RESUMO
It is critical to understand how human quiescent long-term hematopoietic stem cells (LT-HSCs) sense demand from daily and stress-mediated cues and then transition into bioenergetically active progeny to differentiate and meet these cellular needs. However, the demand-adapted regulatory circuits of these early steps of hematopoiesis are largely unknown. Here we show that lysosomes, sophisticated nutrient-sensing and signaling centers, are regulated dichotomously by transcription factor EB (TFEB) and MYC to balance catabolic and anabolic processes required for activating LT-HSCs and guiding their lineage fate. TFEB-mediated induction of the endolysosomal pathway causes membrane receptor degradation, limiting LT-HSC metabolic and mitogenic activation, promoting quiescence and self-renewal, and governing erythroid-myeloid commitment. In contrast, MYC engages biosynthetic processes while repressing lysosomal catabolism, driving LT-HSC activation. Our study identifies TFEB-mediated control of lysosomal activity as a central regulatory hub for proper and coordinated stem cell fate determination.
Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos , Hematopoese , Células-Tronco Hematopoéticas , Diferenciação Celular , Células-Tronco Hematopoéticas/citologia , Humanos , Lisossomos , Transdução de SinaisRESUMO
Children with Down syndrome have a 150-fold increased risk of developing myeloid leukemia, but the mechanism of predisposition is unclear. Because Down syndrome leukemogenesis initiates during fetal development, we characterized the cellular and developmental context of preleukemic initiation and leukemic progression using gene editing in human disomic and trisomic fetal hematopoietic cells and xenotransplantation. GATA binding protein 1 (GATA1) mutations caused transient preleukemia when introduced into trisomy 21 long-term hematopoietic stem cells, where a subset of chromosome 21 microRNAs affected predisposition to preleukemia. By contrast, progression to leukemia was independent of trisomy 21 and originated in various stem and progenitor cells through additional mutations in cohesin genes. CD117+/KIT proto-oncogene (KIT) cells mediated the propagation of preleukemia and leukemia, and KIT inhibition targeted preleukemic stem cells.
Assuntos
Proteínas de Ciclo Celular/genética , Síndrome de Down/genética , Fator de Transcrição GATA1/genética , Células-Tronco Hematopoéticas/fisiologia , Leucemia Mieloide/genética , Pré-Leucemia/genética , Animais , Antígenos CD34/análise , Proteínas de Ciclo Celular/metabolismo , Linhagem da Célula , Proliferação de Células , Transformação Celular Neoplásica , Proteínas Cromossômicas não Histona/genética , Cromossomos Humanos Par 21/genética , Cromossomos Humanos Par 21/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Síndrome de Down/complicações , Feminino , Fator de Transcrição GATA1/metabolismo , Hematopoese , Transplante de Células-Tronco Hematopoéticas , Xenoenxertos , Humanos , Leucemia Mieloide/metabolismo , Leucemia Mieloide/patologia , Fígado/embriologia , Masculino , Megacariócitos/fisiologia , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Mutação , Pré-Leucemia/metabolismo , Pré-Leucemia/patologia , Inibidores de Proteínas Quinases/farmacologia , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas c-kit/análise , Proteínas Proto-Oncogênicas c-kit/antagonistas & inibidores , CoesinasRESUMO
In the human hematopoietic system, rare self-renewing multipotent long-term hematopoietic stem cells (LT-HSCs) are responsible for the lifelong production of mature blood cells and are the rational target for clinical regenerative therapies. However, the heterogeneity in the hematopoietic stem cell compartment and variable outcomes of CRISPR/Cas9 editing make functional interrogation of rare LT-HSCs challenging. Here, we report high efficiency LT-HSC editing at single-cell resolution using electroporation of modified synthetic gRNAs and Cas9 protein. Targeted short isoform expression of the GATA1 transcription factor elicit distinct differentiation and proliferation effects in single highly purified LT-HSC when analyzed with functional in vitro differentiation and long-term repopulation xenotransplantation assays. Our method represents a blueprint for systematic genetic analysis of complex tissue hierarchies at single-cell resolution.
