Bmi-1-green fluorescent protein (GFP)-knock-in mice reveal the dynamic regulation of Bmi-1 expression in normal and leukemic hematopoietic cells

¹ Department of Pathology, Stanford University School of Medicine, Stanford, California; Department of Developmental Biology, Stanford University School of Medicine, Stanford, California; Stanford Institute for Stem Cell Biology and Regenerative Medicine, Palo Alto, California
² Stanford Institute for Stem Cell Biology and Regenerative Medicine, Palo Alto, California

^* To whom correspondence should be addressed. E-mail: hnaoki{at}stanford.edu.

	Abstract

The ability to self-renew is essential for all kinds of stem cells regardless of tissue types. One of the best candidate genes involved in conferring self-renewal capacity is Bmi-1, which has been proven to be essential for the maintenance of both normal adult hematopoietic and leukemia stem cells as well as adult neural stem cells. In order to investigate the possible role of Bmi-1 in other cell types that also self-renew, we generated Bmi-1-green fluorescent protein (GFP)-knock-in mice, in which GFP was expressed under the endogenous transcriptional regulatory elements of the Bmi-1 gene. Using these targeted reporter mice, we demonstrate that Bmi-1 is expressed in hematopoietic stem cells (HSCs) at its highest levels and down-regulated upon commitment to differentiation. An in-vivo reconstitution assay revealed that the frequency of HSCs was 1/16 in Bmi-1^high c-kit⁺lin^- Sca-1⁺ bone marrow (BM) cells and 1/49 in Bmi-1^high lin^- BM cells, suggesting that Bmi-1 may serve as a marker for normal HSCs. In murine leukemia models induced by P210BCR/ABL or TEL/PDGFR + AML1/ETO, Bmi-1 was not over-expressed in leukemic HSC, despite the increase in the HSC numbers. Bmi-1 was expressed at its highest levels in undifferentiated leukemia cells. Furthermore, in several other non-hematopoietic tissues, cells could be separated into distinct subpopulations with differential Bmi-1 expression. Thus, these mice allow for the isolation of viable Bmi-1-expressing cells and have the potential to become a useful tool for understanding the role of Bmi-1 in normal and cancer stem cells in multiple tissue types.

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