RESUMO
Extensive molecular profiling of leukemias and preleukemic diseases has revealed that distinct clinical entities, like acute myeloid (AML) and T-lymphoblastic leukemia (T-ALL), share similar pathogenetic mutations. It is not well understood how the cell of origin, accompanying mutations, extracellular signals or structural differences in a mutated gene determine the phenotypic identity of leukemias. We dissected the functional aspects of different protein regions of the MN1 oncogene and their effect on the leukemic phenotype, building on the ability of MN1 to induce leukemia without accompanying mutations. We found that the most C-terminal region of MN1 was required to block myeloid differentiation at an early stage, and deletion of an extended C-terminal region resulted in loss of myeloid identity and cell differentiation along the T-cell lineage in vivo. Megakaryocytic/erythroid lineage differentiation was blocked by the N-terminal region. In addition, the N-terminus was required for proliferation and leukemogenesis in vitro and in vivo through upregulation of HoxA9, HoxA10 and Meis2. Our results provide evidence that a single oncogene can modulate cellular identity of leukemic cells based on its active gene regions. It is therefore likely that different mutations in the same oncogene may impact cell fate decisions and phenotypic appearance of malignant diseases.
Assuntos
Hematopoese/genética , Leucemia/genética , Leucemia/patologia , Fenótipo , Proteínas Supressoras de Tumor/genética , Animais , Células da Medula Óssea/metabolismo , Células da Medula Óssea/patologia , Diferenciação Celular/genética , Transformação Celular Neoplásica/genética , Análise por Conglomerados , Perfilação da Expressão Gênica , Humanos , Leucemia/metabolismo , Células Progenitoras de Megacariócitos e Eritrócitos/metabolismo , Células Progenitoras de Megacariócitos e Eritrócitos/patologia , Camundongos , Mutação , Células Mieloides/metabolismo , Células Mieloides/patologia , Domínios e Motivos de Interação entre Proteínas/genética , Linfócitos T/metabolismo , Linfócitos T/patologia , Transativadores , Proteínas Supressoras de Tumor/química , Proteínas Supressoras de Tumor/metabolismoRESUMO
Pathways defining susceptibility of normal cells to oncogenic transformation may be valuable therapeutic targets. We characterized the cell of origin and its critical pathways in MN1-induced leukemias. Common myeloid (CMP) but not granulocyte-macrophage progenitors (GMP) could be transformed by MN1. Complementation studies of CMP-signature genes in GMPs demonstrated that MN1-leukemogenicity required the MEIS1/AbdB-like HOX-protein complex. ChIP-sequencing identified common target genes of MN1 and MEIS1 and demonstrated identical binding sites for a large proportion of their chromatin targets. Transcriptional repression of MEIS1 targets in established MN1 leukemias demonstrated antileukemic activity. As MN1 relies on but cannot activate expression of MEIS1/AbdB-like HOX proteins, transcriptional activity of these genes determines cellular susceptibility to MN1-induced transformation and may represent a promising therapeutic target.
Assuntos
Transformação Celular Neoplásica/metabolismo , Transformação Celular Neoplásica/patologia , Proteínas de Homeodomínio/metabolismo , Leucemia Mieloide Aguda/patologia , Complexos Multiproteicos/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Perfilação da Expressão Gênica , Regulação Leucêmica da Expressão Gênica , Genes Dominantes/genética , Células Progenitoras de Granulócitos e Macrófagos/metabolismo , Células Progenitoras de Granulócitos e Macrófagos/patologia , Humanos , Leucemia Mieloide Aguda/genética , Camundongos , Camundongos Endogâmicos C57BL , Modelos Biológicos , Proteína Meis1 , Regiões Promotoras Genéticas/genética , Ligação ProteicaRESUMO
To determine the contribution of the common N-terminal truncation of NUP98 in NUP98-translocations resulting in acute myeloid leukemia, we have conducted a structure-function analysis of NUP98 in the context of NUP98-HOXA10HD, a novel, canonical NUP98-Hox fusion that significantly enhances the self-renewal capacity of hematopoietic stem cells and collaborates with Meis1 to induce AML in our mouse models. Our results identify that NUP98 functions by transcriptional activation likely by recruitment of CBP/p300 via its FG/GLFG repeats. In contrast, the functional interaction of NUP98 with Rae1 or the anaphase promoting complex appears non-essential for its role in NUP98-leukemogenic fusions.
Assuntos
Proteínas de Homeodomínio/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Doença Aguda , Sequência de Aminoácidos , Animais , Sítios de Ligação , Células da Medula Óssea/metabolismo , Transformação Celular Neoplásica/genética , Células Cultivadas , Feminino , Proteínas Homeobox A10 , Proteínas de Homeodomínio/genética , Estimativa de Kaplan-Meier , Leucemia Mieloide/genética , Leucemia Mieloide/metabolismo , Leucemia Mieloide/patologia , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Proteína Meis1 , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Proteínas Associadas à Matriz Nuclear/genética , Proteínas Associadas à Matriz Nuclear/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/genética , Proteínas de Transporte Nucleocitoplasmático/genética , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Proteínas de Fusão Oncogênica/genética , Ligação Proteica , Sequências Repetitivas de Aminoácidos , Fatores de Transcrição de p300-CBP/genética , Fatores de Transcrição de p300-CBP/metabolismoRESUMO
Although the cancer stem cell (CSC) concept implies that CSCs are rare, recent reports suggest that CSCs may be frequent in some cancers. We hypothesized that the proportion of leukemia stem cells would vary as a function of the number of dysregulated pathways. Constitutive expression of MN1 served as a 1-oncogene model, and coexpression of MN1 and a HOX gene served as a 2-oncogene model. Leukemia-initiating cell (LIC) number and in vitro expansion potential of LICs were functionally assessed by limiting dilution analyses. LIC expansion potential was 132-fold increased in the 2- compared with the 1-oncogene model, although phenotypically, both leukemias were similar. The 2-oncogene model was characterized by granulocyte-macrophage colony-stimulating factor (GM-CSF) hypersensitivity and activated STAT/ERK signaling. GM-CSF hypersensitivity of the 2-oncogene model (MN1/HOXA9) was lost in Stat5b(-/-) cells, and the LIC expansion potential was reduced by 86- and 28-fold in Stat5b(-/-) and Stat1(-/-) cells, respectively. Interestingly, in 201 acute myeloid leukemia (AML) patients, coexpression of MN1 and HOXA9 was restricted to patients with the poorest prognosis and was associated with highly active STAT signaling. Our data demonstrate the functional heterogeneity of LICs and show that STAT signaling is critical for leukemia stem cell self-renewal in MN1- and HOXA9-expressing leukemias.
Assuntos
Leucemia Experimental/metabolismo , Leucemia Experimental/patologia , Modelos Biológicos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fator de Transcrição STAT5/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Técnicas In Vitro , Leucemia Experimental/genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Células-Tronco Neoplásicas/efeitos dos fármacos , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/metabolismo , Oncogenes , Proteínas Recombinantes , Fator de Transcrição STAT1/deficiência , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT5/deficiência , Fator de Transcrição STAT5/genética , Transdução de Sinais/efeitos dos fármacos , Transativadores , Proteínas Supressoras de TumorAssuntos
Fator Estimulador de Colônias de Granulócitos/administração & dosagem , Leucemia/terapia , Receptores CXCR4/antagonistas & inibidores , Condicionamento Pré-Transplante/métodos , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Benzilaminas , Transplante de Medula Óssea , Movimento Celular/efeitos dos fármacos , Movimento Celular/fisiologia , Ciclamos , Esquema de Medicação , Compostos Heterocíclicos/farmacologia , Compostos Heterocíclicos/uso terapêutico , Humanos , Camundongos , Terapia Neoadjuvante/métodos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologiaRESUMO
MLL5 is a divergent member of the Drosophila Trithorax-related (SET) domain and plant homeodomain (PHD) domain-containing chromatin regulators that are involved in the regulation of transcriptional "memory" during differentiation. Human MLL5 is located on chromosome 7q22, which frequently is deleted in myeloid leukemias, suggesting a possible role in hemopoiesis. To address this question, we generated a loss-of-function allele (Mll5(tm1Apa)) in the murine Mll5 locus. Unlike other Mll genes, Mll5(tm1Apa) homozygous mice are viable but display defects in immunity and hematopoiesis. First, Mll5(tm1Apa) homozygous mice show increased susceptibility to spontaneous eye infections, associated with a cell-autonomous impairment of neutrophil function. Second, Mll5(tm1Apa/tm1Apa) mice exhibit a mild impairment of erythropoiesis. Third, Mll5(tm1Apa/tm1Apa) hematopoietic stem cells (HSCs) have impaired competitive repopulating capacity both under normal conditions and when subjected to self-renewal stimulation by NUP98-HOXA10. Fourth, Mll5(tm1Apa) homozygous HSCs show a dramatic sensitivity to DNA demethylation-induced differentiation (5-azadeoxycytidine). Taken together, our data show that MLL5 is involved in terminal myeloid differentiation and the regulation of HSC self-renewal by a mechanism that involves DNA methylation. These data warrant investigation of MLL5 expression levels as a predictive marker of demethylating-agent response in patients with myelodysplastic syndromes and leukemias and identify MLL5 as a key regulator of normal hematopoiesis.
