Cell Cycle Phase Dependence of Estrogen and Epidermal Growth Factor (EGF) Receptor Expression in MCF-7 Cells: Implications in Antiestrogen and EGF Cell Responsiveness*

Treatment of A431 human epidermoid cells with epidermal growth factor (EGF; 20 nM) results in decreased proliferation. This is associated with blockage of the cells in the S and/or G2 phases of the cell cycle. We found that tyrphostin, a putative tyrosine kinase inhibitor, in the range of 50 to 100 microM, partially reversed the growth-inhibitory and cell cycle changes induced by EGF. By using high-pressure liquid chromatography with electrochemical detection, we found that tyrphostin was readily incorporated into A431 cells, reaching maximal levels within 1 h. Although tyrphostin (50 to 100 microM) had no effect on high-affinity binding of EGF to its receptor in A431 cells for up to 24 h, the compound partially inhibited EGF-stimulated EGF receptor tyrosine kinase activity. However, this effect was evident only after prolonged treatment of the cells (4 to 24 h) with the drug. When the peak intracellular concentration of tyrphostin occurred (1 h), no inhibition of tyrosine kinase activity was observed. After both 1 and 24 h, tyrphostin was a less effective inhibitor of tyrosine kinase activity than the potent tumor promoter 12-O-tetradecanoyl phorbol-13-acetate, which almost completely blocked EGF receptor autophosphorylation. On the basis of our data, we hypothesize that tyrphostin is not a competitive inhibitor of the EGF receptor tyrosine kinase in intact cells and that it functions by an indirect mechanism.

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Regulation of EGFR Endocytosis by CBL During Mitosis

Cells ◽

10.3390/cells7120257 ◽

2018 ◽

Vol 7 (12) ◽

pp. 257 ◽

Cited By ~ 6

Author(s):

Ping Wee ◽

Zhixiang Wang

Keyword(s):

Growth Factor ◽

Epidermal Growth Factor ◽

Cell Signaling ◽

Binding Site ◽

Egf Receptor ◽

Dominant Negative ◽

Low Doses ◽

Interphase Cells ◽

Epidermal Growth ◽

Mcf 7

The overactivation of epidermal growth factor (EGF) receptor (EGFR) is implicated in various cancers. Endocytosis plays an important role in EGFR-mediated cell signaling. We previously found that EGFR endocytosis during mitosis is mediated differently from interphase. While the regulation of EGFR endocytosis in interphase is well understood, little is known regarding the regulation of EGFR endocytosis during mitosis. Here, we found that contrary to interphase cells, mitotic EGFR endocytosis is more reliant on the activation of the E3 ligase CBL. By transfecting HeLa, MCF-7, and 293T cells with CBL siRNA or dominant-negative 70z-CBL, we found that at high EGF doses, CBL is required for EGFR endocytosis in mitotic cells, but not in interphase cells. In addition, the endocytosis of mutant EGFR Y1045F-YFP (mutation at the direct CBL binding site) is strongly delayed. The endocytosis of truncated EGFR Δ1044-YFP that does not bind to CBL is completely inhibited in mitosis. Moreover, EGF induces stronger ubiquitination of mitotic EGFR than interphase EGFR, and mitotic EGFR is trafficked to lysosomes for degradation. Furthermore, we showed that, different from interphase, low doses of EGF still stimulate EGFR endocytosis by non-clathrin mediated endocytosis (NCE) in mitosis. Contrary to interphase, CBL and the CBL-binding regions of EGFR are required for mitotic EGFR endocytosis at low doses. This is due to the mitotic ubiquitination of the EGFR even at low EGF doses. We conclude that mitotic EGFR endocytosis exclusively proceeds through CBL-mediated NCE.

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Epiregulin (EPR) Reduces Epidermal Growth Factor (EGF) Receptor Expression In Oral Squamous Cell Carcinoma (OSCC) Cell Lines

Oral Surgery Oral Medicine Oral Pathology and Oral Radiology ◽

10.1016/j.oooo.2014.07.331 ◽

2015 ◽

Vol 119 (3) ◽

pp. e177

Author(s):

Darren Chyi Hsiang Kong ◽

Kenneth Yew Choy Chew ◽

Eng Lai Tan ◽

Suan Phaik Khoo

Keyword(s):

Squamous Cell Carcinoma ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Oral Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Cell Lines ◽

Squamous Cell ◽

Egf Receptor ◽

Receptor Expression ◽

Epidermal Growth

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Quantitative analysis of the EGF receptor autocrine system reveals cryptic regulation of cell response by ligand capture

Journal of Cell Science ◽

10.1242/jcs.114.12.2301 ◽

2001 ◽

Vol 114 (12) ◽

pp. 2301-2313 ◽

Cited By ~ 3

Author(s):

Ann E. DeWitt ◽

Jian Ying Dong ◽

H. Steven Wiley ◽

Douglas A. Lauffenburger

Keyword(s):

Growth Factor ◽

Epidermal Growth Factor ◽

Egf Receptor ◽

Expression System ◽

Receptor Occupancy ◽

Receptor Activation ◽

Quantitative Model ◽

Receptor Expression ◽

Extracellular Medium ◽

Epidermal Growth

Autocrine signaling is important in normal tissue physiology as well as pathological conditions. It is difficult to analyze these systems, however, because they are both self-contained and recursive. To understand how parameters such as ligand production and receptor expression influence autocrine activity, we investigated a human epidermal growth factor/epidermal growth factor receptor (EGF/EGFR) loop engineered into mouse B82 fibroblasts. We varied the level of ligand production using the tet-off expression system and used metalloprotease inhibitors to modulate ligand release. Receptor expression was varied using antagonistic blocking antibodies. We compared autocrine ligand release with receptor activation using a microphysiometer-based assay and analyzed our data using a quantitative model of ligand release and receptor dynamics. We found that the activity of our autocrine system could be described in terms of a simple ratio between the rate of ligand production (VLT) and the rate of receptor production (VR). At a VLT/VR ratio of <0.3, essentially no ligand was found in the extracellular medium, but a significant number of cell receptors (30-40%) were occupied. As the VLT/VR ratio increased from 0.3 towards unity, receptor occupancy increased and significant amounts of ligand appeared in the medium. Above a VLT/VR ratio of 1.0, receptor occupancy approached saturation and most of the released ligand was lost into the medium. Analysis of human mammary epithelial cells showed that a VLT/VR ratio of <5×10−4was sufficient to evoke >20% of a maximal proliferative response. This demonstrates that natural autocrine systems can be active even when no ligand appears in the extracellular medium.

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Rapid uptake of tyrphostin into A431 human epidermoid cells is followed by delayed inhibition of epidermal growth factor (EGF)-stimulated EGF receptor tyrosine kinase activity

Molecular and Cellular Biology ◽

10.1128/mcb.11.5.2697-2703.1991 ◽

1991 ◽

Vol 11 (5) ◽

pp. 2697-2703

Author(s):

C A Faaland ◽

F H Mermelstein ◽

J Hayashi ◽

J D Laskin

Keyword(s):

Cell Cycle ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Tyrosine Kinase ◽

Receptor Tyrosine Kinase ◽

Kinase Activity ◽

Egf Receptor ◽

Tyrosine Kinase Activity ◽

A431 Cells ◽

Epidermal Growth

Treatment of A431 human epidermoid cells with epidermal growth factor (EGF; 20 nM) results in decreased proliferation. This is associated with blockage of the cells in the S and/or G2 phases of the cell cycle. We found that tyrphostin, a putative tyrosine kinase inhibitor, in the range of 50 to 100 microM, partially reversed the growth-inhibitory and cell cycle changes induced by EGF. By using high-pressure liquid chromatography with electrochemical detection, we found that tyrphostin was readily incorporated into A431 cells, reaching maximal levels within 1 h. Although tyrphostin (50 to 100 microM) had no effect on high-affinity binding of EGF to its receptor in A431 cells for up to 24 h, the compound partially inhibited EGF-stimulated EGF receptor tyrosine kinase activity. However, this effect was evident only after prolonged treatment of the cells (4 to 24 h) with the drug. When the peak intracellular concentration of tyrphostin occurred (1 h), no inhibition of tyrosine kinase activity was observed. After both 1 and 24 h, tyrphostin was a less effective inhibitor of tyrosine kinase activity than the potent tumor promoter 12-O-tetradecanoyl phorbol-13-acetate, which almost completely blocked EGF receptor autophosphorylation. On the basis of our data, we hypothesize that tyrphostin is not a competitive inhibitor of the EGF receptor tyrosine kinase in intact cells and that it functions by an indirect mechanism.

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Epidermal Growth Factor is a Motility Factor for Microglial CellsIn Vitro:Evidence for EGF Receptor Expression

European Journal of Neuroscience ◽

10.1111/j.1460-9568.1997.tb01526.x ◽

1997 ◽

Vol 9 (8) ◽

pp. 1690-1698 ◽

Cited By ~ 64

Author(s):

Christiané Nolte ◽

Frank Kirchhoff ◽

Helmut Kettenmann

Keyword(s):

Growth Factor ◽

Epidermal Growth Factor ◽

Egf Receptor ◽

Receptor Expression ◽

Motility Factor ◽

Epidermal Growth

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Argonaute-2 Expression Is Regulated by Epidermal Growth Factor Receptor and Mitogen-Activated Protein Kinase Signaling and Correlates with a Transformed Phenotype in Breast Cancer Cells

Endocrinology ◽

10.1210/en.2008-0984 ◽

2008 ◽

Vol 150 (1) ◽

pp. 14-23 ◽

Cited By ~ 97

Author(s):

Brian D. Adams ◽

Kevin P. Claffey ◽

Bruce A. White

Keyword(s):

Breast Cancer ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Egf Receptor ◽

Growth Factor Receptor ◽

Mapk Signaling ◽

Full Length ◽

Breast Cancer Cell Lines ◽

Epidermal Growth ◽

Mcf 7

Argonaute (Ago) 2 is the catalytic engine of mammalian RNA interference, but little is known concerning the regulation of Ago2 by cell-signaling pathways. In this study we show that expression of Ago2, but not Ago1, Ago3, or Ago4, is elevated in estrogen receptor (ER) α-negative (ERα−) vs. ERα-positive (ERα+) breast cancer cell lines, and in ERα− breast tumors. In MCF-7 cells the low level of Ago2 was found to be dependent upon active ERα/estrogen signaling. Interestingly, the high expression of Ago2 in ERα− cells was severely blunted by inhibition of the epidermal growth factor (EGF) receptor/MAPK signaling pathway, using either a pharmacological MAPK kinase inhibitor, U0126, or a small interfering RNA directed against EGF receptor. Half-life studies using cycloheximide indicated that EGF enhanced, whereas U0126 decreased, Ago2 protein stability. Furthermore, a proteosome inhibitor, MG132, blocked Ago2 protein turnover. The functional consequences of elevated Ago2 levels were examined by stable transfection of ERα+ MCF-7 cells with full-length and truncated forms of Ago2. The full-length Ago2 transfectants displayed enhanced proliferation, reduced cell-cell adhesion, and increased migratory ability, as shown by proliferation, homotypic aggregation, and wound healing assays, respectively. Overexpression of full-length Ago2, but not truncated forms of Ago2 or an empty vector control, reduced the levels of E-cadherin, β-catenin, and β-actin, as well as enhanced endogenous miR-206 activity. These data indicate that Ago2 is regulated at both the transcriptional and posttranslational level, and also implicate Ago2 and enhanced micro-RNA activity in the tumorigenic progression of breast cancer cell lines. Argonaute-2 is elevated in ERα- breast cancer cells due to epidermal growth factor receptor/MAPK signaling, and overexpression of this gene induces a more transformed phenotype in ERα+ MCF-7 cells.

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Studies of the effects of new progestins from the pentarane series on viability, epidermal growth factor receptor expression, and apoptosis of cultured HeLa and MCF-7 cells

Pharmaceutical Chemistry Journal ◽

10.1007/s11094-008-0102-1 ◽

2008 ◽

Vol 42 (5) ◽

pp. 258-261 ◽

Cited By ~ 1

Author(s):

A. V. Semeikin ◽

I. S. Levina ◽

L. E. Kulikova ◽

A. V. Kamernitskii ◽

V. B. Vetchinkina ◽

...

Keyword(s):

Epidermal Growth Factor Receptor ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Growth Factor Receptor ◽

Receptor Expression ◽

Epidermal Growth ◽

Mcf 7

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Bradykinin-induced growth inhibition of normal rat kidney (NRK) cells is paralleled by a decrease in epidermal-growth-factor receptor expression

Biochemical Journal ◽

10.1042/bj2980335 ◽

1994 ◽

Vol 298 (2) ◽

pp. 335-340 ◽

Cited By ~ 17

Author(s):

E J J Van Zoelen ◽

P H J Peters ◽

G B Afink ◽

S Van Genesen ◽

A D G De Roos ◽

...

Keyword(s):

Retinoic Acid ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Egf Receptor ◽

Transforming Growth Factor ◽

Rat Kidney ◽

Receptor Expression ◽

Binding Studies ◽

Normal Rat ◽

Epidermal Growth

Normal rat kidney fibroblasts, grown to density arrest in the presence of epidermal growth factor (EGF), can be induced to undergo phenotypic transformation by treatment with transforming growth factor beta or retinoic acid. Here we show that bradykinin blocks this growth-stimulus-induced loss of density-dependent growth arrest by a specific receptor-mediated mechanism. The effects of bradykinin are specific, and are not mimicked by other phosphoinositide-mobilizing agents such as prostaglandin F2 alpha. Northern-blot analysis and receptor-binding studies demonstrate that bradykinin also inhibits the retinoic acid-induced increase in EGF receptor levels in these cells. These studies provide additional evidence that EGF receptor levels modulate EGF-induced expression of the transformed phenotype in these cells.

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