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Expression of the SNT-1/FRS2 phosphotyrosine binding domain inhibits activation of MAP kinase and PI3-kinase pathways and antiestrogen resistant growth induced by FGF-1 in human breast carcinoma cells

Oncogene volume 25, pages 6003–6014 (2006)Cite this article

Abstract

Fibroblast growth factor (FGF) signaling can bypass the requirement for estrogen receptor (ER) activation in the growth of ER-positive (ER+) breast cancer cells. Fibroblast growth factor-1 stimulation leads to phosphorylation of the adaptor protein Suc1-associated neurotrophic factor-induced tyrosine-phosphorylated target (SNT-1) on C-terminal tyrosine residues, whereas it is constitutively bound through its N-terminal phosphotyrosine-binding domain (PTB) to FGF receptors (FGFRs). By expressing the PTB domain of SNT-1 (SNT-1 PTB) in an inducible manner in an ER+ breast carcinoma line, ML20, we asked whether we could uncouple FGFR activation from its downstream signaling components and abrogate FGF-1-induced antiestrogen-resistant growth. Induction of SNT-1 PTB resulted in a significant decrease of FGF-1-dependent tyrosine phosphorylation of endogenous SNT-1, strong inhibition of complex formation between SNT-1, Gab-1 and Sos-1, and reduced activation of Ras, mitogen-activated protein kinase (MAP kinase), and Akt. SNT-1 PTB also inhibited the phosphorylation of p70S6K on Thr421/Ser424 and Ser411, which may result from the abrogation of MAP kinase activity. Moreover, we also observed a decreased phosphorylation of the MAP kinase-independent site Thr389. This may reflect both inhibition of PI-3 kinase pathways and mammalian target of rapamycin (mTOR)-dependent signaling, as the phosphorylation of Thr389 site was sensitive to treatment with the PI3-K and mTOR inhibitors, LY294002 and rapamycin, respectively. Collectively these results suggest that SNT-1 plays a pivotal role in FGF-dependent activation of the Ras-MAP kinase, PI-3 kinase, and mTOR pathways in these cells. Fibroblast growth factor-1 dependent colony formation of ML20 cells in media containing the pure antiestrogen ICI 182,780 was also markedly inhibited upon induction of SNT-1 PTB, suggesting that blockade of FGFR–SNT-1 interactions might abrogate FGF-mediated antiestrogen resistance in breast cancers.

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Accession codes

Accessions

GenBank/EMBL/DDBJ

Abbreviations

CCS:

charcoal-stripped calf serum

FBS:

fetal bovine serum

MAP kinase:

mitogen-activated protein kinase

Erk:

extracellular signal-regulated kinase

IMEM:

Improved Minimal Eagle's Medium

ER:

estrogen receptor

EGF:

epidermal growth factor

FGF:

fibroblast growth factor

HRG-β1:

heregulin β1

PAGE:

polyacrylamide gel electrophoresis

PTB:

phosphotyrosine-binding domain

PRF:

phenol red free

RBD:

Ras-binding domain

SNT-1:

Suc1-associated neurotrophic factor-induced tyrosine-phosphorylated target. FRS2α, FGF receptor substrate 2α

IRES:

internal ribosome entry site

GPCR:

G-protein-coupled receptor

NGF:

nerve growth factor

BDNF:

brain-derived neurophophic factor

GDNF:

glial-derived neurotrophic factor

VEGF:

vascular endothelial growth factor

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Acknowledgements

We thank Dr Bernard Vose (AstraZeneca, Cheshire, UK) for ICI182,780. This work was supported in part by NIH Grant CA50376 (to FGK) and US Army Department of Defense Breast Cancer Research Program DAMD 17-00-1-0434 (to FGK); UAB Comprehensive Cancer Center Core grant P30 CA13148 and UAB Center for AIDS Research Core grant P30 AI 27767. This work was also supported in part by the Adolph Weil Endowed Chair in Cancer Biology at Southern Research Institute (FGK). We also thank Sylvia and David McPherson at the Molecular Biology Core at UAB Center for AIDS for help with some vector constructions. JVT is the recipient of a Susan G Komen Breast Cancer Foundation grant (BCTR00-000456).

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Authors and Affiliations

  1. Drug Discovery Division, Biochemistry and Molecular Biology Department, Southern Research Institute, Birmingham, AL, USA

    M Manuvakhova, J V Thottassery, S Hays, Z Qu, S S Rentz, L Westbrook & F G Kern

  2. UAB Comprehensive Cancer Center, Birmingham, AL, USA

    M Manuvakhova, J V Thottassery, Z Qu & F G Kern

  3. Lexicon Genetics, Inc., The Woodlands, TX, USA

    F G Kern

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  1. M Manuvakhova
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Correspondence to F G Kern.

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Manuvakhova, M., Thottassery, J., Hays, S. et al. Expression of the SNT-1/FRS2 phosphotyrosine binding domain inhibits activation of MAP kinase and PI3-kinase pathways and antiestrogen resistant growth induced by FGF-1 in human breast carcinoma cells. Oncogene 25, 6003–6014 (2006). https://doi.org/10.1038/sj.onc.1209592

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