**2.2.2 Scl, Lyl1**

Scl/Tal1 is a basic helix-loop-helix (bHLH) transcription factor that is essential for the development of HSCs in the embryo (Robb et al., 1995; Shivdasani et al., 1995). During adult hematopoiesis, Scl/Tal1 is highly expressed in LT-HSCs compared with short-term HSCs and progenitor cells (Lacombe et al., 2010). However, a study using conditional Scl/Tal1 knockout mice revealed that Scl/Tal1 is required for the generation of, but not the maintenance of, adult HSCs (Mikkola et al., 2003). Another group showed that conditional deletion of Scl/Tal1 in adult HSCs has a relatively mild effect: Scl-null HSCs show impaired short-term repopulating ability, but no defect in long-term repopulating capacity (Curtis et al., 2004). Redundant activity caused by the expression of Lyl1, a related bHLH transcription factor, in adult HSCs may provide an explanation for these "mild" phenotypes. While adult HSCs in single-knockout mice show no or only a mild phenotype, Lyl1;Scl-conditional double-knockout mice show a gene dosage defect on HSC survival, as HSCs and progenitor cells are immediately lost due to apoptosis (Souroullas et al., 2009).

Recently, Lacombe et al. demonstrated that Scl/Tal1 is required for the maintenance of the quiescent stem cell pool (Lacombe et al., 2010). Cell-cycle analyses revealed that Scl/Tal1 negatively regulates the G0-G1 transit of LT-HSCs; however, these phenomena were specific to adult HSCs and were not observed in perinatal HSCs. The reconstituting ability of *Scl+/-* HSCs or HSCs with decreased Scl protein expression induced by RNA interference was impaired in various transplantation assays. Furthermore, gene expression analysis and chromatin immunoprecipitation experiments revealed that the Cdkn1a and Id1 genes are direct SCL targets.

#### **2.2.3 Tel/Etv6**

The transcription factor Tel (also known as Etv6), an Ets-related transcriptional repressor, is a frequent target of the diverse chromosomal translocations observed in leukemias (Golub et al., 1994). Tel/ETV6 is also required for HSC survival in adult hematopoiesis. Following conditional inactivation of Tel/Etv6, HSCs are rapidly lost from the adult BM. However, Tel/Etv6 is not required for the maintenance of lineage-committed progenitors. Conditional deletion of Tel/Etv6 after lineage commitment does not affect the differentiation or survival of these progenitors, although it does impair the maturation of megakaryocytes (Hock et al., 2004).

#### **2.2.4 Zfx**

48 Advances in Hematopoietic Stem Cell Research

Accumulating evidence suggests that the suppression of apoptosis is required for HSC survival. Forced expression of Bcl-2 increases the number of HSCs and provides them with enhanced competitive repopulation ability (Domen et al., 1998, 2000), suggesting that cell

Mcl-1, another anti-apoptotic Bcl-2 family member, is also an essential regulator of HSC survival. Mcl-1 is highly expressed in LT-HSCs, and conditional deletion of MCl-1 results in the loss of the early BM progenitor population, including HSCs, leading to fatal hematopoietic failure (Opferman et al., 2005). Recently, it was reported that Mcl-1 is an indispensable regulator of self-renewal in human stem cells and that functional dependence

Scl/Tal1 is a basic helix-loop-helix (bHLH) transcription factor that is essential for the development of HSCs in the embryo (Robb et al., 1995; Shivdasani et al., 1995). During adult hematopoiesis, Scl/Tal1 is highly expressed in LT-HSCs compared with short-term HSCs and progenitor cells (Lacombe et al., 2010). However, a study using conditional Scl/Tal1 knockout mice revealed that Scl/Tal1 is required for the generation of, but not the maintenance of, adult HSCs (Mikkola et al., 2003). Another group showed that conditional deletion of Scl/Tal1 in adult HSCs has a relatively mild effect: Scl-null HSCs show impaired short-term repopulating ability, but no defect in long-term repopulating capacity (Curtis et al., 2004). Redundant activity caused by the expression of Lyl1, a related bHLH transcription factor, in adult HSCs may provide an explanation for these "mild" phenotypes. While adult HSCs in single-knockout mice show no or only a mild phenotype, Lyl1;Scl-conditional double-knockout mice show a gene dosage defect on HSC survival, as HSCs and progenitor

Recently, Lacombe et al. demonstrated that Scl/Tal1 is required for the maintenance of the quiescent stem cell pool (Lacombe et al., 2010). Cell-cycle analyses revealed that Scl/Tal1 negatively regulates the G0-G1 transit of LT-HSCs; however, these phenomena were specific to adult HSCs and were not observed in perinatal HSCs. The reconstituting ability of *Scl+/-* HSCs or HSCs with decreased Scl protein expression induced by RNA interference was impaired in various transplantation assays. Furthermore, gene expression analysis and chromatin immunoprecipitation experiments revealed that the Cdkn1a and Id1 genes are

The transcription factor Tel (also known as Etv6), an Ets-related transcriptional repressor, is a frequent target of the diverse chromosomal translocations observed in leukemias (Golub et al., 1994). Tel/ETV6 is also required for HSC survival in adult hematopoiesis. Following conditional inactivation of Tel/Etv6, HSCs are rapidly lost from the adult BM. However, Tel/Etv6 is not required for the maintenance of lineage-committed progenitors. Conditional deletion of Tel/Etv6 after lineage commitment does not affect the differentiation or survival of these progenitors, although it does impair the maturation of megakaryocytes (Hock et al., 2004).

on Mcl-1 defines the human stem cell hierarchy (Campbell et al., 2010).

cells are immediately lost due to apoptosis (Souroullas et al., 2009).

**2.2.1 Bcl-2 family** 

**2.2.2 Scl, Lyl1** 

direct SCL targets.

**2.2.3 Tel/Etv6** 

death plays a role in regulating HSC homeostasis.

Zfx is a zinc finger protein belonging to the Zfx/ZFy family. Mammalian Zfx is encoded on the X chromosome and contains an acidic transcriptional activation domain, a nuclear localization sequence, and a DNA binding protein domain consisting of 13 C2H2-type zinc fingers (Schneider-Gadicke et al., 1989). Zfx is highly expressed in both HSCs and undifferentiated embryonic stem cells (ESCs). Using conditional gene targeting, Zfx was identified as an essential transcriptional regulator of HSC function (Galan-Caridad et al., 2007). Constitutive or inducible deletion of Zfx in HSCs (using *Tie2-Cre* and *Mx1-Cre* deletion strains, respectively) impairs self-renewal, resulting in increased apoptosis and the upregulation of stress-inducible genes.
