RESUMO
Recent studies have established that during embryonic development, hematopoietic progenitors and stem cells are generated from hemogenic endothelium precursors through a process termed endothelial to hematopoietic transition (EHT). The transcription factor RUNX1 is essential for this process, but its main downstream effectors remain largely unknown. Here, we report the identification of Gfi1 and Gfi1b as direct targets of RUNX1 and critical regulators of EHT. GFI1 and GFI1B are able to trigger, in the absence of RUNX1, the down-regulation of endothelial markers and the formation of round cells, a morphologic change characteristic of EHT. Conversely, blood progenitors in Gfi1- and Gfi1b-deficient embryos maintain the expression of endothelial genes. Moreover, those cells are not released from the yolk sac and disseminated into embryonic tissues. Taken together, our findings demonstrate a critical and specific role of the GFI1 transcription factors in the first steps of the process leading to the generation of hematopoietic progenitors from hemogenic endothelium.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Hemangioblastos/citologia , Hemangioblastos/fisiologia , Hematopoese/fisiologia , Proteínas Proto-Oncogênicas/fisiologia , Proteínas Repressoras/fisiologia , Fatores de Transcrição/fisiologia , Animais , Diferenciação Celular , Linhagem Celular , Subunidade alfa 2 de Fator de Ligação ao Core/deficiência , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/fisiologia , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/fisiologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos , Camundongos Knockout , Modelos Biológicos , Neovascularização Fisiológica , Gravidez , Proteínas Proto-Oncogênicas/deficiência , Proteínas Proto-Oncogênicas/genética , Proteínas Repressoras/deficiência , Proteínas Repressoras/genética , Fatores de Transcrição/deficiência , Fatores de Transcrição/genética , Saco Vitelino/citologia , Saco Vitelino/embriologia , Saco Vitelino/fisiologiaRESUMO
The endothelial to haematopoietic transition (EHT) is a key developmental process where a drastic change of endothelial cell morphology leads to the formation of blood stem and progenitor cells during embryogenesis. As TGFß signalling triggers a similar event during embryonic development called epithelial to mesenchymal transition (EMT), we hypothesised that TGFß activity could play a similar role in EHT as well. We used the mouse embryonic stem cell differentiation system for in vitro recapitulation of EHT and performed gain and loss of function analyses of the TGFß pathway. Quantitative proteomics analysis showed that TGFß treatment during EHT increased the secretion of several proteins linked to the vascular lineage. Live cell imaging showed that TGFß blocked the formation of round blood cells. Using gene expression profiling we demonstrated that the TGFß signalling activation decreased haematopoietic genes expression and increased the transcription of endothelial and extracellular matrix genes as well as EMT markers. Finally we found that the expression of the transcription factor Sox17 was up-regulated upon TGFß signalling activation and showed that its overexpression was enough to block blood cell formation. In conclusion we showed that triggering the TGFß pathway does not enhance EHT as we hypothesised but instead impairs it.