The Experts below are selected from a list of 5646 Experts worldwide ranked by ideXlab platform
Masayuki Yamamoto - One of the best experts on this subject based on the ideXlab platform.
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mouse tryptase gene expression is coordinately regulated by GATA1 and gata2 in bone marrow derived mast cells
International Journal of Molecular Sciences, 2019Co-Authors: Kinuko Ohneda, Shin'ya Ohmori, Masayuki YamamotoAbstract:Mast cell tryptases have crucial roles in allergic and inflammatory diseases. The mouse tryptase genes represent a cluster of loci on chromosome 16p3.3. While their functional studies have been extensively performed, transcriptional regulation of tryptase genes is poorly understood. In this study, we examined the molecular basis of the tryptase gene expression in bone marrow-derived mast cells (BMMCs) of C57BL/6 mice and in MEDMC-BRC6 mast cells. The expression of the Tpsb2 and Tpsg1 genes, which reside at the 3′-end of the tryptase locus, is significantly decreased by the reduction of the GATA transcription factors GATA1 or GATA2. Chromatin immunoprecipitation assays have shown that the GATA factors bind at multiple regions within the locus, including 1.0 and 72.8 kb upstream of the Tpsb2 gene, and that GATA1 and GATA2 facilitate each other’s DNA binding activity to these regions. Deletion of the −72.8 kb region by genome editing significantly reduced the Tpsb2 and Tpsg1 mRNA levels in MEDMC-BRC6 cells. Furthermore, binding of CTCF and the cohesin subunit Rad21 was found upstream of the −72.8 kb region and was significantly reduced in the absence of GATA1. These results suggest that mouse tryptase gene expression is coordinately regulated by GATA1 and GATA2 in BMMCs.
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Derepression of the DNA Methylation Machinery of the GATA1 Gene Triggers the Differentiation Cue for Erythropoiesis
Molecular and cellular biology, 2017Co-Authors: Jun Takai, Takashi Moriguchi, Mikiko Suzuki, James Douglas Engel, Akihito Otsuki, Fumiki Katsuoka, Saori Katayama, Masahiro Nezu, Masayuki YamamotoAbstract:GATA1 is a critical regulator of erythropoiesis. While the mechanisms underlying the high-level expression of GATA1 in maturing erythroid cells have been studied extensively, the initial activation of the GATA1 gene in early hematopoietic progenitors remains to be elucidated. We previously identified a hematopoietic stem and progenitor cell (HSPC)-specific silencer element (the GATA1 methylation-determining region [G1MDR]) that recruits DNA methyltransferase 1 (Dnmt1) and provokes methylation of the GATA1 gene enhancer. In the present study, we hypothesized that removal of the G1MDR-mediated silencing machinery is the molecular basis of the initial activation of the GATA1 gene and erythropoiesis. To address this hypothesis, we generated transgenic mouse lines harboring a GATA1 bacterial artificial chromosome in which the G1MDR was deleted. The mice exhibited abundant GATA1 expression in HSPCs, in a GATA2-dependent manner. The ectopic GATA1 expression repressed Gata2 transcription and induced erythropoiesis and apoptosis of HSPCs. Furthermore, genetic deletion of Dnmt1 in HSPCs activated GATA1 expression and depleted HSPCs, thus recapitulating the HSC phenotype associated with GATA1 gain of function. These results demonstrate that the G1MDR holds the key to HSPC maintenance and suggest that release from this suppressive mechanism is a fundamental requirement for subsequent initiation of erythroid differentiation.
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GATA1 Binding Kinetics on Conformation-Specific Binding Sites Elicit Differential Transcriptional Regulation.
Molecular and cellular biology, 2016Co-Authors: Atsushi Hasegawa, Masayuki Yamamoto, Hiroshi Kaneko, Daishi Ishihara, Masahiro Nakamura, Akira Watanabe, Cecelia D. Trainor, Ritsuko ShimizuAbstract:GATA1 organizes erythroid and megakaryocytic differentiation by orchestrating the expression of multiple genes that show diversified expression profiles. Here, we demonstrate that GATA1 monovalently binds to a single GATA motif (Single-GATA) while a monomeric GATA1 and a homodimeric GATA1 bivalently bind to two GATA motifs in palindromic (Pal-GATA) and direct-repeat (Tandem-GATA) arrangements, respectively, and form higher stoichiometric complexes on respective elements. The amino-terminal zinc (N) finger of GATA1 critically contributes to high occupancy of GATA1 on Pal-GATA. GATA1 lacking the N finger-DNA association fails to trigger a rate of target gene expression comparable to that seen with the wild-type GATA1, especially when expressed at low level. This study revealed that Pal-GATA and Tandem-GATA generate transcriptional responses from GATA1 target genes distinct from the response of Single-GATA. Our results support the notion that the distinct alignments in binding motifs are part of a critical regulatory strategy that diversifies and modulates transcriptional regulation by GATA1.
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the human GATA1 gene retains a 5 insulator that maintains chromosomal architecture and GATA1 expression levels in splenic erythroblasts
Molecular and Cellular Biology, 2015Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Makiko Hayashi, Hironori Satoh, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a key transcription factor for erythropoiesis. GATA1 gene expression is strictly regulated at the transcriptional level. While the regulatory mechanisms governing mouse GATA1 (mGATA1) gene expression have been studied extensively, how expression of the human GATA1 (hGATA1) gene is regulated remains to be elucidated. To address this issue, we generated hGATA1 bacterial artificial chromosome (BAC) transgenic mouse lines harboring a 183-kb hGATA1 locus covering the hGATA1 exons and distal flanking sequences. Transgenic hGATA1 expression coincides with endogenous mGATA1 expression and fully rescues hematopoietic deficiency in mGATA1 knockdown mice. The transgene exhibited copy number-dependent and integration position-independent expression of hGATA1, indicating the presence of chromatin insulator activity within the transgene. We found a novel insulator element at 29 kb 5′ to the hGATA1 gene and refer to this element as the 5′ CCCTC-binding factor (CTCF) site. Substitution mutation of the 5′ CTCF site in the hGATA1 BAC disrupted the chromatin architecture and led to a reduction of hGATA1 expression in splenic erythroblasts under conditions of stress erythropoiesis. Our results demonstrate that expression of the hGATA1 gene is regulated through the chromatin architecture organized by 5′ CTCF site-mediated intrachromosomal interactions in the hGATA1 locus.
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The Human GATA1 Gene Retains a 5’ Insulator that Maintains Chromosomal Architecture and GATA1 Expression Levels in Splenic Erythroblasts
Molecular and cellular biology, 2015Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Makiko Hayashi, Hironori Satoh, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a key transcription factor for erythropoiesis. GATA1 gene expression is strictly regulated at the transcriptional level. While the regulatory mechanisms governing mouse GATA1 (mGATA1) gene expression have been studied extensively, how expression of the human GATA1 (hGATA1) gene is regulated remains to be elucidated. To address this issue, we generated hGATA1 bacterial artificial chromosome (BAC) transgenic mouse lines harboring a 183-kb hGATA1 locus covering the hGATA1 exons and distal flanking sequences. Transgenic hGATA1 expression coincides with endogenous mGATA1 expression and fully rescues hematopoietic deficiency in mGATA1 knockdown mice. The transgene exhibited copy number-dependent and integration position-independent expression of hGATA1, indicating the presence of chromatin insulator activity within the transgene. We found a novel insulator element at 29 kb 5′ to the hGATA1 gene and refer to this element as the 5′ CCCTC-binding factor (CTCF) site. Substitution mutation of the 5′ CTCF site in the hGATA1 BAC disrupted the chromatin architecture and led to a reduction of hGATA1 expression in splenic erythroblasts under conditions of stress erythropoiesis. Our results demonstrate that expression of the hGATA1 gene is regulated through the chromatin architecture organized by 5′ CTCF site-mediated intrachromosomal interactions in the hGATA1 locus.
Takashi Moriguchi - One of the best experts on this subject based on the ideXlab platform.
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Derepression of the DNA Methylation Machinery of the GATA1 Gene Triggers the Differentiation Cue for Erythropoiesis
Molecular and cellular biology, 2017Co-Authors: Jun Takai, Takashi Moriguchi, Mikiko Suzuki, James Douglas Engel, Akihito Otsuki, Fumiki Katsuoka, Saori Katayama, Masahiro Nezu, Masayuki YamamotoAbstract:GATA1 is a critical regulator of erythropoiesis. While the mechanisms underlying the high-level expression of GATA1 in maturing erythroid cells have been studied extensively, the initial activation of the GATA1 gene in early hematopoietic progenitors remains to be elucidated. We previously identified a hematopoietic stem and progenitor cell (HSPC)-specific silencer element (the GATA1 methylation-determining region [G1MDR]) that recruits DNA methyltransferase 1 (Dnmt1) and provokes methylation of the GATA1 gene enhancer. In the present study, we hypothesized that removal of the G1MDR-mediated silencing machinery is the molecular basis of the initial activation of the GATA1 gene and erythropoiesis. To address this hypothesis, we generated transgenic mouse lines harboring a GATA1 bacterial artificial chromosome in which the G1MDR was deleted. The mice exhibited abundant GATA1 expression in HSPCs, in a GATA2-dependent manner. The ectopic GATA1 expression repressed Gata2 transcription and induced erythropoiesis and apoptosis of HSPCs. Furthermore, genetic deletion of Dnmt1 in HSPCs activated GATA1 expression and depleted HSPCs, thus recapitulating the HSC phenotype associated with GATA1 gain of function. These results demonstrate that the G1MDR holds the key to HSPC maintenance and suggest that release from this suppressive mechanism is a fundamental requirement for subsequent initiation of erythroid differentiation.
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the human GATA1 gene retains a 5 insulator that maintains chromosomal architecture and GATA1 expression levels in splenic erythroblasts
Molecular and Cellular Biology, 2015Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Makiko Hayashi, Hironori Satoh, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a key transcription factor for erythropoiesis. GATA1 gene expression is strictly regulated at the transcriptional level. While the regulatory mechanisms governing mouse GATA1 (mGATA1) gene expression have been studied extensively, how expression of the human GATA1 (hGATA1) gene is regulated remains to be elucidated. To address this issue, we generated hGATA1 bacterial artificial chromosome (BAC) transgenic mouse lines harboring a 183-kb hGATA1 locus covering the hGATA1 exons and distal flanking sequences. Transgenic hGATA1 expression coincides with endogenous mGATA1 expression and fully rescues hematopoietic deficiency in mGATA1 knockdown mice. The transgene exhibited copy number-dependent and integration position-independent expression of hGATA1, indicating the presence of chromatin insulator activity within the transgene. We found a novel insulator element at 29 kb 5′ to the hGATA1 gene and refer to this element as the 5′ CCCTC-binding factor (CTCF) site. Substitution mutation of the 5′ CTCF site in the hGATA1 BAC disrupted the chromatin architecture and led to a reduction of hGATA1 expression in splenic erythroblasts under conditions of stress erythropoiesis. Our results demonstrate that expression of the hGATA1 gene is regulated through the chromatin architecture organized by 5′ CTCF site-mediated intrachromosomal interactions in the hGATA1 locus.
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The Human GATA1 Gene Retains a 5’ Insulator that Maintains Chromosomal Architecture and GATA1 Expression Levels in Splenic Erythroblasts
Molecular and cellular biology, 2015Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Makiko Hayashi, Hironori Satoh, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a key transcription factor for erythropoiesis. GATA1 gene expression is strictly regulated at the transcriptional level. While the regulatory mechanisms governing mouse GATA1 (mGATA1) gene expression have been studied extensively, how expression of the human GATA1 (hGATA1) gene is regulated remains to be elucidated. To address this issue, we generated hGATA1 bacterial artificial chromosome (BAC) transgenic mouse lines harboring a 183-kb hGATA1 locus covering the hGATA1 exons and distal flanking sequences. Transgenic hGATA1 expression coincides with endogenous mGATA1 expression and fully rescues hematopoietic deficiency in mGATA1 knockdown mice. The transgene exhibited copy number-dependent and integration position-independent expression of hGATA1, indicating the presence of chromatin insulator activity within the transgene. We found a novel insulator element at 29 kb 5′ to the hGATA1 gene and refer to this element as the 5′ CCCTC-binding factor (CTCF) site. Substitution mutation of the 5′ CTCF site in the hGATA1 BAC disrupted the chromatin architecture and led to a reduction of hGATA1 expression in splenic erythroblasts under conditions of stress erythropoiesis. Our results demonstrate that expression of the hGATA1 gene is regulated through the chromatin architecture organized by 5′ CTCF site-mediated intrachromosomal interactions in the hGATA1 locus.
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Progenitor stage-specific activity of a cis-acting double GATA motif for GATA1 gene expression.
Molecular and cellular biology, 2014Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a master regulator of erythropoiesis, expression of which is regulated by multiple discrete cis-acting elements. In this study, we examine the activity of a promoter-proximal double GATA (dbGATA) motif, using a GATA1 bacterial artificial chromosome (BAC)-transgenic green fluorescent protein (GFP) reporter (G1BAC-GFP) mouse system. Deletion of the dbGATA motif led to significant reductions in GFP expression in hematopoietic progenitors, while GFP expression was maintained in erythroblasts. Consistently, in mice with a germ line deletion of the dbGATA motif (GATA1(ΔdbGATA) mice), GATA1 expression in progenitors was significantly decreased. The suppressed GATA1 expression was associated with a compensatory increase in GATA2 levels in progenitors. When we crossed GATA1(ΔdbGATA) mice with Gata2 hypomorphic mutant mice (Gata2(fGN/fGN) mice), the GATA1(ΔdbGATA)::Gata2(fGN/fGN) compound mutant mice succumbed to a significant decrease in the progenitor population, whereas both groups of single mutant mice maintained progenitors and survived to adulthood, indicating the functional redundancy between GATA1 and GATA2 in progenitors. Meanwhile, the effects of the dbGATA site deletion on GATA1 expression were subtle in erythroblasts, which showed increased GATA1 binding and enhanced accumulation of active histone marks around the 1st-intron GATA motif of the ΔdbGATA locus. These results thus reveal a novel role of the dbGATA motif in the maintenance of GATA1 expression in hematopoietic progenitors and a functional compensation between the dbGATA site and the 1st-intron GATA motif in erythroblasts.
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A regulatory network governing GATA1 and Gata2 gene transcription orchestrates erythroid lineage differentiation
International Journal of Hematology, 2014Co-Authors: Takashi Moriguchi, Masayuki YamamotoAbstract:GATA transcription factor family members GATA1 and GATA2 play crucial roles in the regulation of lineage-restricted genes during erythroid differentiation. GATA1 is indispensable for survival and terminal differentiation of erythroid, megakaryocytic and eosinophilic progenitors, whereas GATA2 regulates proliferation and maintenance of hematopoietic stem and progenitor cells. Expression levels of GATA1 and GATA2 are primarily regulated at the transcriptional level through auto- and reciprocal regulatory networks formed by these GATA factors. The dynamic and strictly controlled change of expression from GATA2 to GATA1 during erythropoiesis has been referred to as GATA factor switching, which plays a crucial role in erythropoiesis. The regulatory network comprising GATA1 and GATA2 gives rise to the stage-specific changes in GATA1 and Gata2 gene expression during erythroid differentiation, which ensures specific expression of early and late erythroid genes at each stage. Recent studies have also shed light on the genome-wide binding profiles of GATA1 and GATA2, and the significance of epigenetic modification of GATA1 gene during erythroid differentiation. This review summarizes the current understanding of network regulation underlying stage-dependent GATA1 and Gata2 gene expressions and the functional contribution of these GATA factors in erythroid differentiation.
Mikiko Suzuki - One of the best experts on this subject based on the ideXlab platform.
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Derepression of the DNA Methylation Machinery of the GATA1 Gene Triggers the Differentiation Cue for Erythropoiesis
Molecular and cellular biology, 2017Co-Authors: Jun Takai, Takashi Moriguchi, Mikiko Suzuki, James Douglas Engel, Akihito Otsuki, Fumiki Katsuoka, Saori Katayama, Masahiro Nezu, Masayuki YamamotoAbstract:GATA1 is a critical regulator of erythropoiesis. While the mechanisms underlying the high-level expression of GATA1 in maturing erythroid cells have been studied extensively, the initial activation of the GATA1 gene in early hematopoietic progenitors remains to be elucidated. We previously identified a hematopoietic stem and progenitor cell (HSPC)-specific silencer element (the GATA1 methylation-determining region [G1MDR]) that recruits DNA methyltransferase 1 (Dnmt1) and provokes methylation of the GATA1 gene enhancer. In the present study, we hypothesized that removal of the G1MDR-mediated silencing machinery is the molecular basis of the initial activation of the GATA1 gene and erythropoiesis. To address this hypothesis, we generated transgenic mouse lines harboring a GATA1 bacterial artificial chromosome in which the G1MDR was deleted. The mice exhibited abundant GATA1 expression in HSPCs, in a GATA2-dependent manner. The ectopic GATA1 expression repressed Gata2 transcription and induced erythropoiesis and apoptosis of HSPCs. Furthermore, genetic deletion of Dnmt1 in HSPCs activated GATA1 expression and depleted HSPCs, thus recapitulating the HSC phenotype associated with GATA1 gain of function. These results demonstrate that the G1MDR holds the key to HSPC maintenance and suggest that release from this suppressive mechanism is a fundamental requirement for subsequent initiation of erythroid differentiation.
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the human GATA1 gene retains a 5 insulator that maintains chromosomal architecture and GATA1 expression levels in splenic erythroblasts
Molecular and Cellular Biology, 2015Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Makiko Hayashi, Hironori Satoh, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a key transcription factor for erythropoiesis. GATA1 gene expression is strictly regulated at the transcriptional level. While the regulatory mechanisms governing mouse GATA1 (mGATA1) gene expression have been studied extensively, how expression of the human GATA1 (hGATA1) gene is regulated remains to be elucidated. To address this issue, we generated hGATA1 bacterial artificial chromosome (BAC) transgenic mouse lines harboring a 183-kb hGATA1 locus covering the hGATA1 exons and distal flanking sequences. Transgenic hGATA1 expression coincides with endogenous mGATA1 expression and fully rescues hematopoietic deficiency in mGATA1 knockdown mice. The transgene exhibited copy number-dependent and integration position-independent expression of hGATA1, indicating the presence of chromatin insulator activity within the transgene. We found a novel insulator element at 29 kb 5′ to the hGATA1 gene and refer to this element as the 5′ CCCTC-binding factor (CTCF) site. Substitution mutation of the 5′ CTCF site in the hGATA1 BAC disrupted the chromatin architecture and led to a reduction of hGATA1 expression in splenic erythroblasts under conditions of stress erythropoiesis. Our results demonstrate that expression of the hGATA1 gene is regulated through the chromatin architecture organized by 5′ CTCF site-mediated intrachromosomal interactions in the hGATA1 locus.
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The Human GATA1 Gene Retains a 5’ Insulator that Maintains Chromosomal Architecture and GATA1 Expression Levels in Splenic Erythroblasts
Molecular and cellular biology, 2015Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Makiko Hayashi, Hironori Satoh, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a key transcription factor for erythropoiesis. GATA1 gene expression is strictly regulated at the transcriptional level. While the regulatory mechanisms governing mouse GATA1 (mGATA1) gene expression have been studied extensively, how expression of the human GATA1 (hGATA1) gene is regulated remains to be elucidated. To address this issue, we generated hGATA1 bacterial artificial chromosome (BAC) transgenic mouse lines harboring a 183-kb hGATA1 locus covering the hGATA1 exons and distal flanking sequences. Transgenic hGATA1 expression coincides with endogenous mGATA1 expression and fully rescues hematopoietic deficiency in mGATA1 knockdown mice. The transgene exhibited copy number-dependent and integration position-independent expression of hGATA1, indicating the presence of chromatin insulator activity within the transgene. We found a novel insulator element at 29 kb 5′ to the hGATA1 gene and refer to this element as the 5′ CCCTC-binding factor (CTCF) site. Substitution mutation of the 5′ CTCF site in the hGATA1 BAC disrupted the chromatin architecture and led to a reduction of hGATA1 expression in splenic erythroblasts under conditions of stress erythropoiesis. Our results demonstrate that expression of the hGATA1 gene is regulated through the chromatin architecture organized by 5′ CTCF site-mediated intrachromosomal interactions in the hGATA1 locus.
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Progenitor stage-specific activity of a cis-acting double GATA motif for GATA1 gene expression.
Molecular and cellular biology, 2014Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a master regulator of erythropoiesis, expression of which is regulated by multiple discrete cis-acting elements. In this study, we examine the activity of a promoter-proximal double GATA (dbGATA) motif, using a GATA1 bacterial artificial chromosome (BAC)-transgenic green fluorescent protein (GFP) reporter (G1BAC-GFP) mouse system. Deletion of the dbGATA motif led to significant reductions in GFP expression in hematopoietic progenitors, while GFP expression was maintained in erythroblasts. Consistently, in mice with a germ line deletion of the dbGATA motif (GATA1(ΔdbGATA) mice), GATA1 expression in progenitors was significantly decreased. The suppressed GATA1 expression was associated with a compensatory increase in GATA2 levels in progenitors. When we crossed GATA1(ΔdbGATA) mice with Gata2 hypomorphic mutant mice (Gata2(fGN/fGN) mice), the GATA1(ΔdbGATA)::Gata2(fGN/fGN) compound mutant mice succumbed to a significant decrease in the progenitor population, whereas both groups of single mutant mice maintained progenitors and survived to adulthood, indicating the functional redundancy between GATA1 and GATA2 in progenitors. Meanwhile, the effects of the dbGATA site deletion on GATA1 expression were subtle in erythroblasts, which showed increased GATA1 binding and enhanced accumulation of active histone marks around the 1st-intron GATA motif of the ΔdbGATA locus. These results thus reveal a novel role of the dbGATA motif in the maintenance of GATA1 expression in hematopoietic progenitors and a functional compensation between the dbGATA site and the 1st-intron GATA motif in erythroblasts.
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the GATA1 5 region harbors distinct cis regulatory modules that direct gene activation in erythroid cells and gene inactivation in hscs
Blood, 2013Co-Authors: Jun Takai, Kinuko Ohneda, Takashi Moriguchi, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a master regulator of hematopoietic differentiation, but GATA1 expression is inactivated in hematopoietic stem cells (HSCs). Using a bacterial artificial chromosome containing the GATA1 gene modified with green fluorescent protein (GFP) reporter, we explored the function of the 3.7-kb GATA1 upstream region (GdC region) that harbors 3 core cis-elements: GATA1 hematopoietic enhancer, double GATA-motif, and CACCC-motif. Transgenic GFP expression directed by the GATA1-BAC faithfully recapitulated the endogenous GATA1 expression pattern. However, deletion of the GdC-region eliminated reporter expression in all hematopoietic cells. To test whether the combination of the core cis-elements represents the regulatory function of the GdC-region, we replaced the region with a 659-bp minigene that linked the three cis-elements (MG-GFP). The GFP reporter expression directed by the MG-GFP BAC fully recapitulated the erythroid-megakaryocytic GATA1 expression. However, the GFP expression was aberrantly increased in the HSCs and was associated with decreases in DNA methylation and abundant GATA2 binding to the transgenic MG-GFP allele. The 3.2-kb sequences interspaced between the GATA1 hematopoietic enhancer and the double GATA-motif were able to recruit DNA methyltransferase 1, thereby exerting a cis-repressive function in the HSC-like cell line. These results indicate that the 3.2-kb interspacing sequences inactivate GATA1 by maintaining DNA-methylation in the HSCs.
Gerd A. Blobel - One of the best experts on this subject based on the ideXlab platform.
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Pluripotent stem cells reveal erythroid-specific activities of the GATA1 N-terminus
Journal of Clinical Investigation, 2015Co-Authors: Marta Byrska-bishop, Daniel Vandorn, Amy E. Campbell, Marisol Betensky, Philip R. Arca, Fernando F Costa, Richard L Nemiroff, Paul Gadue, Gerd A. BlobelAbstract:Germline GATA1 mutations that result in the production of an amino-truncated protein termed GATA1s (where s indicates short) cause congenital hypoplastic anemia. In patients with trisomy 21, similar somatic GATA1s-producing mutations promote transient myeloproliferative disease and acute megakaryoblastic leukemia. Here, we demonstrate that induced pluripotent stem cells (iPSCs) from patients with GATA1-truncating mutations exhibit impaired erythroid potential, but enhanced megakaryopoiesis and myelopoiesis, recapitulating the major phenotypes of the associated diseases. Similarly, in developmentally arrested GATA1-deficient murine megakaryocyte-erythroid progenitors derived from murine embryonic stem cells (ESCs), expression of GATA1s promoted megakaryopoiesis, but not erythropoiesis. Transcriptome analysis revealed a selective deficiency in the ability of GATA1s to activate erythroid-specific genes within populations of hematopoietic progenitors. Although its DNA-binding domain was intact, chromatin immunoprecipitation studies showed that GATA1s binding at specific erythroid regulatory regions was impaired, while binding at many nonerythroid sites, including megakaryocytic and myeloid target genes, was normal. Together, these observations indicate that lineage-specific GATA1 cofactor associations are essential for normal chromatin occupancy and provide mechanistic insights into how GATA1s mutations cause human disease. More broadly, our studies underscore the value of ESCs and iPSCs to recapitulate and study disease phenotypes.
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dynamic shifts in occupancy by tal1 are guided by gata factors and drive large scale reprogramming of gene expression during hematopoiesis
Genome Research, 2014Co-Authors: Christapher S Morrissey, Gerd A. Blobel, Cheryl A Keller, Tejaswini Mishra, Maxim Pimkin, Mitchell J Weiss, Ross C. HardisonAbstract:We used mouse ENCODE data along with complementary data from other laboratories to study the dynamics of occupancy and the role in gene regulation of the transcription factor TAL1, a critical regulator of hematopoiesis, at multiple stages of hematopoietic differentiation. We combined ChIP-seq and RNA-seq data in six mouse cell types representing a progression from multilineage precursors to differentiated erythroblasts and megakaryocytes. We found that sites of occupancy shift dramatically during commitment to the erythroid lineage, vary further during terminal maturation, and are strongly associated with changes in gene expression. In multilineage progenitors, the likely target genes are enriched for hematopoietic growth and functions associated with the mature cells of specific daughter lineages (such as megakaryocytes). In contrast, target genes in erythroblasts are specifically enriched for red cell functions. Furthermore, shifts in TAL1 occupancy during erythroid differentiation are associated with gene repression (dissociation) and induction (co-occupancy with GATA1). Based on both enrichment for transcription factor binding site motifs and co-occupancy determined by ChIP-seq, recruitment by GATA transcription factors appears to be a stronger determinant of TAL1 binding to chromatin than the canonical E-box binding site motif. Studies of additional proteins lead to the model that TAL1 regulates expression after being directed to a distinct subset of genomic binding sites in each cell type via its association with different complexes containing master regulators such as GATA2, ERG, and RUNX1 in multilineage cells and the lineage-specific master regulator GATA1 in erythroblasts.
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Tissue-Specific Mitotic Bookmarking by Hematopoietic Transcription Factor GATA1
Cell, 2012Co-Authors: Stephan Kadauke, Jan M. Pawlicki, Maheshi Udugama, Jordan C. Achtman, Yong Cheng, Ross C. Hardison, Deepti Jain, Gerd A. BlobelAbstract:Tissue-specific transcription patterns are preserved throughout cell divisions to maintain lineage fidelity. We investigated whether transcription factor GATA1 plays a role in transmitting hematopoietic gene expression programs through mitosis when transcription is transiently silenced. Live-cell imaging revealed that a fraction of GATA1 is retained focally within mitotic chromatin. ChIP-seq of highly purified mitotic cells uncovered that key hematopoietic regulatory genes are occupied by GATA1 in mitosis. The GATA1 coregulators FOG1 and TAL1 dissociate from mitotic chromatin, suggesting that GATA1 functions as platform for their postmitotic recruitment. Mitotic GATA1 target genes tend to reactivate more rapidly upon entry into G1 than genes from which GATA1 dissociates. Mitosis-specific destruction of GATA1 delays reactivation selectively of genes that retain GATA1 during mitosis. These studies suggest a requirement of mitotic "bookmarking" by GATA1 for the faithful propagation of cell-type-specific transcription programs through cell division.
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Lineage-Specific Mitotic Bookmarking by Hematopoietic Transcription Factor GATA1
Blood, 2011Co-Authors: Stephan Kadauke, Jan M. Pawlicki, Maheshi Udugama, Jordan C. Achtman, Yong Cheng, Ross C. Hardison, Gerd A. BlobelAbstract:Abstract 547 Hematopoietic lineage choice decisions are stably maintained throughout many cell divisions. For example, erythroid precursor cells undergo several rounds of cell division during their maturation. During each mitosis, most transcription factors separate from chromatin causing transcription to cease globally. Mitosis therefore poses a challenge for transcription factors to re-associate with the appropriate target sites in chromatin of newborn cells. The epigenetic mechanisms that cement lineage stability and resist cell reprogramming during mitosis are poorly understood, although recent evidence supports the existence of “bookmarking” factors that remain bound to mitotic chromatin. Since the hematopoietic transcription factor GATA1 controls the expression of essentially all erythroid-specific genes, we asked whether it might play a role in maintaining erythroid gene expression programs throughout the cell cycle. Live cell confocal imaging revealed that foci of high GATA1 density are present within mitotic chromatin. Using a novel approach that combines mitotic cell sorting with ChIP-Seq, we defined mitotic GATA1 binding sites on a genome-wide scale. Remarkably, whereas GATA1 vacated the great majority of its target sites during mitosis, including the archetypical GATA1 regulated genes α- and β-globin, those target sites where GATA1 was maintained during mitosis showed a strong tendency to reside near genes encoding key developmental regulators of hematopoiesis (e.g., Zfpm1, Nfe2, Klf1, GATA1, Gata2, Runx1). Tissue-specific GATA1 co-regulators such as FOG-1 and the SCL complex dissociated from GATA1-occupied elements during mitosis, suggesting that GATA1 persists at these sites to facilitate their spatially and temporally appropriate reassembly upon exit from mitosis. Consistent with the notion that GATA1 acts as a mitotic bookmark for its mitotic target genes, timed primary transcript analysis revealed that genes that are marked by GATA1 during mitosis re-activate more rapidly upon G1 entry than those that are not. To directly address the functional importance of mitotic chromatin binding, we developed a version of GATA1 that is selectively degraded during mitosis but remains stable during interphase. This strategy allowed us to prove, for the first time, that the presence of a transcription factor is required specifically during mitosis for timely reactivation of its mitotic target genes. In addition, mitotically disrupted GATA1 failed to fully repress markers of immature erythroid precursors (e.g., Kit, Lyl1), highlighting a potential role of mitotic GATA1 bookmarking for establishing and maintaining lineage- and developmental stage-specific transcriptional programs. Follow-up mechanistic experiments to define the mode by which GATA1 operates during mitosis are underway and will be discussed at the meeting. Together, these studies establish GATA1 as a bona fide mitotic bookmarking factor and provide a deeper understanding by which transcription programs are faithfully perpetuated through cell divisions to maintain lineage stability. Disclosures: No relevant conflicts of interest to declare.
Kinuko Ohneda - One of the best experts on this subject based on the ideXlab platform.
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mouse tryptase gene expression is coordinately regulated by GATA1 and gata2 in bone marrow derived mast cells
International Journal of Molecular Sciences, 2019Co-Authors: Kinuko Ohneda, Shin'ya Ohmori, Masayuki YamamotoAbstract:Mast cell tryptases have crucial roles in allergic and inflammatory diseases. The mouse tryptase genes represent a cluster of loci on chromosome 16p3.3. While their functional studies have been extensively performed, transcriptional regulation of tryptase genes is poorly understood. In this study, we examined the molecular basis of the tryptase gene expression in bone marrow-derived mast cells (BMMCs) of C57BL/6 mice and in MEDMC-BRC6 mast cells. The expression of the Tpsb2 and Tpsg1 genes, which reside at the 3′-end of the tryptase locus, is significantly decreased by the reduction of the GATA transcription factors GATA1 or GATA2. Chromatin immunoprecipitation assays have shown that the GATA factors bind at multiple regions within the locus, including 1.0 and 72.8 kb upstream of the Tpsb2 gene, and that GATA1 and GATA2 facilitate each other’s DNA binding activity to these regions. Deletion of the −72.8 kb region by genome editing significantly reduced the Tpsb2 and Tpsg1 mRNA levels in MEDMC-BRC6 cells. Furthermore, binding of CTCF and the cohesin subunit Rad21 was found upstream of the −72.8 kb region and was significantly reduced in the absence of GATA1. These results suggest that mouse tryptase gene expression is coordinately regulated by GATA1 and GATA2 in BMMCs.
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the human GATA1 gene retains a 5 insulator that maintains chromosomal architecture and GATA1 expression levels in splenic erythroblasts
Molecular and Cellular Biology, 2015Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Makiko Hayashi, Hironori Satoh, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a key transcription factor for erythropoiesis. GATA1 gene expression is strictly regulated at the transcriptional level. While the regulatory mechanisms governing mouse GATA1 (mGATA1) gene expression have been studied extensively, how expression of the human GATA1 (hGATA1) gene is regulated remains to be elucidated. To address this issue, we generated hGATA1 bacterial artificial chromosome (BAC) transgenic mouse lines harboring a 183-kb hGATA1 locus covering the hGATA1 exons and distal flanking sequences. Transgenic hGATA1 expression coincides with endogenous mGATA1 expression and fully rescues hematopoietic deficiency in mGATA1 knockdown mice. The transgene exhibited copy number-dependent and integration position-independent expression of hGATA1, indicating the presence of chromatin insulator activity within the transgene. We found a novel insulator element at 29 kb 5′ to the hGATA1 gene and refer to this element as the 5′ CCCTC-binding factor (CTCF) site. Substitution mutation of the 5′ CTCF site in the hGATA1 BAC disrupted the chromatin architecture and led to a reduction of hGATA1 expression in splenic erythroblasts under conditions of stress erythropoiesis. Our results demonstrate that expression of the hGATA1 gene is regulated through the chromatin architecture organized by 5′ CTCF site-mediated intrachromosomal interactions in the hGATA1 locus.
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The Human GATA1 Gene Retains a 5’ Insulator that Maintains Chromosomal Architecture and GATA1 Expression Levels in Splenic Erythroblasts
Molecular and cellular biology, 2015Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Makiko Hayashi, Hironori Satoh, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a key transcription factor for erythropoiesis. GATA1 gene expression is strictly regulated at the transcriptional level. While the regulatory mechanisms governing mouse GATA1 (mGATA1) gene expression have been studied extensively, how expression of the human GATA1 (hGATA1) gene is regulated remains to be elucidated. To address this issue, we generated hGATA1 bacterial artificial chromosome (BAC) transgenic mouse lines harboring a 183-kb hGATA1 locus covering the hGATA1 exons and distal flanking sequences. Transgenic hGATA1 expression coincides with endogenous mGATA1 expression and fully rescues hematopoietic deficiency in mGATA1 knockdown mice. The transgene exhibited copy number-dependent and integration position-independent expression of hGATA1, indicating the presence of chromatin insulator activity within the transgene. We found a novel insulator element at 29 kb 5′ to the hGATA1 gene and refer to this element as the 5′ CCCTC-binding factor (CTCF) site. Substitution mutation of the 5′ CTCF site in the hGATA1 BAC disrupted the chromatin architecture and led to a reduction of hGATA1 expression in splenic erythroblasts under conditions of stress erythropoiesis. Our results demonstrate that expression of the hGATA1 gene is regulated through the chromatin architecture organized by 5′ CTCF site-mediated intrachromosomal interactions in the hGATA1 locus.
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Progenitor stage-specific activity of a cis-acting double GATA motif for GATA1 gene expression.
Molecular and cellular biology, 2014Co-Authors: Takashi Moriguchi, Kinuko Ohneda, Jun Takai, Mikiko Suzuki, Masayuki YamamotoAbstract:GATA1 is a master regulator of erythropoiesis, expression of which is regulated by multiple discrete cis-acting elements. In this study, we examine the activity of a promoter-proximal double GATA (dbGATA) motif, using a GATA1 bacterial artificial chromosome (BAC)-transgenic green fluorescent protein (GFP) reporter (G1BAC-GFP) mouse system. Deletion of the dbGATA motif led to significant reductions in GFP expression in hematopoietic progenitors, while GFP expression was maintained in erythroblasts. Consistently, in mice with a germ line deletion of the dbGATA motif (GATA1(ΔdbGATA) mice), GATA1 expression in progenitors was significantly decreased. The suppressed GATA1 expression was associated with a compensatory increase in GATA2 levels in progenitors. When we crossed GATA1(ΔdbGATA) mice with Gata2 hypomorphic mutant mice (Gata2(fGN/fGN) mice), the GATA1(ΔdbGATA)::Gata2(fGN/fGN) compound mutant mice succumbed to a significant decrease in the progenitor population, whereas both groups of single mutant mice maintained progenitors and survived to adulthood, indicating the functional redundancy between GATA1 and GATA2 in progenitors. Meanwhile, the effects of the dbGATA site deletion on GATA1 expression were subtle in erythroblasts, which showed increased GATA1 binding and enhanced accumulation of active histone marks around the 1st-intron GATA motif of the ΔdbGATA locus. These results thus reveal a novel role of the dbGATA motif in the maintenance of GATA1 expression in hematopoietic progenitors and a functional compensation between the dbGATA site and the 1st-intron GATA motif in erythroblasts.
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Transcription factor GATA1 is dispensable for mast cell differentiation in adult mice.
Molecular and cellular biology, 2014Co-Authors: Kinuko Ohneda, Yasushi Ishijima, Shin'ya Ohmori, Takashi Moriguchi, Hironori Satoh, Sjaak Philipsen, Masayuki YamamotoAbstract:Although previous studies have shown that GATA1 is required for mast cell differentiation, the effects of the complete ablation of GATA1 in mast cells have not been examined. Using conditional GATA1 knockout mice (GATA1−/y), we demonstrate here that the complete ablation of GATA1 has a minimal effect on the number and distribution of peripheral tissue mast cells in adult mice. The GATA1−/y bone marrow cells were capable of differentiating into mast cells ex vivo. Microarray analyses showed that the repression of GATA1 in bone marrow mast cells (BMMCs) has a small impact on the mast cell-specific gene expression in most cases. Interestingly, however, the expression levels of mast cell tryptases in the mouse chromosome 17A3.3 were uniformly reduced in the GATA1 knockdown cells, and GATA1 was found to bind to a 500-bp region at the 5′ end of this locus. Revealing a sharp contrast to that observed in the GATA1-null BMMCs, GATA2 deficiency resulted in a significant loss of the c-Kit+ FceRIα+ mast cell fraction and a reduced expression of several mast cell-specific genes. Collectively, GATA2 plays a more important role than GATA1 in the regulation of most mast cell-specific genes, while GATA1 might play specific roles in mast cell functions.