Abstract
BackgroundThe Bmi1 polycomb ring finger oncogene, a transcriptional repressor belonging to the Polycomb group of proteins plays an important role in the regulation of stem cell self-renewal and is elevated in several cancers. In the current study, we have explored the role of Bmi1 in regulating the stemness and drug resistance of breast cancer cells.MethodsUsing real time PCR and immunohistochemistry primary breast tissues were analyzed. Retro- and lentiviruses were utilized to overexpress and knockdown Bmi1, RT-PCR and Western blot was performed to evaluate mRNA and protein expression. Stemness properties were analyzed by flow cytometry and sphere-formation and tumor formation was determined by mouse xenograft experiments. Dual luciferase assay was employed to assess promoter activity and MTT assay was used to analyze drug response.ResultsWe found Bmi1 overexpression in 64% of grade III invasive ductal breast adenocarcinomas compared to normal breast tissues. Bmi1 overexpression in immortalized and transformed breast epithelial cells increased their sphere-forming efficiency, induced epithelial to mesenchymal transition (EMT) with an increase in the expression of stemness-related genes. Knockdown of Bmi1 in tumorigenic breast cells induced epithelial morphology, reduced expression of stemness-related genes, decreased the IC50 values of doxorubicin and abrogated tumor-formation. Bmi1-high tumors showed elevated Nanog expression whereas the tumors with lower Bmi1 showed reduced Nanog levels. Overexpression of Bmi1 increased Nanog levels whereas knockdown of Bmi1 reduced its expression. Dual luciferase promoter-reporter assay revealed Bmi1 positively regulated the Nanog and NFκB promoter activity. RT-PCR analysis showed that Bmi1 overexpression activated the NFκB pathway whereas Bmi1 knockdown reduced the expression of NFκB target genes, suggesting that Bmi1 might regulate Nanog expression through the NFκB pathway.ConclusionsOur study showed that Bmi1 is overexpressed in several high-grade, invasive ductal breast adenocarcinomas, thus supporting its role as a prognostic marker. While Bmi1 overexpression increased self-renewal and promoted EMT, its knockdown reversed EMT, reduced stemness, and rendered cells drug sensitive, thus highlighting a crucial role for Bmi1 in regulating the stemness and drug response of breast cancer cells. Bmi1 may control self-renewal through the regulation of Nanog expression via the NFκB pathway.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-785) contains supplementary material, which is available to authorized users.
Highlights
IntroductionThe Bmi polycomb ring finger oncogene, a transcriptional repressor belonging to the Polycomb group of proteins plays an important role in the regulation of stem cell self-renewal and is elevated in several cancers
The B lymphoma Mo-MLV insertion region 1 homolog (Bmi1) polycomb ring finger oncogene, a transcriptional repressor belonging to the Polycomb group of proteins plays an important role in the regulation of stem cell self-renewal and is elevated in several cancers
Normal breast tissues either lacked Bmi1 or showed lower cytoplasmic expression (Figure 1B). These results indicated that compared to normal breast tissue, Bmi1 expression is higher in breast tumor tissues both at mRNA and protein levels
Summary
The Bmi polycomb ring finger oncogene, a transcriptional repressor belonging to the Polycomb group of proteins plays an important role in the regulation of stem cell self-renewal and is elevated in several cancers. We have explored the role of Bmi in regulating the stemness and drug resistance of breast cancer cells. Cancer cells undergoing epithelial to mesenchymal transition, considered to be a pre-requisite for solid tumor metastasis, have been shown to acquire stem-like properties [10]. We have recently shown that the very transcription factors that bring about an EMT lead to an increased expression of ABC family of transporters, thereby increasing drug resistance [11]. These data suggest that the properties of selfrenewal, EMT, and drug resistance may all be linked [12]. Understanding the pathways and mechanisms that regulate the stem-like properties of cancer cells is fundamental for their effective therapeutic targeting
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