Single-cell analysis of megakaryopoiesis in peripheral CD34+ cells: insights into ETV6-related thrombocytopenia
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ABSTRACT:
PROVIDER: PRJNA849164 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR19649505_1.fastq.gz | Fastqsanger.gz | |||
| SRR19649505_2.fastq.gz | Fastqsanger.gz | |||
| SRR19649506_1.fastq.gz | Fastqsanger.gz | |||
| SRR19649506_2.fastq.gz | Fastqsanger.gz | |||
| SRR19649507_1.fastq.gz | Fastqsanger.gz |
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Project description:Using single-cell RNA sequencing (scRNAseq), we investigated ETV6-variant (P214L and F417LTer4) carriers who are affected by constitutional thrombopenia. We demonstrate that ETV6-variant carriers presented transcriptomic aberrant populations of megakaryocytes (MK) and mega-erythroid progenitors (MEP) with affected pathways. We confirmed this bioinformatic analyzes by functional assays. We also demonstrate a bypass of common myeloid progenitors (CMP) during physiologic differentiation from hematopoietic stem/progenitors cells (HSPC) to MK and we identified a new population of MK-primed CMP in our culture model. We raise this analysis as a new exploratory modus operandi for constitutional diseases.
2023-06-15 | GSE206089 | GEO
Project description:Inherited thrombocytopenia results in low platelet counts and increased bleeding. Subsets of these patients have monoallelic germline mutations in ETV6 or RUNX1 and a heightened risk of developing hematologic malignancies. Utilizing CRISPR/Cas9, we compared the in vitro phenotype of hematopoietic progenitor cells and megakaryocytes derived from induced pluripotent stem cell (iPSC) lines harboring mutations in either ETV6 or RUNX1. Both mutant lines display phenotypes consistent with a platelet-bleeding disorder. Surprisingly, these cellular phenotypes were largely distinct. The ETV6-mutant iPSCs yield more hematopoietic progenitor cells and megakaryocytes, but the megakaryocytes are immature and less responsive to agonist stimulation. On the contrary, RUNX1-mutant iPSCs yield fewer hematopoietic progenitor cells and megakaryocytes, but the megakaryocytes are more responsive to agonist stimulation. However, both mutant iPSC lines display defects in proplatelet formation. Our work highlights that while patients harboring germline ETV6 or RUNX1 mutations have similar clinical phenotypes, the molecular mechanisms may be distinct.
2021-03-06 | GSE168326 | GEO
Project description:Exparession profiling of ETV6-NCOA2 at different stages of disease dEmpty vectorelopment: CD34 cord blood cells transduced with ETV6-NCOA2-GFP or with empty vector-GFP, CD34 cord blood cells transduced with ETV6-NCOA2-NGFR or ETV6-NCOA2-NGFR + NOTCH1-L1601PdP-GFP and transplanted in NSGS mice, and ETV6-NCOA2 patient derived xenografts.
2022-05-16 | GSE161702 | GEO
Project description:S78454 (Abexinostat, PCI-24781) is a pan histone deacetylase inhibitor (HDACi) that demonstrates efficacy in malignancy treatment. Like other HDACi, this drug induces a profound thrombocytopenia, whose mechanism is partially understood. We have tested its effect at doses reached in patients plasma on in vitro megakaryopoiesis derived from human CD34+ cells. When added at day 0 in culture, S78454 inhibited CFU-MK growth, megakaryocyte (MK) proliferation and differentiation. This effect only required a short incubation period. When added later on (day 8), the compound induced a dose-dependent decrease (up to 10-fold) in proplatelet formation. Decreased proliferation was due to induction of apoptosis and was not related to a defect in TPO/MPL/JAK2 signaling. MK gene profiling revealed a silencing in the expression of DNA repair genes with a marked RAD51 decrease at protein level. Double DNA strand breaks were increased as attested by elevated gH2AX phosphorylation level. Consequently, ATM was phosphorylated leading to p53 stabilization and increased BAX and p21 expression. The use of a p53 shRNA rescued apoptosis, but only partially the defect in proplatelet formation. These results suggest that HDACi induces a thrombocytopenia by a p53 dependent mechanism along MK differentiation and a p53 independent mechanism for proplatelet formation.
2013-06-30 | E-MTAB-1180 | biostudies-arrayexpress
Project description:Mixed-phenotype acute leukemia is a rare subtype of leukemia in which both myeloid and lymphoid markers are co-expressed on the same malignant cells. Their pathogenesis is largely unknown, and their treatment is challenging. We have previously discovered the specific association between recurrent t(8;12)(q13;p13) chromosomal translocation, creating ETV6-NCOA2 fusion, with T/Myeloid leukemias. Here we report that the ETV6-NCOA2 initiates T/Myeloid leukemia in preclinical models and examine the expression profile induced by ETV6-NCOA2 in CD34+ cord blood progenitors in-vitro.
2022-05-16 | GSE162230 | GEO
Project description:This SuperSeries is composed of the SubSeries listed below. ETS Variant 6 (ETV6) encodes an essential transcriptional repressor abundantly expressed in hematopoietic stem and progenitor cells (HSPCs), where it is required for adult hematopoiesis. Heterozygous pathogenic germline ETV6 variants are associated with Thrombocytopenia 5 (T5), a poorly understood genetic condition predisposing to thrombocytopenia and hematologic malignancies. To elucidate how germline ETV6 variants impact the HSPC compartment and contribute to disease, we generated a knock-in mouse model harboring an Etv6R355X loss-of-function variant, which is equivalent to the T5-associated variant ETV6R359X. Under homeostatic conditions, all HSPC subpopulations are present in the bone marrow (BM) of Etv6R355X/+ mice; however, these animals display subtle shifts in the proportions and/or numbers of specific progenitor subtypes. To examine whether the Etv6R355X/+ mutation impacts HSPC function, we carried out serial competitive transplantation and observed that Etv6R355X/+ lineage-sca1+cKit+ (LSK) cells exhibit significantly impaired reconstitution compared to Etv6+/+ LSK cells with near complete failure to repopulate irradiated-recipients by the tertiary transplant. Mechanistic studies incorporating CUT&RUN, ATAC-Seq and Hi-C identify ETV6 binding at inflammatory gene loci, including those within the TNF signaling pathway, in Etv6+/+ HSPCs, mouse BM-progenitor-derived HPC5 cells, and human CD34+ cells. Further, single-cell RNA-Seq of BM cells isolated post-competitive transplantation reveals upregulation of inflammatory genes in Etv6R355X/+ compared to Etv6+/+ progenitors. Corroborating these findings, Etv6R355X/+ HSPCs produce significantly more TNF than Etv6+/+ cells post-transplantation. From these studies, we conclude that ETV6 is required to repress inflammatory gene expression in HSPCs under conditions of hematopoietic stress and this mechanism may be critical to sustain HSPC function.
2023-08-24 | GSE213597 | GEO
Project description:ETV6-NCOA2 expression in CD34 positive cord blood cells
| PRJNA681553 | ENA
Project description:Purpose and Methods: Immune thrombocytopenia (ITP), an autoimmune disorder, is mainly caused by megakaryocyte dysfunction, although the underlying mechanisms remain unclear. Here, we performed single-cell transcriptome profiling of bone marrow (BM) CD34+ hematopoietic stem and progenitor cells (HSPCs) to determine defects in megakaryopoiesis in ITP. Results: Gene expression, cell-cell interactions, and transcriptional regulatory networks varied in HSPCs of ITP, particularly in natural killer/T cell progenitors and a pre-B cell subset, highlighting the selective immune aberratixon associated with defective megakaryopoiesis in ITP. CD9 can be used to enrich megakaryocyte-biased HSPCs, and CD9+Lin−CD34+CD45RA− HSPCs have the potential to be novel diagnostic and therapeutic targets in ITP. Further analysis revealed that megakaryocytic progenitors (MkP) can be divided into seven subclusters with different gene expression patterns and functions. The ITP-associated DEGs were MkP subtype-specific, with most DEGs concentrated in the subcluster possessing dual functions of immunomodulation and platelet generation. Conclusions: This study comprehensively dissects defective hematopoiesis and provides novel therapeutic targets for ITP.
2022-09-03 | GSE196676 | GEO
Project description:ETS Variant 6 (ETV6) encodes an essential transcriptional repressor abundantly expressed in hematopoietic stem and progenitor cells (HSPCs), where it is required for adult hematopoiesis. Heterozygous pathogenic germline ETV6 variants are associated with Thrombocytopenia 5 (T5), a poorly-understood genetic condition predisposing to thrombocytopenia and hematologic malignancies. To elucidate how germline ETV6 variants impact the HSPC compartment and contribute to disease, we generated a knock-in mouse harboring an Etv6R355X loss-of-function variant, which represents the mouse equivalent to the T5-associated variant ETV6R359X. All HSPC subpopulations are present in the bone marrow (BM) of Etv6R355X/+ mice under homeostatic conditions; however, these animals exhibit subtle shifts in the proportions and/or numbers of specific progenitor subtypes. To examine whether the Etv6R355X/+ mutation impacts HSPC function, we carried out serial competitive transplantation and observed that Etv6R355X/+ lineagesca1+cKit+ (LSK) cells exhibit significantly impaired reconstitution, with near complete failure to repopulate irradiated-recipients by the tertiary transplant. Mechanistic studies incorporating CUT&RUN, ATAC-Seq and Hi-C identify ETV6 binding at inflammatory gene loci, including those within the TNF pathway in Etv6+/+ HSPCs, the mouse BM-progenitor derived HPC5 cell line, and G-CSF-mobilized human CD34+ cells. Further, single-cell RNA sequencing of mouse LSK cells isolated six-weeks post-competitive transplantation reveals upregulation of inflammatory gene pathways. Corroborating these findings, we observe significantly increased production of TNF by Etv6R355X/+ versus Etv6+/+ HSPCs post-transplantation. From these studies, we conclude that ETV6 represses inflammatory response genes within HSPCs under conditions of hematopoietic stress, and that this mechanism may be critical to sustain HSPC function.
2023-08-24 | GSE213595 | GEO
Project description:ETS Variant 6 (ETV6) encodes an essential transcriptional repressor abundantly expressed in hematopoietic stem and progenitor cells (HSPCs), where it is required for adult hematopoiesis. Heterozygous pathogenic germline ETV6 variants are associated with Thrombocytopenia 5 (T5), a poorly-understood genetic condition predisposing to thrombocytopenia and hematologic malignancies. To elucidate how germline ETV6 variants impact the HSPC compartment and contribute to disease, we generated a knock-in mouse harboring an Etv6R355X loss-of-function variant, which represents the mouse equivalent to the T5-associated variant ETV6R359X. All HSPC subpopulations are present in the bone marrow (BM) of Etv6R355X/+ mice under homeostatic conditions; however, these animals exhibit subtle shifts in the proportions and/or numbers of specific progenitor subtypes. To examine whether the Etv6R355X/+ mutation impacts HSPC function, we carried out serial competitive transplantation and observed that Etv6R355X/+ lineagesca1+cKit+ (LSK) cells exhibit significantly impaired reconstitution, with near complete failure to repopulate irradiated-recipients by the tertiary transplant. Mechanistic studies incorporating CUT&RUN, ATAC-Seq and Hi-C identify ETV6 binding at inflammatory gene loci, including those within the TNF pathway in Etv6+/+ HSPCs, the mouse BM-progenitor derived HPC5 cell line, and G-CSF-mobilized human CD34+ cells. Further, single-cell RNA sequencing of mouse LSK cells isolated six-weeks post-competitive transplantation reveals upregulation of inflammatory gene pathways. Corroborating these findings, we observe significantly increased production of TNF by Etv6R355X/+ versus Etv6+/+ HSPCs post-transplantation. From these studies, we conclude that ETV6 represses inflammatory response genes within HSPCs under conditions of hematopoietic stress, and that this mechanism may be critical to sustain HSPC function.
2023-08-24 | GSE213594 | GEO