Investigation of Bmi1 phosphorylation in breast cancer cells
Bmi1 is a polycomb group protein (PcG) frequently over expressed in multiple types of human cancer cells. Bmi1 promotes monoubiquitination of histone H2A, causing gene silencing often leading to the cancer stem cell (CSC) phenotype. Previous studies have shown that Bmi1 displays two mobility forms during electrophoresis presumably due to phosphorylation. Since Bmi1 is an important regulator of tumorigenesis and stem cell self-renewal, any post translational modification regarding the regulation of this protein could be of significance, and elucidate the molecular mechanism of Bmi1 regulation that could contribute to both cancer and stem cell research. For this purpose, we treated mitotic Bmi1 with phosphatase and found that the upshifted Bmi1 was converted to the fast migrating form similar to the interphase Bmi1, confirming that the upshift of Bmi1 is indeed due to phosphorylation. Furthermore, by using synchronized breast cancer cells as a model, we did a timecourse study and found that Bmi1 is phosphorylated at the onset of mitosis, and dephosphorylated at the end mitosis, suggesting that one or more mitotic kinases are responsible for Bmi1 phosphorylation. Since Bmi1 is a ubiquitin ligase, and its target protein, histone H2A, is deubiquitinated in mitosis, we hypothesized that the phosphorylation of Bmi1 might contribute to the inactivation of Bmi1. This study has set the foundation for the identification of the upstream kinase(s) that phosphorylate Bmi1 in mitosis, allowing us to better understand the molecular mechanism of breast cancer development.
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