**2.2.5 ADAR1**

ADAR (adenosine deaminase acting on RNA) catalyzes the deamination of adenosine to inosine in double-stranded RNA. Conventional *Adar*-/- mice die around embryonic day 11.5–12 because of widespread apoptosis and defective hematopoiesis (Hartner et al., 2004; Wang et al., 2004). Conditional deletion of Adar in HSCs shows that ADAR1 is essential for the maintenance of both fetal and adult HSCs, and leads to global upregulation of type I and II interferon-inducible transcripts and rapid apoptosis (Hartner et al., 2009). Interferon regulatory factor-2 (Irf2), a transcriptional suppressor of type I interferon signaling, is a positive regulator of HSC quiescence (Sato et al., 2009). Irf2-deficient HSCs are unable to restore hematopoiesis in irradiated mice, but the reconstituting capacity of *Irf2*-/- HSCs can be restored in these cells by disabling type I IFN signaling.

#### **2.3 Response to hematopoietic emergency**

Various external stresses, such as myelosuppressive chemotherapy, bleeding, infection, and total body irradiation, put HSCs under stress, as they must proliferate to produce large numbers of primitive progenitor cells, thereby enabling rapid hematologic regeneration. Although this property has long been recognized, the molecular basis underlying the reaction of HSCs to hematologic emergency remains enigmatic. However, some key players have been identified.

#### **2.3.1 Heme oxygenase-1**

Heme promotes the proliferation and differentiation of hematopoietic progenitor cells (HPCs) (Chertkov et al., 1991) and stimulates hematopoiesis (Porter et al., 1979; Abraham, 1991). The degradation of heme is catalyzed by heme oxygenase (HO). HO-1, encoded by the *Hmox1* gene, is the stress-inducible isozyme of HO and is highly expressed in the spleen and BM (Abraham, 1991). Heterozygous HO-1–deficient mice (*HO-1*+/-) show accelerated hematologic recovery from myelotoxic injury induced by 5-FU treatment, and mice transplanted with *HO-1*+/- BM cells show more rapid hematopoietic repopulation than those transplanted with *Ho-1*+/+ BM cells. However, *HO-1*+/- HSCs show a reduced capacity to rescue lethally irradiated mice and to serially repopulate irradiated recipients (Cao et al., 2008). These results suggest that HO-1 limits the proliferation and differentiation of HPCs under stressful conditions, and that the failure of this mechanism can lead to the premature exhaustion of the HSC pool.

Regulation of Hematopoietic Stem Cell Fate: Self-Renewal, Quiescence and Survival 51

therapies (Dick, 2008). In an acute myelogenous leukemia (AML) xenograft model, AML leukemic stem cells (LSCs) localized in the endosteal region of the BM show cellular quiescence and resistance to chemotherapy (Ishikawa et al., 2007; Saito et al., 2010). In patients with chronic myelogenous leukemia (CML), CD34+ progenitor cells contain dormant cells that are resistant to BCR/ABL tyrosine kinase inhibitors (Bhatia et al., 2003). It is well documented that regulators of HSC maintenance are also involved in the development of leukemias (Rizo et al., 2006). A number of cancer-related proteins, such as Bmi1, c-Myc, p53, Gfi-1, and PTEN, are key participants in HSC regulation, demonstrating the close relationship between normal HSCs and CSCs. Therefore, further understanding the mechanisms regulating HSC fate is needed if we are to develop new strategies for

In this review, we have briefly summarized a number of critical regulators involved in the control of HSC self-renewal, quiescence, survival, and responses to external insults. Recent evidence strongly suggests that the BM niche also plays an integral role by providing critical signals that maintain HSCs in a stat of hibernation, thus preventing them from exhausting themselves. However, HSCs are critical for the maintenance and regeneration of an organism after injury/illness. This process must be tightly regulated and coordinated. Intensive studies have uncovered the molecular signatures and key molecules regulating HSC behavior. Moreover, new systems approaches, such as microRNA expression profiling and protein expression profiling, are expected to provide further useful information about HSC biology in the future. However, the overall picture of the molecular mechanisms that govern HSC fate is still unclear. Further understanding of the systems that regulate HSCs will enable the manipulation of stem cells for use in tissue engineering and cell-based

This work was supported by a Grant-in-Aid for Young Scientists to Y.K. (no. 23791083) from

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the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan.

targeting CSCs and successfully treat cancer.

**4. Conclusions** 

therapies.

**5. Acknowledgments** 

*Rev* 5(1):19-28.

**6. References** 

389.

#### **2.3.2 Necdin**

Necdin is a member of the melanoma antigen family of molecules, whose physiological roles have not been well characterized (Xiao et al., 2004). Necdin acts as a cell cycle regulator in post-mitotic neurons (Yoshikawa, 2000). Intriguingly, recent genetic analyses show that aberrant genomic imprinting of *NDN* on the human 15q11-q13 chromosomal region is, at least in part, responsible for the pathogenesis of Prader-Willi syndrome (MacDonald & Wevrick, 1997; Nakada et al., 1998; Barker et al., 2002), a disorder associated with a mildly increased risk of myeloid leukemia (Davies et al., 2003). Necdin interacts with multiple cell-cycle related proteins, such as SV-40 large T antigen, adenovirus E1A, E2F1, and p53 (Taniura et al., 1998, 1999, 2005; Hu et al., 2003). As shown in Table 2, necdin is one of 32 genes that show higher expression in HSCs than in differentiated hematopoietic cells (Kubota et al., 2009). Other groups also found that necdin is highly expressed in HSCs (Forsberg et al., 2005; Liu et al., 2009). Necdindeficient mice show accelerated recovery of hematopoietic systems after myelosuppressive stress, such as 5-FU treatment and BM transplantation, whereas no overt abnormality is seen under conditions of steady-state hematopoiesis. Considering necdin as a potential negative cell-cycle regulator, it was reasoned that the enhanced hematologic recovery in necdin-null mice could be the result of an increased number of proliferating HSCs and progenitor cells. As expected, after 5-FU treatment, necdindeficient mice had an increased number of HSCs, but this was only transiently observed during the recovery phase (Kubota et al., 2009). These data suggest that the repression of necdin function in HSCs may present a novel strategy for accelerating hematopoietic recovery, thus providing therapeutic benefits after clinical myelosuppressive treatments (e.g., cytoablative chemotherapy or HSC transplantation).

#### **2.3.3 Slug**

Slug belongs to the highly conserved Slug/Snail family of zinc-finger transcriptional repressors found in diverse species ranging from *C. elegans* to humans. SLUG is a target gene for the E2A-HLF chimeric oncoprotein in pro-B cell acute leukemia (Inukai et al., 1999). Slug-deficient mice show normal peripheral blood counts, but they are very sensitive to γirradiation (Inoue et al., 2002). Slug is induced by p53 and protects primitive hematopoietic cells from apoptosis triggered by DNA damage. Slug exerts this function by repressing Puma, a proapoptotic target of p53 (Wu et al., 2005). Sun et al. recently showed that Slug negatively regulates the repopulating ability of HSCs under conditions of stress. Slug deficiency increases HSC proliferation and reconstitution potential *in vivo* after myelosuppressive treatment, and accelerates HSC expansion during *in vitro* culture (Sun et al., 2010).
