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Ubiquitin-proteasome degradation of KLF5 transcription factor in cancer and untransformed epithelial cells

Department of Oncology and Hematology, Winship Cancer Institute, Emory University School of Medicine, 1365-C Clifton Road, Atlanta, GA, USA.
Oncogene (Impact Factor: 8.56). 06/2005; 24(20):3319-27. DOI: 10.1038/sj.onc.1208497
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ABSTRACT Ubiquitin-mediated proteolysis plays a central role in controlling intracellular levels of essential regulatory molecules such as p53, cyclins, myc, BRCA1, HIF-1alpha, etc. The Kruppel-like factor 5 (KLF5) transcription factor regulates biological processes involved in carcinogenesis, angiogenesis, and smooth muscle cell differentiation. In carcinogenesis, KLF5's role has been indicated by frequent genetic deletion as well as functional studies. Here we show that KLF5 is an unstable protein with a short half-life. Destruction of KLF5 was prevented by each of the proteasome-specific inhibitors tested but not by an inhibitor for trypsin-like proteases and cysteine proteases or by a lysosome inhibitor in epithelial cells. Furthermore, KLF5 underwent ubiquitination, and deletion of a 56-amino-acid sequence adjacent to a known transactivation domain of KLF5 significantly reduced its ubiquitination and degradation. Interestingly, cancer cells appeared to be more active in KLF5 degradation than untransformed epithelial cells, yet their proteasome activity was not higher. These results suggest that KLF5 protein is degraded at least in part through ubiquitination-proteasome pathway, which may have become hyperactive for KLF5 in cancer cells.

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Available from: Keith D Wilkinson, Nov 17, 2014
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    • "during cancer development [10] [11] [12]. In addition, KLF5 protein is degraded by the ubiquitin proteasome pathway, and one E3 ubiquitin ligase that degrades KLF5, WWP1, is amplified and overexpressed in human prostate and breast cancers, causing excessive protein degradation and functional insufficiency of KLF5 [13] [14] [15]. These findings indicate that KLF5 is frequently inactivated during human carcinogenesis and thus could be a tumor suppressor gene, and some functional studies indeed support a tumor suppressor function of KLF5. "
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    ABSTRACT: Krüppel-like factor 5 (KLF5) regulates multiple biologic processes. Its function in tumorigenesis appears contradictory though, showing both tumor suppressor and tumor promoting activities. In this study, we examined whether and how Klf5 functions in prostatic tumorigenesis using mice with prostate-specific deletion of Klf5 and phosphatase and tensin homolog (Pten), both of which are frequently inactivated in human prostate cancer. Histologic analysis demonstrated that when one Pten allele was deleted, which causes mouse prostatic intraepithelial neoplasia (mPIN), Klf5 deletion accelerated the emergence and progression of mPIN. When both Pten alleles were deleted, which causes prostate cancer, Klf5 deletion promoted tumor growth, increased cell proliferation, and caused more severe morphologic and molecular alterations. Homozygous deletion of Klf5 was more effective than hemizygous deletion. Unexpectedly, while Pten deletion alone expanded basal cell population in a tumor as reported, Klf5 deletion in the Pten-null background clearly reduced basal cell population while expanding luminal cell population. Global gene expression profiling, pathway analysis, and experimental validation indicate that multiple mechanisms could mediate the tumor-promoting effect of Klf5 deletion, including the up-regulation of epidermal growth factor and its downstream signaling molecules AKT and ERK and the inactivation of the p15 cell cycle inhibitor. KLF5 also appears to cooperate with several transcription factors, including CREB1, Sp1, Myc, ER and AR, to regulate gene expression. These findings validate the tumor suppressor function of KLF5. They also yield a mouse model that shares two common genetic alterations with human prostate cancer—mutation/deletion of Pten and deletion of Klf5.
    Neoplasia (New York, N.Y.) 11/2014; 16(11). DOI:10.1016/j.neo.2014.09.006 · 5.40 Impact Factor
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    • "The western blot and anti-KLF5 antibody have been described in our previous study (Chen et al., 2005b). The anti-FGF-BP antibody was purchased from R&D Systems (Minneapolis, MN, USA; AF1593) and diluted 500 times for western blot. "
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    ABSTRACT: The Krüppel-like factor 5 (KLF5) is a zinc-finger transcription factor promoting cell proliferation, cell-cycle progression and survival. 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 both KLF5 and its downstream gene fibroblast growth factor binding protein 1 (FGF-BP) are co-expressed in breast cell lines and primary tumors. Manipulation of the KLF5 expression can positively regulate the FGF-BP mRNA and protein levels in multiple breast cell lines. In addition, the secreted FGF-BP protein in the conditional medium is also regulated by KLF5. Furthermore, we demonstrated that KLF5 binds and activates the FGF-BP promoter through a GC box by luciferase reporter, oligo pull down and chromatin immunoprecipitation (ChIP) assays. When FGF-BP is depleted by siRNA, KLF5 fails to promote cell proliferation in MCF10A, SW527 and TSU-Pr1. We further demonstrated that overexpression or addition of FGF-BP rescues the KLF5-knockdown-induced growth arrest in MCF10A cells. Finally, KLF5 significantly promotes MCF7 breast cancer cell xenograft growth in athymic nude mice. These findings suggest that KLF5 may promote breast cancer cell proliferation at least partially through directly activating the FGF-BP mRNA transcription. Understanding the mechanism of KLF5 action in breast cancer may result in useful diagnostic and therapeutic targets.
    Oncogene 09/2009; 28(42):3702-13. DOI:10.1038/onc.2009.235 · 8.56 Impact Factor
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    • "Western blot analysis was performed as described in our previous study (Chen et al., 2005b). Antibodies for Smad2 and Smad3 were purchased from Zymed Laboratories, and antibodies for Smad4, TbRI and p15 were purchased from Cell Signaling. "
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