Evi-1 has been recognized as one of the dominant oncogenes associated with murine and human myeloid leukemia. Here, we show that hematopoietic stem cells (HSCs) in Evi-1-deficient embryos are severely reduced in number with defective proliferative and repopulating capacity. Selective ablation of Evi-1 in Tie2(+) cells mimics Evi-1 deficiency, suggesting that Evi-1 function is required in Tie2(+) hematopoietic stem/progenitors. Conditional deletion of Evi-1 in the adult hematopoietic system revealed that Evi-1-deficient bone marrow HSCs cannot maintain hematopoiesis and lose their repopulating ability. In contrast, Evi-1 is dispensable for blood cell lineage commitment. Evi-1(+/-) mice exhibit the intermediate phenotype for HSC activity, suggesting a gene dosage requirement for Evi-1. We further demonstrate that disruption of Evi-1 in transformed leukemic cells leads to significant loss of their proliferative activity both in vitro and in vivo. Thus, Evi-1 is a common and critical regulator essential for proliferation of embryonic/adult HSCs and transformed leukemic cells.

A new mouse in which an IRES-GFP cassette is knocked-in to the Evi1 locus reveals that HSC long-term multilineage repopulating activity specifically segregates with expression of the Evi1 transcription factor.

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Ecotropic viral integration site-1 (Evi-1) is a nuclear transcription factor that plays an essential role in the regulation of hematopoietic stem cells. Aberrant expression of Evi-1 has been reported in up to 10% of patients with acute myeloid leukemia and is a diagnostic marker that predicts a poor outcome. Although chromosomal rearrangement involving the Evi-1 gene is one of the major causes of Evi-1 activation, overexpression of Evi-1 is detected in a subgroup of acute myeloid leukemia patients without any chromosomal abnormalities, which indicates the presence of other mechanisms for Evi-1 activation. In this study, we found that Evi-1 is frequently up-regulated in bone marrow cells transformed by the mixed-lineage leukemia ( IntroductionThe ecotropic viral integration site-1 (Evi-1) is a nuclear transcription factor that plays an essential role in the proliferation and maintenance of hematopoietic stem cells (HSCs). 1-3 There are 2 major alternative forms generated from the Evi-1 gene, Evi-1a and Mds1-Evi-1 (also called Evi-1c). Mds1-Evi-1 is a fusion variant of Evi-1 generated through intergenic splicing with Mds1, 4 a gene located approximately 140 and 500 kb upstream of Evi-1 in the human and mouse genome, respectively. In contrast to Evi-1a, Mds1-Evi-1 possesses the PRDI-BF1-RIZ1 homologous (PR) domain in the N-terminus, which regulates oligomerization of the Evi-1 proteins. 5 Both Evi-1a and Mds1-Evi-1 are normally coexpressed in several developing and adult tissues, 6 and differences in the normal function between these proteins remain to be elucidated. Like all other PR domain proteins, Evi-1 contains several zinc finger motifs. They are grouped into N-terminal 7 and C-terminal 3 clusters, which are called the first and second zinc finger domain, respectively. 7,8 Between these 2 zinc finger domains lie the C-terminal binding protein (CtBP) domain and the repression domain. The first zinc finger, the repression, and the CtBP-binding domains exhibit a growth-promoting effect by blocking transforming growth factor-␤ signaling. 9 The first zinc finger domain also exhibits an antiapoptotic effect by repressing c-Jun N-terminal kinase signaling. 10 The second zinc finger domain stimulates proliferation by increasing activator protein-1 activity. 11 Thus, Evi-1 possesses diverse functions as an oncoprotein.Aberrant expression of EVI-1 frequently has been found in myeloid leukemia and in several solid tumors and is associated with poor prognosis of patients with leukemia. 12-15 Rearrangements of chromosome 3q26, which contains the EVI-1 gene, lead to overexpression of EVI-1 and are implicated in the development or progression of high-risk acute myeloid leukemia (AML). 16 Importantly, EVI-1 is also highly expressed in a subgroup of AML patients without 3q26 rearrangements, 12 which indicates the presence of other mechanisms of EVI-1 activation. Recently, several clinical studies revealed a positive correlation between EVI-1 (both EVI-1a and MDS1-EVI-1) overexpression and rearrangements of the mixed-lineage ...

The ecotropic viral integration site-1 (EVI-1) is a nuclear transcription factor and has an essential function in the proliferation/maintenance of haematopoietic stem cells. Aberrant expression of EVI-1 has been frequently found in myeloid leukaemia as well as in several solid tumours, and is associated with a poor patient survival. It was recently shown that EVI-1 associates with two different histone methyltransferases (HMTs), SUV39H1 and G9a. However, the functional roles of these HMTs in EVI-1-mediated leukemogenesis remain unclear. In this study, we showed that EVI-1 physically interacts with SUV39H1 and G9a, but not with Set9. Immunofluorescence analysis revealed that EVI-1 colocalizes with these HMTs in nuclei. We also found that the catalytically inactive form of SUV39H1 abrogates the transcriptional repression mediated by EVI-1, suggesting that SUV39H1 is actively involved in EVI-1-mediated transcriptional repression. Furthermore, RNAi-based knockdown of SUV39H1 or G9a in Evi-1-expressing progenitors significantly reduced their colony-forming activity. In contrast, knockdown of these HMTs did not impair bone marrow immortalization by E2A/HLF. These results indicate that EVI-1 forms higher-order complexes with HMTs, and this association has a role in the transcription repression and bone marrow immortalization. Targeting these HMTs may be of therapeutic benefit in the treatment for EVI-1-related haematological malignancies.