Id-1 induces cyclin D1 expression and cytoplasmic localization of p27Kip1 via activation of PI3K/Akt/Wnt pathway in breast cancer cells

Abstract

Recent evidence has revealed that inhibitor of DNA binding and differentiation (Id) proteins, especially Id-1, are able to promote invasion, angiogenesis and metastasis. Wnt signaling is suggested as an essential pathway in epithelial-mesenchymal transition (EMT), which is indicated to play an important role in the invasion and progression of human cancer. In this regard, the relationship between Id-1 overexpression and Wnt signaling has not been investigated. Therefore, we examined the regulation of Wnt signaling pathway in Id-1 overexpressing human breast cancer cells. MCF7 stable cells overexpressing Id-1 (MCF7 Id-1) showed increased phosphorylation of Akt at Ser473 which was associated with activation of Akt signaling pathway. Phosphorylated Akt inhibited GSK-3β by phosphorylation at Ser9. Furthermore, stabilization and nuclear localization of β-catenin was enhanced by Id-1 as shown by nuclear/cytoplasmic fractionation, immunoblotting and immnofluorescence microscopy. In addition, TCF/LEF transactivation activity was also augmented as artificial reporter (TOP) activity and Cyclin D1 expression were enhanced by Id-1. Such changes were reversed upon reduced Id-1 expression by retroviral antisense expression or PI3K inhibitor, LY294002. Thus, Akt-mediated canonical Wnt signaling pathway was upregulated by Id-1. On the other hand, p27Kip1 phosphorylation at Thr157, which is mediated by Akt, was enhanced in MCF7 Id-1 cells and Akt-mediated cytoplasmic localization of p27Kip1 was also increased. These effects were reversed by blockade of Id-1 expression and treatment with PI3K inhibitor. Therefore, cytoplasmic localization of p27Kip1 may have additionally contributed to enhanced cell proliferation by Id-1. In conclusion, activation of PI3K/Akt-mediated Wnt signaling may be essential for Id-1-induced cell proliferation, and also provide a possible mechanism of EMT on the molecular basis of breast cancer.