In this study we investigated whether the Ednra signaling
In this study, we investigated whether the Ednra signaling is involved in Hoxa9/Meis1-mediated leukemogenesis. We found an upregulation of Ednra in leukemic murine bone marrow (BM) dmxaa overexpressing Hoxa9 and Meis1 compared to cells overexpressing Hoxa9 alone. We showed that overexpression of Ednra together with Hoxa9 shortens the survival of mice in comparison to mice transplanted with cells overexpressing Hoxa9 alone. Furthermore, overexpression of Ednra gives proliferation advantage as well as apoptosis resistance to the Hoxa9-expressing cells in vitro. We also showed that Meis1 binds to the Ednra promoter region. Our findings demonstrate a role for Ednra in the progression of Hoxa9/Meis1-induced leukemia in vivo as well as in proliferation and apoptosis in vitro.
Materials and methods
Discussion This study presents the first evidence that Ednra is upregulated in leukemic cells with concurrent high expression of Hoxa9 and Meis1. We showed that overexpression of Hoxa9 and Meis1 is associated with high expression of Ednra in murine BM model in vitro as well as in a leukemia transplantation model in vivo. In addition, our data indicated that Hoxa9/Meis1 regulates the expression of Ednra through direct binding of Meis1 to the Ednra regulatory region. Transplantation of Ednra expressing cells together with expression of Hoxa9 shortens the survival of mice when compared to mice transplanted with cells overexpressing Hoxa9 alone. Increased organ infiltration, enlarged spleen and augmented number of leukemic cells in mice BM led to the induction of leukemia similar to the disease induced by Hoxa9+Meis1 but with a slower onset. Finally, we showed that overexpression of Ednra together with Hoxa9 gives a proliferative advantage to cells and infers apoptosis resistance. Taken together our data suggest a functional role for Ednra in Hoxa9+Meis1-induced leukemia. The induction of leukemia by Hoxa9+Ednra underscores the possible importance of Ednra in the progression of leukemia. The later onset of the disease relative to Hoxa9+Meis1-induced leukemia, is most likely due to the fact that Ednra is only one of several other factors regulated by the Hoxa9/Meis1 complex. Ednra’s upregulation contributed to at least two important aspects of oncogenesis namely, enhanced cell proliferation and increased resistance to apoptosis of the cells. In order to activate signaling of Ednra as a receptor, its ligand Et-1is required . ET-1 is a known survival factor in a variety of normal and cancer cell types and acts mainly through EDNRA . Some studies have reported that ET-1 levels are elevated in solid tumors [26,27]. In our cell cultures (Hoxa9, Hoxa9+Meis1 and Hoxa9+Ednra), almost equal levels of secreted Et-1 was detected in the media collected from all transfected cells. Overexpression of Ednra led to increased proliferation and repopulating capacity of Hoxa9 expressing cells. Furthermore, resistance to apoptosis is critical for cancer progression and metastasis. In a variety of cancers, particularly in leukemia, it is important for leukemia inducing cells to be in close contact with their niche. This contact is supported and maintained by several surface molecules which modulate the homing capacity of leukemic cells as well as their proliferation and quiescence. These cells benefit from the niche microenvironment because a contact with the niche leads to protection of quiescent leukemia inducing cells the population which gives rise to chemo-resistant leukemic cells . Identification of such surface molecules mediating this effect may be helpful in targeting the residual leukemia inducing cells after therapy . Here, we evaluated whether Ednra signaling may infer protection against drug-induced apoptosis as reported in solid tumors. We used daunorubicin, a chemotherapeutic agent which is widely used in leukemia treatment to induce apoptosis in the cells. Our results suggest that Ednra upregulation may contribute to chemo-resistance in leukemic cells in response to daunorubicin. In this context, Ednra may be a functional signaling molecule both by stimulating cell growth and inducing protection against cytotoxic effects. Further studies will be necessary to investigate the mechanism of Ednra function using additional chemotherapeutic agents and Ednra knockdown model as a control.