Hematopoietic progenitor cell lines with erythroid and megakaryocyte potential
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ABSTRACT:
PROVIDER: PRJNA1026548 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| Other | ||||
| SRR26338057_1.fastq.gz | Fastqsanger.gz | |||
| SRR26338057_2.fastq.gz | Fastqsanger.gz |
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Project description:Red blood cells (RBC) and platelets are generated from a common, bi-potent hematopoietic progenitor cell in the bone marrow. Important events orchestrating the RBC- and platelet cell lineages have been defined, however, more detailed investigation is limited by shortcomings of suitable experimental systems. Here we show that retroviral delivery of an estrogen-regulated form of Hoxa7 into unseparated bone morrow cells along with thrombopoietin (TPO) can be used to immortalize conditionally bi-potent hematopoietic progenitor cells (Hoxa7-TPO). Hoxa7-TPO cells have lost self-renewal capacity and the potential to differentiate into leukocytes, but retain the potential to differentiate into red blood cells and platelets. Consistent with this lineage potential, gene expression and phenotype analysis of non-differentiated Hoxa7-TPO cells show a close relationship to primary bi-potent megakaryocyte erythrocyte progenitors (MEP). Mature Hoxa7-TPO-derived RBC and platelets are phenotypically and functionally indistinguishable from their primary counterparts. Consistent with this close relationship to primary cells, CRISPR/ Cas9-mediated deletion of known differentiation factors, such as Klf1, Nfe2 and Gp1ba mediates the expected impact on RBC and platelet generation in vivo, with GP1ba-deletion recapitulating the human Bernard-Soulier syndrome. The simplicity of this system, including its amenability to genetic manipulation, along with the unlimited proliferative capacity of Hoxa7-TPO cells will facilitate studies related to cell differentiation and function of the RBC- and megakaryocyte lineages.
2025-06-24 | GSE244976 | GEO
Project description:We newly identified prospectively-isolatable unipotent megakaryocyte progenitor population (MegP) as the major source of megakaryocytes, which emerges directly from early stage of hematopoiesis bypassing megakaryocyte-erythroid lineage bifurcation and contributes to physiological and pathological human megakaryopoiesis. To explore gene expression signature of hematopoietic stem/progenitor populrations miroarry-based whole transcriptome analysis was performed. As a result, gene expression signature of MegP clearly reflected its differentiation potential.
2016-02-02 | GSE77439 | GEO
Project description:RNA-seq on mouse megakaryocyte-erythroid progenitor cells (paired end) For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODE_Data_Use_Policy_for_External_Users_03-07-14.pdf
2016-12-07 | GSE90218 | GEO
Project description:Background: Recent advances in single-cell techniques have provided the opportunity to finely dissect cellular heterogeneity within populations previously defined by âbulkâ assays and to uncover rare cell types. In human hematopoiesis, megakaryocytes and erythroid cells differentiate from a shared precursor, the megakaryocyte-erythroid progenitor (MEP), which remains poorly defined.Results: To clarify the cellular pathway in erythro-megakaryocyte differentiation, we correlated the surface immunophenotype, transcriptional profile and differentiation potential of individual MEP cells. Highly purified, single MEP cells (n=681) were analyzed using index fluorescence-activated cell sorting with parallel targeted transcriptional profiling of the same cells performed using a specifically designed panel of 87 genes. Differentiation potential was tested in novel, single-cell differentiation assays. Our results demonstrated that immunophenotypic MEP in fact comprise three distinct subpopulations: (1) âPre-MEPâ, enriched for erythroid/megakaryocyte progenitors but with residual myeloid differentiation capacity (2) âE-MEPâ, strongly biased towards erythroid differentiation, and (3) âMK-MEPâ, a previously undescribed, rare population of cells that are bipotent but primarily generate megakaryocytic progeny. Therefore, conventionally-defined MEP are in fact a mixed population: a minority give rise to mixed-lineage colonies while the majority of cells are transcriptionally-primed to generate exclusively single-lineage output. Conclusions: Our study clarifies the cellular hierarchy in human megakaryocyte/erythroid lineage commitment and highlights the importance of using a combination of single-cell approaches to dissect cellular heterogeneity and identify rare cell types within a population. We present a novel immunophenotyping strategy that enables the prospective identification of specific intermediate progenitor populations in erythro-megakaryopoiesis, allowing for in-depth study of disorders including inherited cytopenias, myeloproliferative disorders and erythromegakaryocytic leukemias. Multiplex RT-PCR gene expression profiling of 807 human megakaryocyte-erythroid progenitor cells (MEP) isolated from three healthy donors by apheresis following G-CSF treatment. Cells were excluded if more than 70 assays did not result in amplification or displayed Ct higer than 13 for B2M or higher than 15 for GAPDH. Furthermore cells with a mean non-dropout Ct value greater than 20 were removed. This resulted in a dataset of 681 cells, which were subsequently normalised to the mean of B2M and GAPDH expression.
2016-04-02 | E-GEOD-79331 | biostudies-arrayexpress
Project description:Background: Recent advances in single-cell techniques have provided the opportunity to finely dissect cellular heterogeneity within populations previously defined by “bulk” assays and to uncover rare cell types. In human hematopoiesis, megakaryocytes and erythroid cells differentiate from a shared precursor, the megakaryocyte-erythroid progenitor (MEP), which remains poorly defined.Results: To clarify the cellular pathway in erythro-megakaryocyte differentiation, we correlated the surface immunophenotype, transcriptional profile and differentiation potential of individual MEP cells. Highly purified, single MEP cells (n=681) were analyzed using index fluorescence-activated cell sorting with parallel targeted transcriptional profiling of the same cells performed using a specifically designed panel of 87 genes. Differentiation potential was tested in novel, single-cell differentiation assays. Our results demonstrated that immunophenotypic MEP in fact comprise three distinct subpopulations: (1) “Pre-MEP”, enriched for erythroid/megakaryocyte progenitors but with residual myeloid differentiation capacity (2) “E-MEP”, strongly biased towards erythroid differentiation, and (3) “MK-MEP”, a previously undescribed, rare population of cells that are bipotent but primarily generate megakaryocytic progeny. Therefore, conventionally-defined MEP are in fact a mixed population: a minority give rise to mixed-lineage colonies while the majority of cells are transcriptionally-primed to generate exclusively single-lineage output. Conclusions: Our study clarifies the cellular hierarchy in human megakaryocyte/erythroid lineage commitment and highlights the importance of using a combination of single-cell approaches to dissect cellular heterogeneity and identify rare cell types within a population. We present a novel immunophenotyping strategy that enables the prospective identification of specific intermediate progenitor populations in erythro-megakaryopoiesis, allowing for in-depth study of disorders including inherited cytopenias, myeloproliferative disorders and erythromegakaryocytic leukemias.
2016-04-02 | GSE79331 | GEO
Project description:Investigation of immune cell differentiation and function is limited by shortcomings of suitable and scalable experimental systems. Although forced expression of certain Hox genes allows immortalization of hematopoietic progenitor cells, their differentiation potential is limited to select myeloid lineages. Here we show that an estrogen-regulated form of Hoxb8 that is retrovirally delivered into bone marrow cells can be used along with FLT3 ligand to conditionally immortalize early hematopoietic progenitor cells (Hoxb8-FL). Hoxb8-FL cells have lost self-renewal capacity and the ability to adopt megakaryocyte/ erythroid lineage fates, but sustain myeloid and lymphoid differentiation potential. Hoxb8-FL cells differentiate in vitro and in vivo into different myeloid and lymphoid cell types, including macrophages, granulocytes, dendritic cells and B- and T-lymphocytes, which are phenotypically and functionally indistinguishable from their primary counterparts. Given the simplicity to generate Hoxb8-FL cells and their unlimited proliferative capacity, this system provides unique opportunities to investigate cell differentiation and immune cell functions. Hoxb8 expressing immortalized cells
2013-06-05 | E-GEOD-45759 | biostudies-arrayexpress
Project description:Helios represses megakaryocyte priming in hematopoietic stem and progenitor cells
| PRJNA628097 | ENA
Project description:The bHLH transcription factor stem cell leukemia gene (Scl) is a master regulator for hematopoiesis essential for hematopoietic specification and proper differentiation of the erythroid and megakaryocyte lineages. However, the critical downstream targets of Scl remain undefined. Here, we identified a novel Scl target gene, transcription factor myocyte enhancer factor 2 C (Mef2C) from Sclfl/fl fetal liver progenitor cell lines. Analysis of Mef2C-/- embryos showed that Mef2C, in contrast to Scl, is not essential for specification into primitive or definitive hematopoietic lineages. However, adult VavCre+Mef2Cfl/fl mice exhibited platelet defects similar to those observed in Scl deficient mice. The platelet counts were reduced, while platelet size was increased and the platelet shape and granularity was altered. Furthermore, megakaryopoiesis was severely impaired in vitro. ChIP-on-chip analysis revealed that Mef2C is directly regulated by Scl in megakaryocytic cells, but not in erythroid cells. In addition, an Scl independent requirement for Mef2C in B-lymphoid homeostasis was observed in Mef2C-deficient mice, characterized as severe age-dependent reduction of specific B cell progenitor populations reminiscent of premature aging. In summary, this work identifies Mef2C as an integral member of hematopoietic transcription factors with distinct upstream regulatory mechanisms and functional requirements in megakaryocyte and B-lymphoid lineages. Experiment Overall Design: Sclfl/fl hematopoietic progenitor lines were generated from fetal liver progenitors from E12.5 Sclfl/fl embryos9 by immortalization with Hox11 retrovirus.17 Sclfl/fl progenitor cells were cultured with IL3, and a clonal line containing cells with megakaryocyte morphology and acetylcholinesterase (AchE) activity was selected. The Sclfl/fl cell line was transduced with Cre- GFP retrovirus to generate the Scl Experiment Overall Design: Î?/Î? cell line, and the Scl Î?/Î? cell line was transduced with Scl retrovirus to re-introduce Scl expression (Scl Î?/Î? +Scl cell line). Megakaryocyte differentiation was enhanced by adding Tpo for 5 days before harvesting the cells. RNA was extracted with Trizol (Gibco BRL) and RNEasy (Qiagen) kits. Differential gene expression between Sclfl/fl, Scl Î?/Î? and Scl Î?/Î? +Scl cell lines was analyzed by Affymetrix MOE430_2 microarray in the microarray core facility at the Dana-Farber Cancer Institute.
2009-02-08 | E-GEOD-14478 | biostudies-arrayexpress
Project description:SCL/TAL1, a tissue-specific transcription factor of the basic helix-loop-helix (bHLH) family, and c-Kit, a tyrosine kinase receptor, control hematopoietic stem cell survival and quiescence. Here we report that SCL and c-Kit signaling control a common gene expression signature, of which 19 genes are associated with apoptosis. In vivo, SCL levels are limiting for the clonal expansion of Kit+ multipotent and erythroid progenitors. In addition, increased SCL expression specifically enhances the sensitivity of multipotent and megakaryocyte/erythroid progenitors to Steel factor (KIT ligand), whilst a DNA binding mutant antagonizes KIT function and induces apoptosis in progenitors. We conclude that Scl operates downstream of Kit to support the survival of megakaryocyte/erythroid progenitors. Finally, higher SCL expression upregulates Kit in normal bone marrow cells and increases chimerism after bone marrow transplantation, indicating that Scl is also upstream of Kit. We conclude that Scl and Kit establish a positive feedback loop in multipotent and megakaryocyte/erythroid progenitors. c-Kit regulated genes were extrapolated from gene expression profiles of TF-1 erythroid progenitor cells (empty MSCV vector) stimulated with SF (Kit ligand), Epo or GM-CSF. Second, SCL-regulated genes were obtained by expressing a DNA binding-defective SCL mutant (DbSCL) and selecting genes that were differentially expressed in M-oM-^AM-^DbSCL cells versus control cells (MSCV) stimulated with the same cytokines.
2013-07-06 | E-GEOD-48557 | biostudies-arrayexpress
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.
2017-05-15 | E-GEOD-74053 | biostudies-arrayexpress