Project description:To facilitate comparative genomic analyses of human fetal and adult cells undergoing erythropoiesis, we employed a serum-free two-phase liquid culture system to expand and differentiate primary human CD34+ hematopoietic stem/progenitor cells (HSPCs) ex vivo. In this experimental context, highly enriched populations of stage-matched, differentiating, primary proerythroblasts (ProEs) were generated. We selected four time points (day 0, CD34+ HSPCs; day 3, 5, and 7, differentiating ProEs) that represented similar stages differentiation for gene expression profiling using microarrays. Primary maturing fetal or adult erythroblasts were generated ex vivo from CD34+ hematopoietic stem/progenitor cells (HSPCs) using a serum-free two-phase liquid culture system. Total RNA from primary fetal and adult HSPCs (day 0) and differentiating proerythroblasts (ProEs; day 3, 5, and 7) were extracted and used to hybridize to Affymetrix expression arrays using the HG-U133 Plus 2.0 platform.
Project description:Advances in sequencing-based genomic profiling present a new challenge of explaining how changes in DNA/RNA are translated into proteins linking genotypes to phenotypes. The developing erythroid cells require highly coordinated gene expression and metabolism, and serve as a unique model in dissecting regulatory events in development and disease. Here we compare the proteomic and transcriptomic changes in human hematopoietic stem/progenitor cells and lineage-committed erythroid progenitors, and uncover pathways related to mitochondrial biogenesis enhanced through post-transcriptional regulation. Two principal mitochondrial factors TFAM and PHB2 are tightly regulated at the protein level and indispensable for mitochondria and erythropoiesis. mTORC1 signaling is progressively enhanced to promote translation of mitochondrial proteins during erythroid specification. Genetic and pharmacological perturbation of mTORC1 or mitochondria impairs erythropoiesis. Our studies suggest a new mechanism for regulation of mitochondrial biogenesis through mTORC1-mediated protein translation, and may have direct relevance to the hematological defects associated with mitochondrial diseases and aging. Transcriptional profiling in human primary fetal and adult CD34+ hematopoietic stem/progenitor cells (HSPCs) erythroid progenitor cells (ProEs) by RNA-seq analysis.
Project description:We describe the proteomic composition of the extracellular environment of fetal and adult hematopoietic progenitors by data-independent acquisition mass spectrometry analysis.
Project description:We describe the proteomic composition of the secretome of fetal and adult hematopoietic progenitors during MLL-rearranged (MLLr) leukemia initiation as well as the pre-leukemic cells’ secretion response to the treatment with Fibulin (Fbln1) and/or Fibronectin (Fn1) using data-independent acquisition mass spectrometry analysis.
Project description:To facilitate comparative genomic analyses of human fetal and adult cells undergoing erythropoiesis, we employed a serum-free two-phase liquid culture system to expand and differentiate primary human CD34+ hematopoietic stem/progenitor cells (HSPCs) ex vivo. In this experimental context, highly enriched populations of stage-matched, differentiating, primary proerythroblasts (ProEs) were generated. We selected four time points (day 0, CD34+ HSPCs; day 3, 5, and 7, differentiating ProEs) that represented similar stages differentiation for gene expression profiling using microarrays.
Project description:This SuperSeries is composed of the following subset Series: GSE36984: Expression Profiling of Primary Human Fetal and Adult Hematopoietic Stem/Progenitor Cells (HSPCs) and Differentiating Proerythroblasts (ProEs) GSE36985: Comparative profiling of chromatin state maps and transcription factor occupancy during human fetal and adult erythropoiesis GSE36988: Expression Profiling of Primary Human Proerythroblasts (ProEs) After IRF2, IRF6, and MYB shRNA Knockdown Refer to individual Series
Project description:The transcription factor SOX17 is expressed by fetal, but not adult hematoipoietic stem cells (HSCs), and is required for the maintenance of fetal and neonatal, but not adult, HSCs. In the current study we show that ectopic expression of Sox17 in adult HSCs and transiently reconstituting multipotent progenitors was sufficient to confer increased self-renewal potential and the expression of fetal HSC genes including fetal HSC surface markers. To assess the mechanisms by which ectopic Sox17 expression in adult hematopoietic progenitors increased self-renewal potential and conferred fetal HSC properties, we compared the gene expression profiles of E16.5 fetal liver HSCs, young adult bone marrow HSCs, young adult bone marrow CD48+LSK cells, and Sox17-expressing CD48+LSK cells isolated from mice that had been transplanted with MSCV-Sox17-infected bone marrow cells 12 weeks earlier.
Project description:Innate immune signaling protects against pathogens, controls hematopoietic development and functions in oncogenesis, yet the relationship between these mechanisms is incompletely defined. Downregulating the GATA2 transcription factor in fetal hematopoietic progenitor cells upregulates genes encoding innate immune regulators, increases Interferon-g (IFNg) signaling and disrupts differentiation. Here, we demonstrate that deletion of an enhancer that confers GATA2 expression in fetal progenitor cells elevated Toll-Like Receptor (TLR) TLR1/2 and TLR2/6 expression and signaling. Genetic rescue by expressing GATA2 downregulated the elevated TLR signaling. IFNg amplified TLR1/2 and TLR2/6 signaling in GATA2-deficient progenitor cells, synergistically activating cytokine/chemokine genes and elevating cytokine/chemokine production in their myeloid cell progeny. Genome-wide analysis of how IFNg and TLR signaling remodels the progenitor cell transcriptome in GATA2-deficient cells revealed exaggerated responses at innate immune genes harboring motifs for signal-dependent transcription factors. Thus, GATA2 establishes a transcriptome that constrains innate immune signaling, and insufficient GATA2 renders fetal progenitor cells hypersensitive to innate immune signaling.
Project description:Fetal hematopoietic stem and progenitor cells (HSPCs) migrate from fetal liver (FL) to bone marrow (BM) around birth. While adult BM HSPCs and their extrinsic regulation is well studied, little is known about the composition, function, and extrinsic regulation of the first HSPCs to enter the BM. Here, we show that HSPCs colonize multiple fetal bones by E15.5, shift from an MPP2 to an MPP3/4-dominant phenotype by birth, and display little function until E18.5, relative to their FL counterparts. We establish a transcriptional atlas of single perinatal HSPCs, and their putative BM niches, from E15.5 through P0 and show that early fetal BM (FBM) lacks HSPCs with intrinsic stem cell programs and niche cells supportive of HSPCs. In contrast, stem cell programs are preserved in neonatal BM HSPCs, which engage with a niche expressing HSC supportive factors distinct from those seen in adult BM (i.e., IGF).