Mouse Gata1 3’UTR modulates Gata1 levels to affect erythropoiesis
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ABSTRACT:
PROVIDER: PRJNA1175664 | ENA |
REPOSITORIES: ENA
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Project description:The 3' untranslated region (3'UTR) of mRNA plays several important roles in post-transcriptional gene regulation. Some of its functions include regulating mRNA stability by polyadenylation and microRNAs. However, the overall function of the Gata1 3’UTR in mammals has not been defined. In this study, we used CRISPR/CAS9 technology to knock out the sequence of the mouse Gata1 3’UTR. We found a defect in erythropoiesis in mutant mice, evidenced by macrocytic anemia at the baseline. Ablation of Gata1 3’UTR also resulted in a reduced number of erythroid precursors that might be associated with the cell cycle, especially G2/M disruption in fetal livers. Mechanistically, deletion of the Gata1 3’UTR destabilizes the Gata1 mRNA and ultimately reduces Gata1 protein levels. The low stability of the Gata1 mRNA is unlikely to be caused by the loss of binding of microRNAs or lack of polyadenylation; rather, in part, by the dissociation of AU-rich elements in the 3’UTR with a trans-activating factor Elavl1. Specifically, we transcribed Gata1 3’UTR in vitro and performed an RNA pulldown assay followed by mass spectrometry to profile the proteins that bind the 3’UTR. Gene Ontology analysis demonstrated that several proteins specifically targeting the 3’UTR were found to potentially bind Gata1 3’UTR, among which Elavl1 was in almost all categories related to mRNA stabilization. Western blotting and RNA immunoprecipitation confirmed the direct interaction of Gata1 3’UTR with Elavl1. Manipulation of Elavl1 activity and protein levels by the small molecule inhibitor Dihydro-tanshinone-I and Elavl1 overexpression in fetal liver erythroblasts confirmed Elavl1 as a stabilizing factor for Gata1 mRNA. Our findings shed light on the functional significance of the 3’UTR of Gata1 mRNA in the context of erythroid development. More importantly, our findings highlight the complexity and diversity of regulatory mechanisms that govern Gata1 mRNA stability and precise expression at the post-transcriptional levels. In addition, our findings prove that miRNAs are not always essential in vivo for controlling transcription factor levels to maintain body homeostasis.
2024-10-27 | GSE279985 | GEO
Project description:Mutations in GATA1, which lead to expression of the GATA1s isoform that lacks the GATA1 N-terminus, are seen in patients with Diamond-Blackfan Anemia (DBA). In our efforts to better understand the connection between GATA1s and DBA, we comprehensively studied erythropoiesis in Gata1s mice. Defects in yolks sac and fetal liver hematopoiesis included impaired terminal maturation and reduced numbers of erythroid progenitors. RNA-sequencing revealed that both erythroid and megakaryocytic genes were altered by the loss of the N-terminus, including aberrant up-regulation of Gata2 and Runx1. Mass spectrometry studies demonstrated there was a global increase in H3K27 methylation in the erythroid progenitors. By contrast, chromatin biding assays revealed that, despite similar occupancy of GATA1 and GATA1s, there was a striking reduction of H3K27me3 at regulatory elements of the Gata2 and Runx1 genes. Consistent with the observation that overexpression of GATA2 has been reported to impair erythropoiesis, we found that haploinsufficiency of Gata2 rescued the erythroid defects of Gata1s fetuses. Together, our integrated genomic analysis of transcriptomic and epigenetic signatures reveals that, although Gata1s mice do not precisely model DBA, they provide novel insights into the role of the N-terminus of GATA1 in transcriptional regulation and red blood cell maturation.
2020-04-14 | GSE130156 | GEO
Project description:Stem Cell Leukemia (Scl or Tal1) and Lymphoblastic Leukemia 1 (Lyl1) are highly related members of the basic helix-loop-helix (bHLH) family of transcription factors that are co- expressed in hematopoietic stem cells and the erythro-megakaryocytic lineages. Previous studies suggest that Scl is essential for hematopoietic development including primitive erythropoiesis. However, analysis of single-cell RNA-sequencing data of early embryos showed that primitive erythroid cells express both Scl and Lyl1. Therefore, to determine whether Lyl1 has a functional role in erythropoiesis, we crossed conditional Scl mice with transgenic mice expressing a Cre recombinase under the control of the Epo receptor, active in erythroid progenitors. Surprisingly, embryos with markedly reduced expression of Scl from E9.5 survived to adulthood. In contrast, mice with reduced expression of Scl and absence of Lyl1 (double knockout; DKO) died at E10.5 due to progressive loss of erythropoiesis. Consistent with a phenocopy of Gata1-null mice, gene expression profiling of DKO yolk sacs prior to the loss of erythrocytes (E9.5) revealed loss of Gata1 and many of the known target genes of the SCL-GATA1 complex. ChIP-seq analyses showed that LYL1 exclusively bound a small subset of SCL targets including GATA1. Together, these data show for the first time that Scl and Lyl1 share functional roles in primitive erythropoiesis.
2018-08-13 | GSE103789 | GEO
Project description:Human erythropoiesis is a stepwise process in which multipotent hematopoietic stem/progenitor cells (HSPC) are initially committed towards the erythroid lineage and then differentiated into mature erythroid precursors. Commitment and differentiation are tightly regulated by the coordinated action of a host of transcription factors, including GATA2 and GATA1. Although the role of GATA factors in erythropoiesis has been extensively studied, how they regulate transcription from early to late stages of human erythropoiesis still remains underinvestigated. Here, we investigated GATA-mediated transcriptional regulation along erythroid development through the integrative analysis of gene expression, chromatin modifications, and GATA factors’ binding in human HSPC, early erythroid progenitors, and late precursors. A progressive loss of H3K27 acetylation, a mark of active regulatory elements, and diminished usage of active enhancers and super-enhancers was observed during erythroid lineage commitment and differentiation. We found that GATA factors mediate the transcriptional changes occurring during erythropoiesis through a stage-specific interplay with regulatory elements. In particular, GATA1 binds different sets of regulatory elements in early progenitors and late precursors, and controls the transcription of distinct genes in commitment and differentiation. By Chromosome Conformation Capture and CRISPR/Cas9-mediated genome editing, we demonstrated that the binding of GATA1 to a stage-specific super-enhancer sustains the expression of the stem cell factor receptor KIT in human erythroid progenitors. This study provides insights into the dynamics of epigenetic and transcriptional interactions during erythroid development and highlights a new layer of GATA1-mediated regulation in erythropoiesis.
2020-04-10 | GSE124163 | GEO
Project description:Human erythropoiesis is a stepwise process in which multipotent hematopoietic stem/progenitor cells (HSPC) are initially committed towards the erythroid lineage and then differentiated into mature erythroid precursors. Commitment and differentiation are tightly regulated by the coordinated action of a host of transcription factors, including GATA2 and GATA1. Although the role of GATA factors in erythropoiesis has been extensively studied, how they regulate transcription from early to late stages of human erythropoiesis still remains underinvestigated. Here, we investigated GATA-mediated transcriptional regulation along erythroid development through the integrative analysis of gene expression, chromatin modifications, and GATA factors’ binding in human HSPC, early erythroid progenitors, and late precursors. A progressive loss of H3K27 acetylation, a mark of active regulatory elements, and diminished usage of active enhancers and super-enhancers was observed during erythroid lineage commitment and differentiation. We found that GATA factors mediate the transcriptional changes occurring during erythropoiesis through a stage-specific interplay with regulatory elements. In particular, GATA1 binds different sets of regulatory elements in early progenitors and late precursors, and controls the transcription of distinct genes in commitment and differentiation. By Chromosome Conformation Capture and CRISPR/Cas9-mediated genome editing, we demonstrated that the binding of GATA1 to a stage-specific super-enhancer sustains the expression of the stem cell factor receptor KIT in human erythroid progenitors. This study provides insights into the dynamics of epigenetic and transcriptional interactions during erythroid development and highlights a new layer of GATA1-mediated regulation in erythropoiesis.
2020-04-10 | GSE124164 | GEO
Project description:5q- syndrome is a somatic ribosomopathy linked to the monoallelic deletion of the RPS14 gene and characterized by a proeminent erythroid phenotype. The mechanism of anemia involves an impaired differentiation and increased apoptosis of erythroblasts. We have analyzed total cell extracts from UT7 cells with or without a decrease of RPS14 proteins induced by shRNAs. Our data show that GATA1 protein expression is low in line with a defect in the representation of its mRNA at the ribosome. A global analysis of transcripts on polysomes indicates that translation is selective with a decreased representation of the transcripts with a short coding sequence and UTRs and a highly structured 3’UTR, a subset of transcripts that includes GATA1. Our whole proteome analysis confirms that post-transcriptionally downregulated proteins were encoded by transcripts with a short length and structured 3’UTR. We identified a subset of post-translationally downregulated proteins including ribosomal proteins and translation elongation factors encoded by 5’TOP mRNAs that were enriched on the ribosome. Our results indicate that the thermodynamic characteristics of 3’UTR and in a lesser extend 5’UTR and the transcript length are the determinants of translation selectivity under RPS14 haploinsufficiency conditions and that a post-translational regulation of ribosomal proteins accounts for their decreased content in the cell.
2021-04-28 | PXD008650 | Pride
Project description:Master regulators, such as the hematopoietic transcription factor GATA1, have numerous roles in lineage commitment and differentiation. While human GATA1 mutations result in several blood diseases, all characterized mutations act relatively early to impair hematopoietic differentiation. Here, we describe a distinct form of hemolytic anemia involving impaired terminal erythropoiesis with a reduced lifespan of circulating red blood cells. We show that this unique blood disorder results from mutations in a poorly characterized and intrinsically disordered C-terminal region of GATA1.
2021-12-31 | GSE133414 | GEO
Project description:Master regulators, such as the hematopoietic transcription factor GATA1, have numerous roles in lineage commitment and differentiation. While human GATA1 mutations result in several blood diseases, all characterized mutations act relatively early to impair hematopoietic differentiation. Here, we describe a distinct form of hemolytic anemia involving impaired terminal erythropoiesis with a reduced lifespan of circulating red blood cells. We show that this unique blood disorder results from mutations in a poorly characterized and intrinsically disordered C-terminal region of GATA1.
2021-12-31 | GSE133409 | GEO
Project description:Master regulators, such as the hematopoietic transcription factor GATA1, have numerous roles in lineage commitment and differentiation. While human GATA1 mutations result in several blood diseases, all characterized mutations act relatively early to impair hematopoietic differentiation. Here, we describe a distinct form of hemolytic anemia involving impaired terminal erythropoiesis with a reduced lifespan of circulating red blood cells. We show that this unique blood disorder results from mutations in a poorly characterized and intrinsically disordered C-terminal region of GATA1.
2021-12-31 | GSE133404 | GEO
Project description:Master regulators, such as the hematopoietic transcription factor GATA1, have numerous roles in lineage commitment and differentiation. While human GATA1 mutations result in several blood diseases, all characterized mutations act relatively early to impair hematopoietic differentiation. Here, we describe a distinct form of hemolytic anemia involving impaired terminal erythropoiesis with a reduced lifespan of circulating red blood cells. We show that this unique blood disorder results from mutations in a poorly characterized and intrinsically disordered C-terminal region of GATA1.
2021-12-31 | GSE133403 | GEO