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Project description:Hematopoietic progenitors respond to developmental and environmental cues to differentiate. The stage-specific steps of differentiation are stereotypic for each cell lineage, and are controlled by transcription. Here we investigate how differential genomic binding of signal-responsive and lineage-restricted transcription factors can specify subsequent stages of erythropoiesis. Using a human erythroid differentiation system, we extensively characterized the co-operation of the BMP signaling transcription factor SMAD1 with the erythroid transcription factors GATA2 and GATA1 in a detailed time-course. BMP signaling enhances erythroid differentiation. SMAD1 is co-recruited with GATA factors at stage-specific genes that are required to have high expression in each stage. Additionally, we observed SMAD1-GATA co-enriched regions are within super-enhancers and span accessible chromatin. Co-bound regions harbor cell-type specific transcription factor motifs that change during commitment, in contrast to GATA-alone regions. Our studies demonstrate that signal-responsive factors together with lineage regulators mark genomic “hotspots” that function to regulate stage-specific gene expression.
2020-09-14 | GSE74482 | GEO
Project description:Hematopoietic progenitors respond to developmental and environmental cues to differentiate. The stage-specific steps of differentiation are stereotypic for each cell lineage, and are controlled by transcription. Here we investigate how differential genomic binding of signal-responsive and lineage-restricted transcription factors can specify subsequent stages of erythropoiesis. Using a human erythroid differentiation system, we extensively characterized the co-operation of the BMP signaling transcription factor SMAD1 with the erythroid transcription factors GATA2 and GATA1 in a detailed time-course. BMP signaling enhances erythroid differentiation. SMAD1 is co-recruited with GATA factors at stage-specific genes that are required to have high expression in each stage. Additionally, we observed SMAD1-GATA co-enriched regions are within super-enhancers and span accessible chromatin. Co-bound regions harbor cell-type specific transcription factor motifs that change during commitment, in contrast to GATA-alone regions. Our studies demonstrate that signal-responsive factors together with lineage regulators mark genomic “hotspots” that function to regulate stage-specific gene expression.
2020-09-14 | GSE74479 | GEO
Project description:Hematopoietic progenitors respond to developmental and environmental cues to differentiate. The stage-specific steps of differentiation are stereotypic for each cell lineage, and are controlled by transcription. Here we investigate how differential genomic binding of signal-responsive and lineage-restricted transcription factors can specify subsequent stages of erythropoiesis. Using a human erythroid differentiation system, we extensively characterized the co-operation of the BMP signaling transcription factor SMAD1 with the erythroid transcription factors GATA2 and GATA1 in a detailed time-course. BMP signaling enhances erythroid differentiation. SMAD1 is co-recruited with GATA factors at stage-specific genes that are required to have high expression in each stage. Additionally, we observed SMAD1-GATA co-enriched regions are within super-enhancers and span accessible chromatin. Co-bound regions harbor cell-type specific transcription factor motifs that change during commitment, in contrast to GATA-alone regions. Our studies demonstrate that signal-responsive factors together with lineage regulators mark genomic “hotspots” that function to regulate stage-specific gene expression.
2020-09-14 | GSE74472 | GEO
Project description:BMP signaling cooperates with GATA factors to govern stage-specific gene expression during erythroid differentiation
| PRJNA300535 | ENA
Project description:Lineage-determination transcription factors coordinate cell differentiation and proliferation by controlling the synthesis of lineage-specific gene products as well as cell cycle regulators. GATA-1 is a master regulator of erythropoiesis. Its role in regulating erythroid-specific genes has been extensively studied, whereas its role in controlling genes that regulate cell proliferation is less understood. Ectopic expression of GATA-1 in erythroleukemia cells releases the block to their differentiation and leads to terminal cell division. An early event in reprogramming the erythroleukemia cells is induction of the cyclin-dependent kinase inhibitor p21. Remarkably, ectopic expression of p21 also induces the erythroleukemia cells to differentiate. We now report that GATA-1 directly regulates transcription of the p21 gene in both erythroleukemia cells and normal erythroid progenitors. Using reporter, electrophoretic mobility shift, and chromatin immunoprecipitation assays, we show that GATA-1 stimulates p21 gene transcription by binding to consensus binding sites in the upstream region of the p21 gene promoter. This activity is also dependent on a binding site for Sp1/KLF-like factors near the transcription start site. Our findings indicate that p21 is a crucial downstream gene target and effector of GATA-1 during red blood cell terminal differentiation.
| S-EPMC3019278 | biostudies-literature
Project description:The first intronic mutations in the intron 1 GATA site (int-1-GATA) of 5-aminolevulinate synthase 2 (ALAS2) have been identified in X-linked sideroblastic anemia (XLSA) pedigrees, strongly suggesting it could be causal mutations of XLSA. However, the function of this int-1-GATA site during in vivo development remains largely unknown. Here, we generated mice lacking a 13 bp fragment, including this int-1-GATA site (T AGATAA: AGCCCC) and found that hemizygous deletion led to an embryonic lethal phenotype due to severe anemia resulting from a lack of ALAS2 expression, indicating that this non-coding sequence is indispensable for ALAS2 expression in vivo Further analyses revealed that this int-1-GATA site anchored the GATA site in intron 8 (int-8-GATA) and the proximal promoter, forming a long-range loop to enhance ALAS2 expression by an enhancer complex including GATA1, TAL1, LMO2, LDB1 and Pol II at least, in erythroid cells. However, compared with the int-8-GATA site, the int-1-GATA site is more essential for regulating ALAS2 expression through CRISPR/Cas9-mediated site-specific deletion. Therefore, the int-1-GATA site could serve as a valuable site for diagnosing XLSA in cases with unknown mutations.
| S-EPMC5314798 | biostudies-literature
Project description:Transcription factor GATA-1 is essential for normal erythropoiesis. GATA-binding sites are consistently found in promoters or enhancers of genes expressed selectively in erythroid cells. To discover novel GATA-1-regulated genes, we searched for GATA-1-activated transcripts in G1E cells, an erythroid line derived from GATA-1(-) embryonic stem cells. By subtractive analysis, we identified a new ATP-binding cassette (ABC) transporter that is strongly and rapidly induced by GATA-1. This protein, named ABC-me (for ABC-mitochondrial erythroid), localizes to the mitochondrial inner membrane and is expressed at particularly high levels in erythroid tissues of embryos and adults. ABC-me is induced during erythroid maturation in cell lines and primary hematopoietic cells, and its overexpression enhances hemoglobin synthesis in erythroleukemia cells. The ABC proteins participate in diverse physiological processes by coupling ATP hydrolysis to the transport of a variety of substrates across cell membranes. We speculate that ABC-me, a newly identified erythroid-expressed ABC superfamily member, may mediate critical mitochondrial transport functions related to heme biosynthesis.
| S-EPMC212759 | biostudies-literature
Project description:BackgroundThe transcription factor GATA-2 is predominantly expressed in hematopoietic stem and progenitor cells and counteracts the erythroid-specific transcription factor GATA-1, to modulate the proliferation and differentiation of hematopoietic cells. During hematopoietic cell differentiation, GATA-2 exhibits dynamic expression patterns, which are regulated by multiple transcription factors.MethodsStable LSD1-knockdown cell lines were established by growing murine erythroleukemia (MEL) or mouse embryonic stem cells together with virus particles, in the presence of Polybrene(®) at 4 ?g/mL, for 24-48 hours followed by puromycin selection (1 ?g/mL) for 2 weeks. Real-time polymerase chain reaction (PCR)-based quantitative chromatin immunoprecipitation (ChIP) analysis was used to test whether the TAL1 transcription factor is bound to 1S promoter in the GATA-2 locus or whether LSD1 colocalizes with TAL1 at the 1S promoter. The sequential ChIP assay was utilized to confirm the role of LSD1 in the regulation of H3K4me2 at the GATA-2 locus during erythroid differentiation. Western blot analysis was employed to detect the protein expression. The alamarBlue(®) assay was used to examine the proliferation of the cells, and the absorbance was monitored at optical density (OD) 570 nm and OD 600 nm.ResultsIn this study, we showed that LSD1 regulates the expression of GATA-2 during erythroid differentiation. Knockdown of LSD1 results in increased GATA-2 expression and inhibits the differentiation of MEL and embryonic stem cells. Furthermore, we demonstrated that LSD1 binds to the 1S promoter of the GATA-2 locus and suppresses GATA-2 expression, via histone demethylation.ConclusionOur data revealed that LSD1 mediates erythroid differentiation, via epigenetic modification of the GATA-2 locus.
| S-EPMC4482369 | biostudies-literature
Project description:Bone morphogenetic proteins (BMPs) belong to the TGF-β superfamily of structurally related signaling proteins that regulate a wide array of cellular functions. The key step in BMP signal transduction is the BMP receptor-mediated phosphorylation of transcription factors Smad1, 5, and 8 (collectively Smad1/5/8), which leads to the subsequent activation of BMP-induced gene transcription in the nucleus. In this study, we describe the identification and characterization of PPM1H as a novel cytoplasm-localized Smad1/5/8-specific phosphatase. PPM1H directly interacts with Smad1/5/8 through its Smad-binding domain, and dephosphorylates phospho-Smad1/5/8 (P-Smad1/5/8) in the cytoplasm. Ectopic expression of PPM1H attenuates BMP signaling, whereas loss of PPM1H activity or expression greatly enhances BMP-dependent gene regulation and mesenchymal differentiation. In conclusion, this study suggests that PPM1H acts as a gatekeeper to prevent excessive BMP signaling through dephosphorylation and subsequent nuclear exclusion of P-Smad1/5/8 proteins.
| S-EPMC4042171 | biostudies-literature
Project description:Expression of Gfi (growth factor-independence)-1B, a Gfi-1-related transcriptional repressor, is restricted to erythroid lineage cells and is essential for erythropoiesis. We have determined the transcription start site of the human Gfi-1B gene and located its first non-coding exon approximately 7.82 kb upstream of the first coding exon. The genomic sequence preceding this first non-coding exon has been identified to be its erythroid-specific promoter region in K562 cells. Using gel-shift and chromatin immunoprecipitation (ChIP) assays, we have demonstrated that NF-Y and GATA-1 directly participate in transcriptional activation of the Gfi-1B gene in K562 cells. Ectopic expression of GATA-1 markedly stimulates the activity of the Gfi-1B promoter in a non-erythroid cell line U937. Interestingly, our results have indicated that this GATA-1-mediated trans-activation is dependent on NF-Y binding to the CCAAT site. Here we conclude that functional cooperation between GATA-1 and NF-Y contributes to erythroid-specific transcriptional activation of Gfi-1B promoter.
| S-EPMC506805 | biostudies-literature