Mecanismos de regulación de la expresión del gen evi1 y análisis funcional de su promotor en leucemia mieloide aguda

Resumen

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). The only known mechanisms that lead to EVI1 overexpression are 3q aberrations, and the MLL-ENL fusion protein. We have studied the promoter region of EVI1. We have demonstrated that an aberrant epigenetic pattern play a role in the transcriptional regulation of this gene in AML. Cell lines with overexpression of EVI1 have no DNA methylation in the promoter region of EVI1 locus, and have 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. Furthermore, we have characterized a minimal functional promoter region of 318 bp close to the transcriptional start site of EVI1. In this region we have identified binding sites for transcription factors with important roles in hematopoiesis, confirming that RUNX1 and ELK1 bind to this region regulating the transcription of EVI1 in AML. Moreover, we found that RUNX1 regulates the transcription of EVI1 through the acetylation of the histone 3 in its promoter region. A positive correlation between the expression of EVI1 and RUNX1 in a series of AML patients confirmed the clinical relevance of these findings. Moreover, we found that EVI1 could regulate its own transcription. The different EVI1 isoforms: EVI1-RP-9, ∆324, and MDS1EVI1 repress its own transcription, while the full length protein EVI1-145kDa activates it. Finally, functional experiments showed that the EVI1 isoforms have different functional roles in cell transformation: EVI1-145kDa and EVI1-RP-9 increase cell viability reducing the number of apoptotic cells, ∆324 has no effects, and MDS1EVI1 induces cell apoptosis. Our results support the complex functional regulation of the EVI1 locus and open new directions to further understand the mechanisms of EVI1 overexpressing leukemias.