RESUMO
The BCR-ABL oncoprotein exhibits deregulated protein tyrosine kinase activity and is implicated in the pathogenesis of Philadelphia chromosome (Ph)-positive human leukemias. Here, we report that ectopic expression of p210(BCR-ABL) in the megakaryoblastic Mo7e cell line and in primary human CD34(+) progenitors trigger erythroid differentiation at the expense of megakaryocyte (MK) differentiation. Clonal culture of purified CD41(+)CD42(-) cells, a population highly enriched in MK progenitors, combined with the conditional expression of p210(BCR-ABL) tyrosine kinase activity by imatinib identified a true lineage reprogramming. In both Mo7e or CD41(+)CD42(-) cells transduced with p210(BCR-ABL), lineage switching was associated with a downregulation of the friend leukemia Integration 1 (FLI-1) transcription factor. Re-expression of FLI-1 in p210(BCR-ABL)-transduced Mo7e cells rescued the megakaryoblastic phenotype. Altogether, these results demonstrate that alteration of signal transduction via p210(BCR-ABL) reprograms MK cells into erythroid cells by a downregulation of FLI-1. In addition, our findings underscore the role of kinases in lineage choice and infidelity in pathology and suggest that downregulation of FLI-1 may have important implications in CML pathogenesis.
Assuntos
Células Eritroides/citologia , Proteínas de Fusão bcr-abl/fisiologia , Células-Tronco Hematopoéticas/citologia , Megacariócitos/citologia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Diferenciação Celular , Regulação para Baixo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Transcrição Gênica , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genéticaRESUMO
Deregulation of transcriptional networks contributes to haematopoietic malignancies. The transcription factor Spi-1/PU.1 is a master regulator of haematopoiesis and its alteration leads to leukaemia. Spi-1 overexpression inhibits differentiation and promotes resistance to apoptosis in erythroleukaemia. Here, we show that Spi-1 inhibits mitochondrial apoptosis in vitro and in vivo through the transcriptional repression of Bim, a proapoptotic factor. BIM interacts with MCL-1 that behaves as a major player in the survival of the preleukaemic cells. The repression of BIM expression reduces the amount of BIM-MCL-1 complexes, thus increasing the fraction of potentially active antiapoptotic MCL-1. We then demonstrate that Spi-1 represses Bim transcription by binding to the Bim promoter and by promoting the trimethylation of histone 3 on lysine 27 (H3K27me3, a repressive histone mark) on the Bim promoter. The PRC2 repressive complex of Polycomb is directly responsible for the deposit of H3K27me3 mark at the Bim promoter. SUZ12 and the histone methyltransferase EZH2, two PRC2 subunits bind to the Bim promoter at the same location than H3K27me3, distinct of the Spi-1 DNA binding site. As Spi-1 interacts with SUZ12 and EZH2, these results indicate that Spi-1 modulates the activity of PRC2 without directly recruiting the complex to the site of its activity on the chromatin. Our results identify a new mechanism whereby Spi-1 represses transcription and provide mechanistic insights on the antiapoptotic function of a transcription factor mediated by the epigenetic control of gene expression.
Assuntos
Proteínas Reguladoras de Apoptose/genética , Apoptose/genética , Proteínas de Ligação a DNA/metabolismo , Leucemia Mieloide Aguda/genética , Proteínas de Membrana/genética , Proteínas Proto-Oncogênicas/genética , Animais , Proteína 11 Semelhante a Bcl-2 , Sítios de Ligação/genética , Diferenciação Celular/genética , Linhagem Celular , Metilação de DNA , Proteínas de Ligação a DNA/biossíntese , Proteína Potenciadora do Homólogo 2 de Zeste , Regulação Leucêmica da Expressão Gênica , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Camundongos , Camundongos Transgênicos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Complexos Multiproteicos/genética , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Complexo Repressor Polycomb 2/metabolismo , Regiões Promotoras Genéticas , Ligação Proteica , Interferência de RNA , RNA Interferente Pequeno , Transcrição Gênica/genéticaRESUMO
Human telomerase, a cellular reverse transcriptase (hTERT), is a nuclear ribonucleoprotein enzyme complex that catalyzes the synthesis and extension of telomeric DNA. This enzyme is specifically activated in most malignant tumors but is usually inactive in normal somatic cells, suggesting that telomerase plays an important role in cellular immortalization and tumorigenesis. Terminal maturation of tumor cells has been associated with the repression of telomerase activity. Using maturation-sensitive and -resistant NB4 cell lines, we analyzed the pattern of telomerase expression during the therapeutic treatment of acute promyelocytic leukemia (APL) by retinoids. Two pathways leading to the down-regulation of hTERT and telomerase activity were identified. The first pathway results in a rapid down-regulation of telomerase that is associated with retinoic acid receptor (RAR)-dependent maturation of NB4 cells. Furthermore, during NB4 cell maturation, obtained independently of RAR by retinoic X receptor (RXR)-specific agonists (rexinoids), no change in telomerase activity was observed, suggesting that hTERT regulation requires a specific signaling and occurs autonomously. A second pathway of hTERT regulation, identified in the RAR-responsive, maturation-resistant NB4-R1 cell line, results in a down-regulation of telomerase that develops slowly during two weeks of all-trans retinoic acid (ATRA) treatment. This pathway leads to telomere shortening, growth arrest, and cell death, all events that are overcome by ectopic expression of hTERT. These findings demonstrate a clear and full dissociation between the process of tumor cell maturation and the regulation of hTERT mRNA expression and telomerase activity by retinoids. We propose telomerase expression as an efficient and selective target of retinoids in the therapy of tumors.