Id2 Is Required for the Self-Renewal and Proliferation of Hematopoietic Stem Cells

The production of mammalian blood cells is sustained throughout life by the self-renewal and differentiation of hematopoietic stem cells (HSCs). Dysregulation in this system leads to different pathologies including anemia, bone marrow failure and hematopoietic malignancies. The Helix-Loop-Helix transcriptional regulator Id2 plays essential roles in regulating proliferation and cell fate of hematopoietic progenitors; however, its role in regulating HSC development remains largely unknown. To assess the function of Id2 in HSCs, we developed two mouse models, including an Id2 conditional knockout model and an Id2-EYFP model, in which EYFP expression is driven by endogenous Id2 promoter. When we examined HSC function by serial transplantation, we found that mice transplanted with Id2^F/F Mx1-Cre+ conditionally deleted bone marrow cells became moribund more rapidly after primary and secondary transplantation, compared to those transplanted with Id2^+/F Mx1-Cre+ bone marrow, suggesting that HSC self-renewal is impaired when Id2 is deleted. To further determine if self-renewal and maintenance of HSCs depends on the expression level of Id2, we purified HSCs with different levels of Id2 expression using Id2-EYFP mice to specifically address the role of Id2 in HSCs. First, we confirmed Id2 is highly expressed in HSCs in this model. Second, when HSCs with either low or high levels of Id2-EYFP were transplanted into irradiated mice, cells with high levels of Id2 reconstituted transplanted recipients faster than those with low levels of Id2 at 3 weeks and longer, suggesting that Id2 expression is associated with repopulation advantage. Furthermore, Ki-67 staining showed that HSCs with high levels of Id2 have 15-fold more cells in G2/M phase, and fewer cells in G0. BrdU staining also suggested that there are 5-fold more BrdU+ cells in HSCs with high levels of Id2, indicating that Id2 expression correlates with cell cycle progression in HSCs. In addition, p57 has been reported to be required for quiescence of HSCs. Our preliminary data showed that p57 is downregulated in HSCs with high levels of Id2, and p57 is correspondingly upregulated in Id2-null HSCs. Altogether, our data demonstrate that Id2 is required for the self-renewal and proliferation of HSCs, and suggest a link between Id2 and the transcriptional regulatory networks that regulate the functional hematopoietic system. Since Id2 is also expressed in other adult stem cells including muscle and neuronal stem cells, as well as cancer cells, we believe our results can improve our understanding of stem cell biology and cancer development, and contribute to the identification of novel molecules that may be targeted to eliminate cancer stem cells.