Open Access
Overexpression of GATA2 predicts an adverse prognosis for patients with acute myeloid leukemia and it is associated with distinct molecular abnormalities
(Springer Nature, 2012) Vicente, Carmen; Vázquez Urio, Iria; Conchillo, Ana; García-Sánchez, M. A.; Marcotegui, Nerea; Fuster, Óscar; González, Marcos; Calasanz, María José; Lahortiga, Idoya; Odero, María D.; Ciencias; Zientziak; © 2011 Springer Nature Limited.
Acute myeloid leukemia (AML) represents a heterogeneous group of hematological neoplasms associated with the accumulation of acquired genetic and epigenetic aberrations in hematopoietic progenitor cells. Myeloid leukemogenesis is directly linked to the disruption of expression of transcription factors that regulate the proliferation, survival and differentiation of myeloid progenitors. Moreover, overexpression of particular genes has been associated with prognosis in AML, enabling patients with different survival rates to be identified within cytogenetic risk groups. The GATA2 gene encodes a transcription factor that mediates essential functions in hematopoietic stem cell and progenitor cell (HSC/HPC) compartments. The essential role of GATA2 in hematopoiesis is evident in mice lacking GATA2, which are anemic, have fewer HSC/HPC cells and that die by day 10–11 of gestation.
Open Access
EVI1 controls proliferation in acute myeloid leukaemia through modulation of miR-1-2
(Nature Publishing Group, 2010-09-14) Gómez-Benito, María; Conchillo, Ana; García, M. A.; Vázquez Urio, Iria; Maicas, Miren; Vicente, Carmen; Cristobal, I.; Marcotegui, Nerea; García-Ortí, L.; Bandrés Elizalde, Eva; Calasanz, María José ; Alonso, M. M.; Odero, María D.; Ciencias; Zientziak; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua
Bakground: the EVI1(ecotropic virus integration site 1) gene codes for a zinc-finger transcription factor, whose transcriptional activation leads to a particularly aggressive form of acute myeloid leukaemia (AML). Although, EVI1 interactions with key proteins in hematopoiesis have been previously described, the precise role of this transcription factor in promoting leukaemic transformation is not completely understood. Recent works have identified specific microRNA (miRNA) signatures in different AML subgroups. However, there is no analysis of miRNAs profiles associated with EVI1 overexpression in humans. Methods: we performed QT-RT-PCR to assess the expression of 250 miRNAs in cell lines with or without EVI1 overexpression and in patient samples. We used ChIP assays to evaluated the possible binding of EVI1 binding to the putative miRNA promoter. Proliferation of the different cell lines transfected with the anti- or pre-miRs was quantified by MTT. Results: our data showed that EVI1 expression was significantly correlated with the expression of miR-1-2 and miR-133-a-1 in established cell lines and in patient samples. ChIP assays confirmed that EVI1 binds directly to the promoter of these two miRNAs. However, only miR-1-2 was involved in abnormal proliferation in EVI1 expressing cell lines. Conclusions: our data showed that EVI1 controls proliferation in AML through modulation of miR-1-2. This study contributes to further understand the transcriptional networks involving transcription factors and miRNAs in AML.
Open Access
Silencing of hsa-miR-124 by EVI1 in cell lines and patients with acute myeloid leukemia
(National Academy of Sciences, 2010-01-07) Vázquez Urio, Iria; Maicas, Miren; Marcotegui, Nerea; Conchillo, Ana; Guruceaga, Elizabeth; Román-Gómez, José; Calasanz, María José; Agirre, Xabier; Prósper, Felipe; Odero, María D.; Ciencias; Zientziak
We read with great interest the work published by Dickstein et al. (1) showing that induced EVI1 expression in a murine model silences miR-124 expression by DNA methylation. The EVI1 gene codes for a transcription factor implicated in the development and progression of high-risk acute myeloid leukemia (AML) (2, 3). We quantify the expression of 250 microRNAs (miRNAs; TaqManHuman miRNA assay set) in 15 myeloid cell lines. Statistical analysis comparing cell lines with and without EVI1 protein identified miRNAs differentially expressed (B > 0)
Open Access
Down-regulation of EVI1 is associated with epigenetic alterations and good prognosis in patients with acute myeloid leukemia
(Ferrata Storti Foundation, 2012-10-18) Vázquez Urio, Iria; Maicas, Miren; Cervera, José; Agirre, Xabier; Marin-Béjar, Oskar; Marcotegui, Nerea; Vicente, Carmen; Lahortiga, Idoya; Gómez-Benito, María; Carranza, Claudia; Valencia, Ana; Brunet, Salut; Lumbreras, Eva; Prósper, Felipe; Gómez-Casares, Teresa; Hernández-Rivas, Jesús M.; Calasanz, María José; Sanz, Miguel A.; Sierra, Jorge; Odero, María D.; Ciencias; Zientziak; Gobierno de Navarra / Nafarroako Gobernua
Background The EVI1 gene (3q26) codes for a zinc finger transcription factor with important roles in both mammalian development and leukemogenesis. Over-expression of EVI1 through either 3q26 rearrangements, MLL fusions, or other unknown mechanisms confers a poor prognosis in acute myeloid leukemia. Design and Methods We analyzed the prevalence and prognostic impact of EVI1 over-expression in a series of 476 patients with acute myeloid leukemia, and investigated the epigenetic modifications of the EVI1 locus which could be involved in the transcriptional regulation of this gene. Results Our data provide further evidence that EVI1 over-expression is a poor prognostic marker in acute myeloid leukemia patients less than 65 years old. Moreover, we found that patients with no basal expression of EVI1 had a better prognosis than patients with expression/over-expression (P=0.036). We also showed that cell lines with over-expression of EVI1 had no DNA methylation in the promoter region of the EVI1 locus, and had marks of active histone modifications: H3 and H4 acetylation, and trimethylation of histone H3 lysine 4. Conversely, cell lines with no expression of EVI1 have DNA hypermethylation and are marked by repressive trimethylation of histone H3 lysine 27 at the EVI1 promoter. Conclusions Our results identify EVI1 over-expression as a poor prognostic marker in a large, independent cohort of acute myeloid leukemia patients less than 65 years old, and show that the total absence of EVI1 expression has a prognostic impact on the outcome of such patients. Furthermore, we demonstrated for the first time that an aberrant epigenetic pattern involving DNA methylation, H3 and H4 acetylation, and trimethylation of histone H3 lysine 4 and histone H3 lysine 27 might play a role in the transcriptional regulation of EVI1 in acute myeloid leukemia. This study opens new avenues for a better understanding of the regulation of EVI1 expression at a transcriptional level.
Open Access
Functional characterization of the promoter region of the human EVI1 gene in acute myeloid leukemia: RUNX1 and ELK1 directly regulate its transcription
(Macmillan Press Ltd, 2012-06-11) Maicas, Miren; Vázquez Urio, Iria; Vicente, Carmen; García-Sánchez, M. A.; Marcotegui, Nerea; Urquiza, L.; Calasanz, María José; Odero, María D.; Ciencias; Zientziak; Gobierno de Navarra / Nafarroako Gobernua
The EVI1 gene (3q26) codes for a transcription factor with important roles in normal hematopoiesis and leukemogenesis. High expression of EVI1 is a negative prognostic indicator of survival in acute myeloid leukemia (AML) irrespective of the presence of 3q26 rearrangements. However, the only known mechanisms that lead to EVI1 overexpression are 3q aberrations, and the MLL-ENL oncoprotein, which activates the transcription of EVI1 in hematopoietic stem cells. Our aim was to characterize the functional promoter region of EVI1, and to identify transcription factors involved in the regulation of this gene. Generation of seven truncated constructs and luciferase reporter assays allowed us to determine a 318-bp region as the minimal promoter region of EVI1. Site-directed mutagenesis and chromatin immunoprecipitation (ChIP) assays identified RUNX1 and ELK1 as putative transcription factors of EVI1. Furthermore, knockdown of RUNX1 and ELK1 led to EVI1 downregulation, and their overexpression to upregulation of EVI1. Interestingly, in a series of patient samples with AML at diagnosis, we found a significant positive correlation between EVI1 and RUNX1 at protein level. Moreover, we identified one of the roles of RUNX1 in the activation of EVI1 during megakaryocytic differentiation. EVI1 knockdown significantly inhibited the expression of megakaryocytic markers after treating K562 cells with TPA, as happens when knocking down RUNX1. In conclusion, we define the minimal promoter region of EVI1 and demonstrate that RUNX1 and ELK1, two proteins with essential functions in hematopoiesis, regulate EVI1 in AML. Furthermore, our results show that one of the mechanisms by which RUNX1 regulates the transcription of EVI1 is by acetylation of the histone H3 on its promoter region. This study opens new directions to further understand the mechanisms of EVI1 overexpressing leukemias.