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ErbB2/neu kinase modulates cellular p27(Kip1) and cyclin D1 through multiple signaling pathways
A E Lenferink1, D Busse, W M Flanagan
1Department of Medicine, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
Abstract:
It is well established that ErbB1 and ErbB2 can cooperate in mammary epithelial cell transformation. Therefore, to understand how ErbB1/ErbB2 signaling contributes to this process, we used the ErbB kinase inhibitor AG1478in ErbB2-dependent BT-474 and SKBR-3 human breast cancer cells. These cells overexpress ErbB2 and also display moderate levels of ErbB1. Treatment with AG1478 resulted in rapid ErbB2 dephosphorylation, reversible G(1) arrest, and interruption of constitutive mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K)/Akt signaling. Consequently, both MAPK-dependent transcription of cyclin D1 and phosphorylation of the cyclin-dependent kinase (Cdk) inhibitor p27 were inhibited. The inhibition of PI3K/Akt resulted in increased activity of glycogen synthase kinase-3beta, which phosphorylated cyclin D1, potentially reducing its steady-state levels. The loss of cyclin D1 reduced the amount of cyclin D1/Cdk4 complexes that can sequester p27 in the cytosol. This plus the reduced phosphorylation of p27 by MAPK enhanced the stability of p27 that associated with nuclear Cdk2 at high stoichiometry and inhibited its kinase activity. Antisense p27 oligonucleotides decreased p27 levels and abrogated the G(1) arrest induced by AG1478. Similarly, infection with an adenovirus encoding inducible cyclin D1 also counteracted the antiproliferative effect of AG1478. These data imply that: (a) modulation of both p27 and cyclin D1 are required for the growth arrest that results from blockade of the ErbB2 kinase; and (b) ErbB2 overexpressing cells use both MAPK and PI3K/Akt to modulate p27 and cyclin D1 and, hence, subvert the G(1)-to-S transition.
Insights
Blocking ErbB2 kinase with AG1478 halts breast cancer cell growth by inhibiting mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K)/Akt signaling, affecting cyclin D1 and p27 protein levels.
Area of Science:
- Oncology
- Molecular Biology
- Cell Signaling
Background:
- ErbB1 and ErbB2 signaling pathways are implicated in mammary epithelial cell transformation.
- ErbB2 overexpression is common in certain breast cancers, driving proliferation.
- Understanding ErbB1/ErbB2 cooperation is crucial for targeted cancer therapies.
Purpose of the Study:
- To investigate the role of ErbB1/ErbB2 signaling in breast cancer cell transformation.
- To elucidate the downstream signaling events affected by ErbB2 kinase inhibition.
- To identify key cell cycle regulators modulated by ErbB2 signaling.
Main Methods:
- Utilized the ErbB kinase inhibitor AG1478 in ErbB2-overexpressing breast cancer cell lines (BT-474 and SKBR-3).
- Assessed effects on ErbB2 dephosphorylation, cell cycle progression (G1 arrest), and key signaling pathways (MAPK, PI3K/Akt).
- Investigated the modulation of cell cycle proteins cyclin D1 and p27, employing antisense oligonucleotides and adenovirus-mediated gene expression.
Main Results:
- AG1478 treatment led to rapid ErbB2 dephosphorylation and G1 cell cycle arrest.
- Inhibition of MAPK and PI3K/Akt signaling pathways was observed.
- Modulation of cyclin D1 and p27 stability and activity was critical for the observed G1 arrest.
Conclusions:
- ErbB2 kinase blockade necessitates the modulation of both p27 and cyclin D1 for growth arrest.
- ErbB2-overexpressing cells utilize MAPK and PI3K/Akt pathways to regulate p27 and cyclin D1, thereby controlling the G1-to-S phase transition.