FBW7-KLF5-FGF-BP: An emerging pathway in breast cancer

The Kruppel-like factor 5 (KLF5) is a zinc-finger transcription factor promoting cell proliferation, cell cycle progression, survival, and angiogenesis. A high expression level of KLF5 mRNA has been shown to be associated with shorter breast cancer patient survival. However, the mechanism of KLF5 action in breast cancer is still not clear. In this study, we found that manipulation of the KLF5 expression positively regulates the fibroblast growth factor binding protein 1 (FGF-BP) mRNA and protein levels. We demonstrated that KLF5 binds and activates the FGF-BP promoter through luciferase reporter, oligo pull down, and chromatin IP (ChIP) assays. When FGF-BP is depleted by siRNA, KLF5 fails to promote cell proliferation in MCF10A, SW527, and TSU-Pr1 cells. We further demonstrated that exogenous FGF-BP rescues the KLF5 knockdown induced growth arrest in MCF10A cells. In addition, KLF5 overexpression significantly promotes MCF7 breast cancer cell xenograft growth in athymic nude mice and KLF5 depletion significantly suppresses SW527 breast cancer cell xenograft growth in severe combined immune deficiency (SCID) mice. Finally, KLF5 and FGF-BP proteins are co-expressed in ER-/PR-/HER2-breast cell lines and primary tumors. These findings suggest that KLF5 promote breast cancer cell proliferation at least partially through directly activating the FGF-BP mRNA transcription.;It has been previously shown that the KLF5 protein is rapidly turned over through the ubiquitin-proteasome pathway. However, the regulation mechanisms of KLF5 protein degradation are still poorly understood. In this study, we demonstrate that the Skp1-Cul1-Fbw7 E3 ubiquitin ligase complex (SCF Fbw7) targets KLF5 for ubiquitin-mediated degradation in a KLF5 phosphorylation dependent manner. The SCFFbw7 E3 ligase interacts with KLF5 through the WD40 repeats of Fbw7 and a CPD motif (303SPPSS) of KLF5 thus promotes KLF5 ubiquitination and degradation. Mutation of the critical Ser-303 residue in the KLF5 CPD motif abolishes the protein interaction, ubiquitination, and degradation by Fbw7. Using a KLF5 phosphorylation specific antibody and the in vitro kinase assay, we demonstrate that the KLF5 Ser-303 residue can be phosphorylated by GSK3beta. Inhibition of GSK3beta activity by LiCl decreases the KLF5 Ser-303 phosphorylation and delays the KLF5 protein degradation. Consistently, inactivation of endogenous Fbw7 remarkably increases the endogenous KLF5 protein abundances through blocking its degradation. Finally, we found that endogenous Fbw7 suppresses the FGF-BP gene expression through targeting KLF5 protein for degradation. These findings suggest that the Fbw7-KLF5-FGF-BP axis regulates breast cancer cell proliferation and tumorigenesis. Intervention of this signaling pathway may be used to treat triple negative basal breast cancer.