High Fractional Occupancy of a Tandem MARE and its Role in Long-Range β-globin Gene Regulation

Enhancers and promoters assemble protein complexes that ultimately regulate the recruitment and activity of RNA polymerases. Previous work has shown that at least some enhancers form stable protein complexes leading to the formation of enhanceosomes. We analyzed protein-DNA interactions in the murine β-globin gene locus using MAPit (methyltransferase accessibility protocol for individual templates). The data show that a tandem MARE (Maf recognition element) in locus control region (LCR) hypersensitive site 2 (HS2) reveals a remarkably high degree of occupancy during differentiation of mouse erythroleukemia cells. Most of the other transcription factor binding sites in LCR HS2 or in the adult β-globin gene promoter regions exhibit low fractional occupancy, suggesting highly dynamic protein-DNA interactions. Targeting of an artificial zinc finger DNA-binding domain (ZF-DBD) to the HS2 tandem MARE caused reduction in the association of MARE binding proteins and transcription complexes at LCR HS2 and the adult βmaj-globin gene promoter but did not affect expression of the βmin-globin gene. The data demonstrate that a stable MARE-associated footprint in LCR HS2 is important for the recruitment of transcription complexes to the adult βmaj-globin gene promoter during erythroid cell differentiation.

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Alterations in Protein-DNA Interactions in the γ-Globin Gene Promoter in Response to Butyrate Therapy

Blood ◽

10.1182/blood.v92.8.2924 ◽

1998 ◽

Vol 92 (8) ◽

pp. 2924-2933 ◽

Cited By ~ 46

Author(s):

Tohru Ikuta ◽

Yuet Wai Kan ◽

Paul S. Swerdlow ◽

Douglas V. Faller ◽

Susan P. Perrine

Keyword(s):

Gene Expression ◽

Globin Gene ◽

Gene Promoter ◽

Dna Interactions ◽

Globin Mrna ◽

Mobility Shift ◽

Protein Dna Interactions ◽

American Society ◽

Globin Gene Expression

Abstract The mechanisms by which pharmacologic agents stimulate γ-globin gene expression in β-globin disorders has not been fully established at the molecular level. In studies described here, nucleated erythroblasts were isolated from patients with β-globin disorders before and with butyrate therapy, and globin biosynthesis, mRNA, and protein-DNA interactions were examined. Expression of γ-globin mRNA increased twofold to sixfold above baseline with butyrate therapy in 7 of 8 patients studied. A 15% to 50% increase in γ-globin protein synthetic levels above baseline γ globin ratios and a relative decrease in β-globin biosynthesis were observed in responsive patients. Extensive new in vivo footprints were detected in erythroblasts of responsive patients in four regions of the γ-globin gene promoter, designated butyrate-response elements gamma 1-4 (BRE-G1-4). Electrophoretic mobility shift assays using BRE-G1 sequences as a probe demonstrated that new binding of two erythroid-specific proteins and one ubiquitous protein, CP2, occurred with treatment in the responsive patients and did not occur in the nonresponder. The BRE-G1 sequence conferred butyrate inducibility in reporter gene assays. These in vivo protein-DNA interactions in human erythroblasts in which γ-globin gene expression is being altered strongly suggest that nuclear protein binding, including CP2, to the BRE-G1 region of the γ-globin gene promoter mediates butyrate activity on γ-globin gene expression. © 1998 by The American Society of Hematology.

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Alterations in Protein-DNA Interactions in the γ-Globin Gene Promoter in Response to Butyrate Therapy

Blood ◽

10.1182/blood.v92.8.2924.420k16_2924_2933 ◽

1998 ◽

Vol 92 (8) ◽

pp. 2924-2933

Author(s):

Tohru Ikuta ◽

Yuet Wai Kan ◽

Paul S. Swerdlow ◽

Douglas V. Faller ◽

Susan P. Perrine

Keyword(s):

Gene Expression ◽

Globin Gene ◽

Gene Promoter ◽

Dna Interactions ◽

Globin Mrna ◽

Mobility Shift ◽

Protein Dna Interactions ◽

American Society ◽

Globin Gene Expression

The mechanisms by which pharmacologic agents stimulate γ-globin gene expression in β-globin disorders has not been fully established at the molecular level. In studies described here, nucleated erythroblasts were isolated from patients with β-globin disorders before and with butyrate therapy, and globin biosynthesis, mRNA, and protein-DNA interactions were examined. Expression of γ-globin mRNA increased twofold to sixfold above baseline with butyrate therapy in 7 of 8 patients studied. A 15% to 50% increase in γ-globin protein synthetic levels above baseline γ globin ratios and a relative decrease in β-globin biosynthesis were observed in responsive patients. Extensive new in vivo footprints were detected in erythroblasts of responsive patients in four regions of the γ-globin gene promoter, designated butyrate-response elements gamma 1-4 (BRE-G1-4). Electrophoretic mobility shift assays using BRE-G1 sequences as a probe demonstrated that new binding of two erythroid-specific proteins and one ubiquitous protein, CP2, occurred with treatment in the responsive patients and did not occur in the nonresponder. The BRE-G1 sequence conferred butyrate inducibility in reporter gene assays. These in vivo protein-DNA interactions in human erythroblasts in which γ-globin gene expression is being altered strongly suggest that nuclear protein binding, including CP2, to the BRE-G1 region of the γ-globin gene promoter mediates butyrate activity on γ-globin gene expression. © 1998 by The American Society of Hematology.

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LSD1-Mediated Epigenetic Modification Is Important for TAL1 Function

Blood ◽

10.1182/blood.v112.11.4757.4757 ◽

2008 ◽

Vol 112 (11) ◽

pp. 4757-4757

Author(s):

Xin Hu ◽

Xingguo Li ◽

River Ybarra ◽

Kristell Valverde ◽

Xueqi Fu ◽

...

Keyword(s):

Target Genes ◽

Protein Complexes ◽

Epigenetic Modification ◽

Es Cells ◽

Gene Promoter ◽

Erythroid Cell ◽

Hematopoietic Stem ◽

Target Gene Promoter ◽

Cell Growth And Differentiation ◽

Erythroid Cell Differentiation

Abstract TAL1/SCL is critical for normal and abnormal hematopoiesis by regulating hematopoietic stem/progenitor cell growth and differentiation. However, it is still unclear how its transcriptional activities are controlled during hematopoiesis. Here, we undertook the biochemical isolation of TAL1-associated protein complexes in erythroleukemia cells and showed that TAL1 interacts with histone demethylase LSD1 complexes containing LSD1, CoREST, HDAC1 and HDAC2. Interestingly, although TAL1 specifically colocalizes with LSD1 at the target gene promoter p4.2 in undifferentiated MEL cells, the recruitment of LSD1 is decreased at the p4.2 promoter upon induced MEL differentiation indicating that LSD1 may differentially regulate TAL1 target genes during differentiation. The siRNA-mediated knockdown of LSD1 in MEL and ES cells resulted in the derepression of p4.2 by increasing dimeH3K4 at their promoter region, respectively. Finally, we demonstrated that TAL1-associated LSD1 complexes, H3K4 demethylase, and histone deacetylase activities are coordinately regulated during erythroid cell differentiation. Thus, the data suggest that LSD1 mediated epigenetic modification may affect hematopoiesis and leukemogenesis through its association with the lineage-specific transcription factor TAL1.

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Human major histocompatibility complex class II-associated invariant chain gene promoter. Functional analysis and in vivo protein/DNA interactions of constitutive and IFN-gamma-induced expression.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(19)74318-x ◽

1993 ◽

Vol 268 (35) ◽

pp. 26328-26333

Author(s):

A M Brown ◽

K L Wright ◽

J P Ting

Keyword(s):

Gene Promoter ◽

Invariant Chain ◽

Chain Gene ◽

Complex Class ◽

Dna Interactions ◽

Human Major Histocompatibility Complex ◽

Ifn Gamma ◽

Histocompatibility Complex ◽

Protein Dna Interactions

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High similarity among ChEC-seq datasets

10.1101/2021.02.04.429774 ◽

2021 ◽

Author(s):

Chitvan Mittal ◽

Matthew J. Rossi ◽

B. Franklin Pugh

Keyword(s):

Rna Polymerase Ii ◽

Transcriptional Regulators ◽

Micrococcal Nuclease ◽

High Similarity ◽

Dna Interactions ◽

Promoter Regions ◽

Genome Wide ◽

A Genome ◽

Protein Dna Interactions ◽

Negative Controls

AbstractChEC-seq is a method used to identify protein-DNA interactions across a genome. It involves fusing micrococcal nuclease (MNase) to a protein of interest. In principle, specific genome-wide interactions of the fusion protein with chromatin result in local DNA cleavages that can be mapped by DNA sequencing. ChEC-seq has been used to draw conclusions about broad gene-specificities of certain protein-DNA interactions. In particular, the transcriptional regulators SAGA, TFIID, and Mediator are reported to generally occupy the promoter/UAS of genes transcribed by RNA polymerase II in yeast. Here we compare published yeast ChEC-seq data performed with a variety of protein fusions across essentially all genes, and find high similarities with negative controls. We conclude that ChEC-seq patterning for SAGA, TFIID, and Mediator differ little from background at most promoter regions, and thus cannot be used to draw conclusions about broad gene specificity of these factors.

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Ligand-inducible retinoid X receptor-mediated protein: DNA interactions in the retinoic acid receptor β2 gene promoter in vivo

Molecular and Cellular Endocrinology ◽

10.1016/s0303-7207(97)00217-7 ◽

1998 ◽

Vol 136 (2) ◽

pp. 109-118 ◽

Cited By ~ 1

Author(s):

Masato Ikeda ◽

Remco A Spanjaard ◽

Elizabeth W Noordhoek ◽

Akio Kawaguchi ◽

Toshimasa Onaya ◽

...

Keyword(s):

Retinoic Acid ◽

Retinoic Acid Receptor ◽

Gene Promoter ◽

Retinoid X Receptor ◽

Dna Interactions ◽

Acid Receptor ◽

Protein Dna Interactions ◽

Retinoic Acid Receptor Β2

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Transcriptional role of a conserved GATA-1 site in the human epsilon-globin gene promoter.

Molecular and Cellular Biology ◽

10.1128/mcb.11.5.2558 ◽

1991 ◽

Vol 11 (5) ◽

pp. 2558-2566 ◽

Cited By ~ 41

Author(s):

Q H Gong ◽

J Stern ◽

A Dean

Keyword(s):

Globin Gene ◽

Point Mutations ◽

Gene Promoter ◽

Globin Genes ◽

Mobility Shift ◽

Dnase I Footprinting ◽

Nuclear Extracts ◽

Protein Dna Interactions ◽

Globin Promoter

The epsilon-globin gene is the first of the human beta-like globin genes to be expressed during development. We have analyzed protein-DNA interactions in the epsilon-globin promoter region by DNase I footprinting and electrophoretic mobility shift experiments using nuclear extracts from K562 human erythroid cells and from nonerythroid HeLa cells. A restricted set of ubiquitous proteins, including Sp1, bound to regions of the promoter including the CACCC and CCAAT sites. Three interactions, at positions -213, -165, and +3 relative to the transcription start site, were erythroid specific and corresponded to binding of GATA-1, a transcription factor highly restricted to the erythroid lineage. Interestingly, the GATA-1 site at -165 has been conserved in the promoters of 10 mammalian embryonic globin genes. Point mutations demonstrate that GATA-1 binding to this site is necessary for interaction with an erythroid-specific enhancer but that in the absence of an enhancer, GATA-1 does not increase transcription.

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