to enter the cell cycle in a synchronous manner associated with a greater than 100-fold increase in DNA synthesis. Analysis of the expression of proto-oncogenes which have been reported to be activated during the cascade of events following growth factor stimulation of fibroblasts or lymphoid cells revealed a very rapid but transient 100-fold increase in c-fos RNA but little or no effect on the other proto-oncogenes analyzed. Exposure of EGF-synchronized BALB/MK cells to high levels of calcium was associated with a striking decrease in the early burst of c-fos RNA as well as the subsequent peak of cell DNA synthesis. Since the inhibitory effect of high calcium on c-fos RNA expression was measurable within 30 min, our studies imply that the EGF proliferative and calcium differentiation signals must interact very early in the pathway of EGF-induced proliferation. Our results also establish that c-fos RNA modulation is an important early marker of cell proliferation in epithelial as well as mesenchymal cells.

scholarly journals The Inhibitory Effect of ErbB2 on Epidermal Growth Factor-induced Formation of Clathrin-coated Pits Correlates with Retention of Epidermal Growth Factor Receptor-ErbB2 Oligomeric Complexes at the Plasma Membrane

2005 ◽  
Vol 16 (12) ◽  
pp. 5832-5842 ◽  
Author(s):  
Camilla Haslekås ◽  
Kamilla Breen ◽  
Ketil W. Pedersen ◽  
Lene E. Johannessen ◽  
Espen Stang ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Plasma Membrane ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Egf Receptor ◽  
Growth Factor Receptor ◽  
Inhibitory Effect ◽  
Coated Pits ◽  
Transfected Cells ◽  
Epidermal Growth

By constructing stably transfected cells harboring the same amount of epidermal growth factor (EGF) receptor (EGFR), but with increasing overexpression of ErbB2, we have demonstrated that ErbB2 efficiently inhibits internalization of ligand-bound EGFR. Apparently, ErbB2 inhibits internalization of EGF-bound EGFR by constitutively driving EGFR-ErbB2 hetero/oligomerization. We have demonstrated that ErbB2 does not inhibit phosphorylation or ubiquitination of the EGFR. Our data further indicate that the endocytosis deficiency of ErbB2 and of EGFR-ErbB2 heterodimers/oligomers cannot be explained by anchoring of ErbB2 to PDZ-containing proteins such as Erbin. Instead, we demonstrate that in contrast to EGFR homodimers, which are capable of inducing new clathrin-coated pits in serum-starved cells upon incubation with EGF, clathrin-coated pits are not induced upon activation of EGFR-ErbB2 heterodimers/oligomers.

scholarly journals RNA Interference Screen Identifies Usp18 as a Regulator of Epidermal Growth Factor Receptor Synthesis

2009 ◽  
Vol 20 (6) ◽  
pp. 1833-1844 ◽  
Author(s):  
Jason E. Duex ◽  
Alexander Sorkin
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Tyrosine Kinases ◽  
Growth Factor Receptor ◽  
Mrna Translation ◽  
Small Interfering Rnas ◽  
Catalytic Cysteine ◽  
Elevated Expression ◽  
Epidermal Growth

Elevated expression of epidermal growth factor receptor (EGFR) contributes to the progression of many types of cancer. Therefore, we developed a high-throughput screen to identify proteins that regulate the levels of EGFR in squamous cell carcinoma. Knocking down various ubiquitination-related genes with small interfering RNAs led to the identification of several novel genes involved in this process. One of these genes, Usp18, is a member of the ubiquitin-specific protease family. We found that knockdown of Usp18 in several cell lines reduced expression levels of EGFR by 50–80%, whereas the levels of other receptor tyrosine kinases remained unchanged. Overexpression of Usp18 elevated EGFR levels in a manner requiring the catalytic cysteine of Usp18. Analysis of metabolically radiolabeled cells showed that the rate of EGFR protein synthesis was reduced up to fourfold in the absence of Usp18. Interestingly, this dramatic reduction occurred despite no change in the levels of EGFR mRNA. This suggests that depletion of Usp18 inhibited EGFR mRNA translation. In fact, this inhibition required the presence of native 5′ and 3′ untranslated region sequences on EGFR mRNA. Together, our data provide evidence for the novel mechanism of EGFR regulation at the translational step of receptor synthesis.

scholarly journals DNA synthesis by ovine mammary alveolar epithelial cells: effects of heparin, epidermal growth factor-related peptides and interaction with stage of pregnancy

1998 ◽  
Vol 156 (2) ◽  
pp. 283-290 ◽  
Author(s):  
IA Forsyth ◽  
JA Taylor ◽  
CD Moorby
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Epithelial Cells ◽  
Dna Synthesis ◽  
Cell Cultures ◽  
Inhibitory Effect ◽  
Alveolar Epithelial Cells ◽  
Alveolar Epithelial ◽  
Pregnant Sheep ◽  
Epidermal Growth

Amphiregulin is a heparin-binding member of the epidermal growth factor (EGF) family, which we have recently shown to be expressed in sheep mammary gland. Uniquely among known EGF-like growth factors, its mitogenic activity is inhibited by soluble heparin, but heparin-like molecules on the cell surface and/or in extracellular matrix appear to be necessary for amphiregulin to exert its biological effect. In primary cultures of sheep mammary alveolar epithelial cells, heparin (1-20 mg/l) inhibited DNA synthesis in a dose-dependent manner. The extent of the inhibition was influenced by physiological state, being greater (P < 0.05) in mammary cell cultures derived from 5- to 10-week pregnant sheep (63.1 +/- 8.2%, mean +/- S.E.M., n = 8) than in cultures derived from sheep which were non-pregnant (35.8 +/- 8.3% inhibition, n = 6) or late, 20-week, pregnant (39.8 +/- 5.6%, n = 6). Both EGF and transforming growth factor-alpha (TGF-alpha) significantly (P < 0.001) increased DNA synthesis in the presence of heparin. The effect of TGF-alpha was dose-related, wholly reversing the inhibitory effect of heparin in cell cultures from non-pregnant and 20-week pregnant sheep. DNA synthesis was stimulated by amphiregulin and TGF-alpha increased the maximum response. The heparin antagonist, hexadimethrine, inhibited DNA synthesis, but, in the presence of amphiregulin, approximately equivalent concentrations of heparin overcame this inhibitory effect. In the presence of heparin, TGF-alpha showed synergistic interactions with insulin or IGF-I. The results indicate interactive effects of EGF and IGF growth factor families in sheep mammary growth.

Sign in / Sign up

Export Citation Format

Share Document