Epidermal Growth Factor (EGF) Receptor Kinase-independent Signaling by EGF

Deb, Tushar B.; Su, Leon; Wong, Lily; Bonvini, Ezio; Wells, Alan; David, Michael; Johnson, Gibbes R.

doi:10.1074/jbc.m100928200

Cited by 60 publications

(59 citation statements)

References 36 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In fact, expression of ErbB4, a protein with limited S C HANLEY and others . EGF induces islet cell dedifferentiation affinity for EGF and not required for EGFmediated activation of the AKT and ERK signaling pathways (Deb et al 2001), was the only receptor transcript to be significantly downregulated in this context. Thus, EGF ligands are constitutively expressed by isolated islets, even after embedding and/or EGF administration.…”

Section: Discussionmentioning

confidence: 99%

Epidermal growth factor induces adult human islet cell dedifferentiation

Hanley¹,

Assouline-Thomas²,

Makhlin³

et al. 2011

Journal of Endocrinology

View full text Add to dashboard Cite

Given the inherent therapeutic potential of the morphogenetic plasticity of adult human islets, the identification of factors controlling their cellular differentiation is of interest. The epidermal growth factor (EGF) family has been identified previously in the context of pancreatic organogenesis. We examined the role of EGF in an in vitro model whereby adult human islets are embedded in a collagen gel and dedifferentiated into duct-like epithelial structures (DLS). We demonstrated that DLS formation was EGF dependent, while residual DLS formation in the absence of added EGF was abrogated by EGF receptor inhibitor treatment. With respect to signaling, EGF administration led to an increase in c-Jun NH2-terminal kinase (JNK) phosphorylation early in DLS formation and in AKT and extracellular signalregulated kinase (ERK) phosphorylation late in the process of DLS formation, concomitant with the increased proliferation of dedifferentiated cells. In the absence of EGF, these phosphorylation changes are not seen and the typical increase in DLS epithelial cell proliferation seen after 10 days in culture is attenuated. Thus, in our model, EGF is necessary for islet cell dedifferentiation, playing an important role in both the onset of DLS formation (through JNK) and in the proliferation of these dedifferentiated cells (through AKT and ERK).

show abstract

Section: Discussionmentioning

confidence: 99%

Epidermal growth factor induces adult human islet cell dedifferentiation

Hanley¹,

Assouline-Thomas²,

Makhlin³

et al. 2011

Journal of Endocrinology

View full text Add to dashboard Cite

show abstract

“…Cells were allowed to recover for 24 h and serum-starved overnight, and lysates were generated as described previously (30). Expression of transfected cDNAs was confirmed by Western blotting analysis (30). To phosphorylate Gab1 or Shc, the cell lysates were mixed with purified EGFR, and the reactions were initiated by the addition of kinase reaction buffer containing ATP.…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…32D cells are excellent for this purpose because they grow rapidly in suspension and do not express any endogenous ErbBs (30,31). A cDNA encoding the 1186-amino acid residue full-length EGFR with a 25-amino acid sequence containing a Myc and hexahistidine epitope sequence at the C terminus was generated.…”

Section: Expression and Purification Of Epitope-tagged Egfr-tomentioning

confidence: 99%

“…However, EGF increased the k cat value ϳ5.6-fold as evidenced by distinct intersections of the plots with ordinate. As summarized in Table I 6) were serum-starved, stimulated with 10 nM EGF for 5 min at 37°C, and lysates generated as described previously (30). Lysates were separated in SDS-PAGE and transferred to a polyvinylidene difluoride membrane, and Western blotting was performed with antibodies against Myc, MAPK, and phosphorylated MAPK.…”

Section: Egf Does Not Affect the Relative Binding Affinities Toward Ementioning

confidence: 99%

See 2 more Smart Citations

Ligand Regulates Epidermal Growth Factor Receptor Kinase Specificity

Fan

Wong

Deb³

et al. 2004

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

The epidermal growth factor receptor (EGFR) kinase catalyzes phosphorylation of tyrosines in its C terminus and in other cellular targets upon epidermal growth factor (EGF) stimulation. Here, by using peptides derived from EGFR autophosphorylation sites and cellular substrates, we tested the hypothesis that ligand may function to regulate EGFR kinase specificity by modulating the binding affinity of peptide sequences to the active site. Measurement of the steady-state kinetic parameters, K m and k cat , revealed that EGF did not affect the binding of EGFR peptides but increased the binding affinity for peptides corresponding to the major EGFRmediated phosphorylation sites of the adaptor proteins Gab1 (Tyr-627) and Shc (Tyr-317), and for peptides containing the previously identified optimal EGFR kinase substrate sequence EEEEYFELV (3-7-fold). Conversely, EGF stimulation increased k cat ϳ5-fold for all peptides. Thus, ligand changed the relative preference of the EGFR kinase for substrates as evidenced by EGF increases of ϳ5-fold in the specificity constants (k cat /K m ) for EGFR peptides, whereas ϳ15-40-fold increases were observed for other peptides, such as Gab1 Tyr-627. Furthermore, we demonstrate that EGF (i) increased the binding affinity of EGFR to Gab1 Tyr-627 and Shc Tyr-317 sites in purified GST fusion proteins ϳ4 -6-fold, and (ii) EGF significantly enhanced the phosphorylation of these sites, relative to EGFR autophosphorylation, in cell lysates containing the full-length Gab1 and Shc proteins. Analysis of peptides containing amino acid substitutions indicated that residues C-terminal to the target tyrosine were critical for EGF-stimulated increases in substrate binding and regulation of kinase specificity. To our knowledge, this represents the first demonstration that ligand can alter specificity of a receptor kinase toward physiologically relevant targets.Protein-tyrosine kinases, which catalyze the transfer of ␥-phosphate from ATP to tyrosine side chains of peptide and protein substrates, play a central role in cellular signaling. Receptor tyrosine kinases (RTKs) 1 constitute one class of protein kinases that consist of an extracellular ligand binding, a transmembrane, an intracellular tyrosine kinase, and a Cterminal domain that contains tyrosine phosphorylation sites (1). The epidermal growth factor receptor (EGFR, ErbB1, and HER1) is the prototypical member of the ErbB family RTKs, which also includes ErbB2 (HER2, neu), ErbB3 (HER3), and ErbB4 (HER4). Binding of growth factors that are structurally related to EGF induces a process of the receptor dimerization, kinase activation and autophosphorylation (2). EGFR activation results in recruitment and phosphorylation of cytosolic downstream targets such as Grb2-associated binding protein 1 (Gab1) (3), Src homology 2 (SH2) domain, collagen-containing protein (Shc) (4), and phospholipase C␥-1 (PLC␥-1) (5, 6). EGFR functions in the proliferation, migration, survival, and differentiation of mammalian cells (2, 7), and dysregulation of signaling by EGFR ...

show abstract

The effects of gefitinib in tamoxifen‐resistant and hormone‐insensitive breast cancer: A phase II study

Gutteridge

Agrawal

Nicholson

et al. 2009

Intl Journal of Cancer

View full text Add to dashboard Cite

Estrogen receptor (ER)-positive acquired tamoxifen-resistant (TAM-R) MCF-7 breast cancer cell lines exhibit epidermal growth factor receptor (EGFR) expression/signaling and are growth-inhibited by gefitinib (IRESSA). We examined the effect of gefitinib on ER-positive TAM-R and ER-negative hormone-insensitive breast cancer in a Phase II study. Fifty-four patients with breast cancer [ER-positive/acquired TAM-R (n 5 28); ER-negative (n 5 26)] received oral gefitinib 500 mg/day. Tumor biopsies were taken pre-(n 5 28) and 8 weeks post-treatment (n 5 14 matched samples). Gefitinib was well tolerated and the clinical benefit rate (objective response or stable disease >24 weeks) was 33.3% overall (n 5 18/54), and 53.6 and 11.5% in ERpositive/TAM-R and ER-negative patients, respectively. Pretreatment ER and progesterone receptor-positivity were associated with response (p < 0.001 and 0.016, respectively) and longer progression-free survival (PFS; p5 0.001 and 0.013, respectively). All patients expressed EGFR, but high pretreatment levels predicted poorer outcome (p 5 0.005) and shorter PFS (p 5 0.012) with gefitinib. In patients with clinical benefit, reduced Ki67 staining during treatment (p 5 0.024) was commonly observed, and those with >10% decline in EGFR phosphorylation demonstrated parallel decreases in ERK1/2 MAPK phosphorylation. Acquired tamoxifen resistance appears in part mediated through EGFR signaling and can be blocked with gefitinib.

show abstract

Epidermal Growth Factor (EGF) Receptor Kinase-independent Signaling by EGF

Cited by 60 publications

References 36 publications

Epidermal growth factor induces adult human islet cell dedifferentiation

Epidermal growth factor induces adult human islet cell dedifferentiation

Ligand Regulates Epidermal Growth Factor Receptor Kinase Specificity

The effects of gefitinib in tamoxifen‐resistant and hormone‐insensitive breast cancer: A phase II study

Contact Info

Product

Resources

About