Abstract
We have examined overexpression of the human epidermal growth factor receptor 2 (HER2) to determine if it modifies the anti-proliferative effect of transforming growth factor (TGF)-beta against MCF-10A human mammary epithelial cells. Exogenous TGF-beta inhibited cell proliferation and induced Smad-dependent transcriptional reporter activity in both MCF-10A/HER2 and MCF-10A/vector control cells. Ligand-induced reporter activity was 7-fold higher in HER2-overexpressing cells. In wound closure and transwell assays, TGF-beta induced motility of HER2-transduced, but not control cells. The HER2-blocking antibody trastuzumab (Herceptin) prevented TGF-beta-induced cell motility. Expression of a constitutively active TGF-beta type I receptor (ALK5(T204D)) induced motility of MCF-10A/HER2 but not MCF-10A/vector cells. TGF-beta-induced motility was blocked by coincubation with either the phosphatidylinositol 3-kinase inhibitor LY294002, the mitogen-activated protein kinase (MAPK) inhibitor U0126, the p38 MAPK inhibitor SB202190, and an integrin beta(1) blocking antibody. Rac1 activity was higher in HER2-overexpressing cells, where both Rac1 and Pak1 proteins were constitutively associated with HER2. Both exogenous TGF-beta and transduction with constitutively active ALK5 enhanced this association. TGF-beta induced actin stress fibers as well as lamellipodia within the leading edge of wounds. Herceptin blocked basal and TGF-beta-stimulated Rac1 activity but did not repress TGF-beta-stimulated transcriptional reporter activity. These data suggest that 1) overexpression of HER2 in nontumorigenic mammary epithelial is permissive for the ability of TGF-beta to induce cell motility and Rac1 activity, and 2) HER2 and TGF-beta signaling cooperate in the induction of cellular events associated with tumor progression.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Actins / metabolism
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Activin Receptors, Type I / metabolism
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Adenoviridae / genetics
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Antibodies, Monoclonal / metabolism
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Antibodies, Monoclonal / pharmacology
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Antibodies, Monoclonal, Humanized
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Blotting, Northern
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Breast Neoplasms / metabolism*
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Bromodeoxyuridine / pharmacology
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Butadienes / pharmacology
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Cell Cycle
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Cell Division
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Cell Line
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Cell Line, Tumor
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Cell Movement
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Chromones / pharmacology
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DNA, Complementary / metabolism
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Disease Progression
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Enzyme Inhibitors / pharmacology
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Epithelial Cells / metabolism*
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Gene Expression Regulation
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Genes, Reporter
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Green Fluorescent Proteins
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Humans
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Imidazoles / pharmacology
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Immunoblotting
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Integrin beta1 / immunology
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Ligands
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Luminescent Proteins / metabolism
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Microscopy, Fluorescence
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Models, Genetic
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Morpholines / pharmacology
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Nitriles / pharmacology
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Phosphoinositide-3 Kinase Inhibitors
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Precipitin Tests
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Protein Serine-Threonine Kinases
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Pseudopodia / metabolism
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Pyridines / pharmacology
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Receptor, ErbB-2 / biosynthesis*
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Receptor, Transforming Growth Factor-beta Type I
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Receptors, Transforming Growth Factor beta / metabolism
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Retroviridae / genetics
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Signal Transduction
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Transcription, Genetic
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Transforming Growth Factor beta / metabolism*
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Trastuzumab
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Wound Healing
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rac1 GTP-Binding Protein / metabolism
Substances
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Actins
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Antibodies, Monoclonal
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Antibodies, Monoclonal, Humanized
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Butadienes
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Chromones
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DNA, Complementary
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Enzyme Inhibitors
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Imidazoles
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Integrin beta1
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Ligands
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Luminescent Proteins
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Morpholines
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Nitriles
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Phosphoinositide-3 Kinase Inhibitors
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Pyridines
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Receptors, Transforming Growth Factor beta
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Transforming Growth Factor beta
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U 0126
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Green Fluorescent Proteins
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2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
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Receptor, ErbB-2
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Protein Serine-Threonine Kinases
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Activin Receptors, Type I
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Receptor, Transforming Growth Factor-beta Type I
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TGFBR1 protein, human
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rac1 GTP-Binding Protein
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Bromodeoxyuridine
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Trastuzumab
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4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole