Abstract LB-21: Regulation of Bmi1 by β-catenin and KLF4 and its implication in colon cancer

B lymphoma Mo-MLV insertion region 1 (Bmi1) is a Polycomb Group (PcG) protein that is important in gene silencing. It is a component of Polycomb Repressive Complex 1 (PRC1), which is required to maintain the transcriptionally repressive state of many genes. Bmi1 was initially identified as an oncogene that regulates cell proliferation and transformation, and is important in hematopoiesis and development of nervous systems. Recently, great attention is drawn upon the role of Bmi1 in stem cell regulation and cancer. It was reported that Bmi1 is a potential marker for the intestinal stem cells. Since Wnt signaling plays a key role in the intestinal stem cells, we analyzed the effects of Wnt signaling on Bmi1 expression in colon cancer cell lines. Kruppel-like factor 4 (KLF4) is a zinc finger protein that is highly expressed in the gut and skin. We recently found that KLF4 crosstalks with Wnt/[[Unsupported Character - Symbol Font ]]-catenin in regulating intestinal homeostasis. In addition, KLF4 is one of the four factors that induce pluripotent stem cells, thus playing a crucial role in stem cell regulation. In this study, the effects of KLF4 on Bmi1 expression were analyzed as well. These studies will provide novel insights into the mechanisms of Bmi1 function and regulation in the intestinal stem cells and colorectal cancers. Since Bmi1 is a stem cell or progenitor cell marker, we analyzed Bmi1 expression in human colorectal cancers tissues by IHC. Bmi1 levels are significantly increased in the tumor tissues from colon cancer patients compared with adjacent normal tissues; and expression of Bmi1 is significantly associated with nuclear [[Unsupported Character - Symbol Font ]]-catenin. Bmi1 knockdown by lentivirus-mediated Bmi1-shRNA repressed growth of different colon cancer cell lines, inhibited xenograft tumor growth, and increased differentiation of colon cancer cells in mouse xenograft. We found that dominant negative TCF (DnTCF), which blocks Wnt signaling, only marginally inhibits Bmi1 expression. However, both Western blot and RT-PCR results demonstrated that the expression of Bmi1 was inhibited by KLF4 in LS174T colon cancer cells. Using luciferase reporter assay, we found that repression of Bmi1 promoter by KLF4 is independent of c-myc. Using ChIP assay, we confirmed that KLF4 binds the endogenous Bmi1 promoter in colon cancer cells. Finally, we found that KLF4 inhibits Bmi1-mediated histone ubiquitination but by regulating more than one protein in the Polycomb complex. We found that Bmi1 is required for colon cancer cell proliferation and tumor growth, and it is up-regulated in colon cancer. Wnt signaling weakly regulates Bmi1; KLF4 inhibits Bmi1 by directly binding to Bmi1 promoter, thus inhibiting Bmi1-mediated H2A ubiquitination. It is of great interest to further study the mechanisms of Bmi1 regulation and the function of Bmi1 in normal intestine and in colon tumorigenesis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-21. doi:1538-7445.AM2012-LB-21