API

Relationship: 964

Title

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Activation, EGFR leads to Activation, Sp1

Upstream event

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Activation, EGFR

Downstream event

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Activation, Sp1

Key Event Relationship Overview

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AOPs Referencing Relationship

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AOP Name Adjacency Weight of Evidence Quantitative Understanding
EGFR Activation Leading to Decreased Lung Function adjacent High Low

Taxonomic Applicability

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Term Scientific Term Evidence Link
human Homo sapiens High NCBI
mouse Mus musculus Moderate NCBI
rat Rattus norvegicus Moderate NCBI
dog Canis lupus familiaris Moderate NCBI

Sex Applicability

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Sex Evidence
Mixed Low

Life Stage Applicability

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Term Evidence
Adult Low

Key Event Relationship Description

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The EGF receptor family comprises 4 members, EGFR (also referred to as ErbB1/HER1), ErbB2/Neu/HER2, ErbB3/HER3 and ErbB4/HER4, all of which are transmembrane glycoproteins with an extracellular ligand binding site and an intracellular tyrosine kinase domain. Receptor-ligand binding induces dimerization and internalization, subsequently leading to activation of the receptor through autophosphorylation (Higashiyama et al., 2008). Classical EGFR downstream signaling involves activation of Ras which subsequently initiates signal transduction through the Raf-1/MEK/ERK pathway. MAP kinase activation in turn promotes airway epithelial cell proliferation and differentiation (Lemjabbar et al., 2003; Kim et al., 2005; Hackel et al, 1999) and facilitates epithelial wound repair (Burgel, 2004; van Winkle et al., 1997; Allahverdian et al., 2010). EGFR signal transduction via the MAPK cascade also activates the  transcription factor Sp1 (Di et al., 2012; Hewson et al., 2004; Lee et al., 2011; Perrais et al., 2002; Barbier et al., 2012; Oyanagi et al, 2016).

Evidence Supporting this KER

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Increased Sp1 activity following EGFR activation is well-documented. EGFR ligands EGF and TGFA or TGFA in combination with polyI:C increase Sp1 activity in NCI-H292 cells, a human pulmonary mucoepidermoid carcinoma cell line, by directly activating EGFR (Perrais et al., 2002; Oyanagi et al., 2016; Song et al., 2017). Sp1 activation through EGFR/MAPK activation was also reported in human airway epithelial cells following stimulation with PMA (Hewson et al. 2004) and in a mouse influenza model (Barbier et al., 2012). In addition, treatment of A549 cells with cigarette smoke extract increased Sp1 expression and nuclear translocation, resulting in enhanced Sp1-DNA binding and promoter transactivation, which could be suppressed by pretreatment with the EGFR inhibitor AG1478 (Di et al., 2012).

Other studies demonstrate EGFR-mediated activation of Sp1 in rat GH4 pituitary tumor cells (Merchant et al., 1995), Madin-Darby canine kidney (MDCK) cells (Ikari et al., 2009), human HepG2 hepatocellular carcinoma cells (Zheng et al., 2001), and human esophageal carcinoma cell lines (Lu et al., 2010).

Biological Plausibility

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Signal transduction via the MAPK cascade subsequent to EGFR activation results in Sp1 activation, and this is well-documented (Di et al., 2012; Hewson et al., 2004; Lee et al., 2011; Perrais et al., 2002; Barbier et al., 2012; Oyanagi et al, 2016).

Empirical Evidence

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There is weak empirical support for EGFR activation of Sp1, because few studies demonstrate the KER directly. In the majority of cases (in the context of this AOP), EGFR activation is shown to result in increased mucin expression, which is associated with the activation (as evidenced by increased expression and by phosphorylation) of Sp1 and/or repressed by the addition of an EGFR (e.g. AG1478) or Sp1 inhibitor (e.g. mithramycin A) (Di et al., 2012; Hewson et al., 2004; Lee et al., 2011; Perrais et al., 2002; Barbier et al., 2012; Oyanagi et al, 2016).

 

Uncertainties and Inconsistencies

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The described mechanism for this KER may specifically apply in the context of increased MUC5AC gene and protein expression, but not to that of MUC5B, another gel-forming mucin associated with mucus hypersecretion in the airways (Wu et al, 2007).

Quantitative Understanding of the Linkage

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20 ng/mL EGF or TGFA induced phosphorylation of Sp1, corresponding to a 2-3-fold increase in MUC2 and MUC5AC promoter activity, after 24 hours which was inhibited by 100 nM mithramycin A (a Sp1 inhibitor) (Perrais et al., 2002).

H292 cells infected for 24 hours with IAV at MOI=1 exhibited Sp1 activation as evidenced by a 1.5- to 5-fold increase in band shift. This was accompanied by an approx. 3-fold increase in EGFR phosphorylation at 24 hours (Barbier et al., 2012).

In human primary bronchial epithelial cells treated with 10 nM TCDD, Sp1 phosphorylation and MUC5AC promoter activity increased by ca. 2-fold and 4-fold, respectively, and increased promoter activity was abrogated in the presence of AG1478 (Lee et al., 2011).

In A549 cells treated with 3% cigarette smoke extract for 3 hours, Sp1 expression increased approx. 2.5-fold in the nuclear fraction, and this correlated with a significant increase in Sp1-DNA complex formation. Pretreatment of cells with the EGFR inhibitor AG1478 decreased Sp1-DNA binding (Di et al., 2012).

Response-response Relationship

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Time-scale

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Infection of H292 cells with influenza A virus (IVA) at MOI=1 resulted in increased EGFR phosphorylation, peaking at 24 hours. This was accompanied by activation of Sp-1 as shown by EMSA (Barbier et al., 2012). 

Treatment of primary bronchial epithelial cells with 10 nM TCDD resulted in maximal EGFR phosphorylation after 30 minutes. TCDD treatment also led to a time-dependent increase in MUC5AC transcriptional promoter activity, peaking between 6 and 12 hours. Sp1 involvement was demonstrated by treatment with mithramycin A (Lee et al., 2011).

20 ng/mL EGF or TGFA induced phosphorylation of Sp1, corresponding to a 2-3-fold increase in MUC2 and MUC5AC promoter activity, after 24 hours which was inhibited by 100 nM mithramycin A (a Sp1 inhibitor) (Perrais et al., 2002).

Treatment of H292 cells with a combination of 4 ng/mL TGFA and 25 microg/mL polyI:C resulted in a ca. 3-fold increase in EGFR phosphorylation at 1 hour. At 12 hours, MUC5AC mRNA expression was induced, intracellular MUC5AC protein expression was increased by nearly 30% and secretion of MUC5AC into the cell culture medium rose approx. 4-fold. MUC5AC mRNA expression could be completely abolished by the Sp1 inhibitor mithramycin A (500 nM) (Oyanagi et al., 2016).

 

Known modulating factors

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Known Feedforward/Feedback loops influencing this KER

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Domain of Applicability

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EGFR-mediated activation of Sp1 was reported in mouse (Hammoud et al., 2009; Lee et al., 2010), rat (Merchant et al., 1995; Mortensen et al., 1997), dog (Ikari et al., 2009; Ford et al., 1997) and human (Di et al., 2012; Hewson et al., 2004; Lee et al., 2011; Perrais et al., 2002; Barbier et al., 2012; Oyanagi et al, 2016).

References

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Allahverdian, S., Wang, A., Singhera, G.K., Wong, B.W., and Dorscheid, D.R. (2010). Sialyl Lewis X modification of the epidermal growth factor receptor regulates receptor function during airway epithelial wound repair. Clin Exp Allergy 40, 607-618.

Barbier, D., Garcia-Verdugo, I., Pothlichet, J., Khazen, R., Descamps, D., Rousseau, K., Thornton, D., Si-Tahar, M., Touqui, L., Chignard, M., et al. (2012). Influenza A Induces the Major Secreted Airway Mucin MUC5AC in a Protease–EGFR–Extracellular Regulated Kinase–Sp1–Dependent Pathway. Am J. Respir Cell Mol Biol 47, 149–157.

Burgel, P., and Nadel, J. (2004). Roles of epidermal growth factor receptor activation in epithelial cell repair and mucin production in airway epithelium. Thorax 59, 992-996.

Di, Y.P., Zhao, J., and Harper, R. (2012). Cigarette smoke induces MUC5AC protein expression through the activation of Sp1. J Biol Chem 287, 27948-27958.

Ford, M.G., Valle, J.D., Soroka, C.J., and Merchant, J.L. (1997). EGF receptor activation stimulates endogenous gastrin gene expression in canine G cells and human gastric cell cultures. J Clin Invest 99, 2762–2771.

Hackel, P.O., Zwick, E., Prenzel, N., and Ullrich, A. (1999). Epidermal growth factor receptors: critical mediators of multiple receptor pathways. Curr Opin Cell Biol 11, 184-189.

Higashiyama, S., Iwabuki, H., Morimoto, C., Hieda, M., Inoue, H., and Matsushita, N. (2008). Membrane-anchored growth factors, the epidermal growth factor family: Beyond receptor ligands. Cancer Sci 99, 214-220.

Hammoud, L., Burger, D.E., Lu, X., and Feng, Q. (2009). Tissue inhibitor of metalloproteinase-3 inhibits neonatal mouse cardiomyocyte proliferation via EGFR/JNK/SP-1 signaling. Am J Physiol Cell Physiol 296, C735–C745.

Hewson, C., Edbrooke, M., and Johnston, S. (2004). PMA induces the MUC5AC respiratory mucin in human bronchial epithelial cells, via PKC, EGF/TGF-alpha, Ras/Raf, MEK, ERK and Sp1-dependent mechanisms. J Mol Biol 344, 683–695.

Ikari, A., Atomi, K., Takiguchi, A., Yamazaki, Y., Miwa, M., and Sugatani, J. (2009). Epidermal growth factor increases claudin-4 expression mediated by Sp1 elevation in MDCK cells. Biochem Biophys Res Commun 384, 306–310.

Kim, S., Schein, A.J., and Nadel, J.A. (2005). E-cadherin promotes EGFR-mediated cell differentiation and MUC5AC mucin expression in cultured human airway epithelial cells. Am J Physiol Lung Cell Mol Physiol 289, L1049-L1060.

Lee, S.J., Kim, C.E., Seo, K.W., and Kim, C.D. (2010). HNE-induced 5-LO expression is regulated by NF-{kappa}B/ERK and Sp1/p38 MAPK pathways via EGF receptor in murine macrophages. Cardiovasc Res 88, 352–359.

Lee, Y.C., Oslund, K.L., Thai, P., Velichko, S., Fujisawa, T., Duong, T., Denison, M.S., and Wu, R. (2011). 2,3,7,8-Tetrachlorodibenzo-p-dioxin–Induced MUC5AC Expression. Am J Respir Cell Mol Biol 45, 270–276.

Lemjabbar, H., Li, D., Gallup, M., Sidhu, S., Drori, E., and Basbaum, C. (2003). Tobacco smoke-induced lung cell proliferation mediated by tumor necrosis factor alpha-converting enzyme and amphiregulin. J Biol Chem 278, 26202-26207.

Lu, X.-F., Li, E.-M., Du, Z.-P., Xie, J.-J., Guo, Z.-Y., Gao, S.-Y., Liao, L.-D., Shen, Z.-Y., Xie, D., and Xu, L.-Y. (2010). Specificity protein 1 regulates fascin expression in esophageal squamous cell carcinoma as the result of the epidermal growth factor/extracellular signal-regulated kinase signaling pathway activation. Cell Mol Life Sci CMLS 67, 3313–3329.

Merchant, J.L., Shiotani, A., Mortensen, E.R., Shumaker, D.K., and Abraczinskas, D.R. (1995). Epidermal growth factor stimulation of the human gastrin promoter requires Sp1. J Biol Chem 270, 6314–6319.

Mortensen, E.R., Marks, P.A., Shiotani, A., and Merchant, J.L. (1997). Epidermal growth factor and okadaic acid stimulate Sp1 proteolysis. J Biol Chem 272, 16540–16547.

Oyanagi, T., Takizawa, T., Aizawa, A., Solongo, O., Yagi, H., Nishida, Y., Koyama, H., Saitoh, A., and Arakawa, H. (2016). Suppression of MUC5AC expression in human bronchial epithelial cells by interferon-γ. Allergol Int 66, 75-82.

Perrais, M., Pigny, P., Copin, M., Aubert, J., and Van Seuningen, I. (2002). Induction of MUC2 and MUC5AC mucins by factors of the epidermal growth factor (EGF) family is mediated by EGF receptor/Ras/Raf/extracellular signal-regulated kinase cascade and Sp1. J Biol Chem 277, 32258–32267.

Van Winkle, L.S., Isaac, J.M., and Plopper, C.G. (1997). Distribution of epidermal growth factor receptor and ligands during bronchiolar epithelial repair from naphthalene-induced Clara cell injury in the mouse. Am J Pathol 151, 443.

Wu, D.Y.-c., Wu, R., Reddy, S.P., Lee, Y.C., and Chang, M.M.-J. (2007). Distinctive epidermal growth factor receptor/extracellular regulated kinase-independent and-dependent signaling pathways in the induction of airway mucin 5B and mucin 5AC expression by phorbol 12-myristate 13-acetate. Am J Pathol 170, 20-32.

Zheng, X.-L., Matsubara, S., Diao, C., Hollenberg, M.D., and Wong, N.C.W. (2001). Epidermal Growth Factor Induction of Apolipoprotein A-I Is Mediated by the Ras-MAP Kinase Cascade and Sp1. J Biol Chem 276, 13822–13829.