Regulation of Chromatin Looping and Transcription by PRMT1 Mediated H4R3 Methylation.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1864-1864
Author(s):  
Xingguo Li ◽  
Xin Hu ◽  
Zhuo Zhou ◽  
Yi Qiu ◽  
Gary Felsenfeld ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Long Range ◽  
Transcriptional Activation ◽  
Globin Gene ◽  
Gene Promoter ◽  
Globin Genes ◽  
Functional Link ◽  
Chromatin Interactions ◽  
Globin Promoter ◽  
Transcription Complexes

Abstract Communication between distal enhancers and proximal promoters is critical in controlling proper transcription of genes. However, the functional link between certain histone modifications and the formation of long-range chromatin interactions involved in transcriptional activation remains unknown. In the globin locus, the b-globin genes are regulated by highly organized chromatin structure that juxtaposes the locus control region (LCR) located far upstream of the genes with the proximal b-major globin promoter (bmajpromoter). We report here that the localized asymmetric dimethylation of Arg3 at histone H4 tails (dimethyl H4R3) catalyzed by the methyltransferase PRMT1 is essential for establishing the long-range chromatin interactions between the LCR and the bmaj-promoter and strongly correlates with the activation of adult b-globin gene transcription. In addition, dimethyl H4R3 potentiates the recruitment of histone acetyltransferases (HATs), CBP and PCAF, and is required for the establishment of subsequent histone acetylation at the globin locus. Suppression of PRMT1 activity disrupts the recruitment of transcription complexes, TBP and RNA polymerase II (RNA Pol II), at the active b-globin promoter, but not at the LCR. Taken together, our data implicate PRMT1-mediated dimethylation of H4R3 in the regulation of long-range enhancer/promoter communications, which are required for the efficient recruitment of transcription complexes to the active gene promoter.

scholarly journals Structural and Functional Cross-Talk between a Distant Enhancer and the ɛ-Globin Gene Promoter Shows Interdependence of the Two Elements in Chromatin

1999 ◽  
Vol 19 (11) ◽  
pp. 7600-7609 ◽  
Author(s):  
Jennifer C. McDowell ◽  
Ann Dean
Keyword(s):  
Globin Gene ◽  
Gene Promoter ◽  
Transcription Activation ◽  
Tata Box ◽  
Globin Genes ◽  
Hypersensitive Site ◽  
Nuclease Digestion ◽  
Dnase I Hypersensitive Site ◽  
Globin Promoter ◽  
Gata Motif

ABSTRACT We investigated the requirements for enhancer-promoter communication by using the human β-globin locus control region (LCR) DNase I-hypersensitive site 2 (HS2) enhancer and the ɛ-globin gene in chromatinized minichromosomes in erythroid cells. Activation of globin genes during development is accompanied by localized alterations of chromatin structure, and CACCC binding factors and GATA-1, which interact with both globin promoters and the LCR, are believed to be critical for globin gene transcription activation. We found that an HS2 element mutated in its GATA motif failed to remodel the ɛ-globin promoter or activate transcription yet HS2 nuclease accessibility did not change. Accessibility and transcription were reduced at promoters with mutated GATA-1 or CACCC sites. Strikingly, these mutations also resulted in reduced accessibility at HS2. In the absence of a globin gene, HS2 is similarly resistant to nuclease digestion. In contrast to observations in Saccharomyces cerevisiae, HS2-dependent promoter remodeling was diminished when we mutated the TATA box, crippling transcription. This mutation also reduced HS2 accessibility. The results indicate that the ɛ-globin promoter and HS2 interact both structurally and functionally and that both upstream activators and the basal transcription apparatus contribute to the interaction. Further, at least in this instance, transcription activation and promoter remodeling by a distant enhancer are not separable.

scholarly journals Substitution of the Human β-Spectrin Promoter for the Human Aγ-Globin Promoter Prevents Silencing of a Linked Human β-Globin Gene in Transgenic Mice

1998 ◽  
Vol 18 (11) ◽  
pp. 6634-6640 ◽  
Author(s):  
Denise E. Sabatino ◽  
Amanda P. Cline ◽  
Patrick G. Gallagher ◽  
Lisa J. Garrett ◽  
George Stamatoyannopoulos ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Fetal Liver ◽  
Globin Gene ◽  
Gene Promoter ◽  
Yolk Sac ◽  
Globin Genes ◽  
Erythroid Cells ◽  
Globin Mrna ◽  
Globin Promoter ◽  
In Erythroid Cells

ABSTRACT During development, changes occur in both the sites of erythropoiesis and the globin genes expressed at each developmental stage. Previous work has shown that high-level expression of human β-like globin genes in transgenic mice requires the presence of the locus control region (LCR). Models of hemoglobin switching propose that the LCR and/or stage-specific elements interact with globin gene sequences to activate specific genes in erythroid cells. To test these models, we generated transgenic mice which contain the human Aγ-globin gene linked to a 576-bp fragment containing the human β-spectrin promoter. In these mice, the β-spectrin Aγ-globin (βsp/Aγ) transgene was expressed at high levels in erythroid cells throughout development. Transgenic mice containing a 40-kb cosmid construct with the micro-LCR, βsp/Aγ-, ψβ-, δ-, and β-globin genes showed no developmental switching and expressed both human γ- and β-globin mRNAs in erythroid cells throughout development. Mice containing control cosmids with the Aγ-globin gene promoter showed developmental switching and expressed Aγ-globin mRNA in yolk sac and fetal liver erythroid cells and β-globin mRNA in fetal liver and adult erythroid cells. Our results suggest that replacement of the γ-globin promoter with the β-spectrin promoter allows the expression of the β-globin gene. We conclude that the γ-globin promoter is necessary and sufficient to suppress the expression of the β-globin gene in yolk sac erythroid cells.

Manipulating Higher Order Chromatin Structure of the β-Globin Locus by Targeted Tethering of a “looping” Factor

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 647-647
Author(s):  
Wulan Deng ◽  
Philip D Gregory ◽  
Andreas Reik ◽  
Gerd Blobel
Keyword(s):  
Transcription Factor ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Conflicts Of Interest ◽  
Chromatin Loop ◽  
Globin Genes ◽  
Erythroid Cells ◽  
Loop Formation ◽  
Pol Ii ◽  
Globin Promoter

Abstract Abstract 647 The mammalian β-globin locus is under the coordinated control of multiple transcription factors to ensure the correct expression of the globin genes during development. The distal β-globin locus control region (LCR) physically interacts with β-like globin promoters to form developmentally dynamic chromatin loops. The hematopoietic transcription factor GATA-1 and its associated cofactor Ldb1 bind to the LCR and the β-globin promoter and are essential for loop formation and β-globin expression. However, the molecular basis of chromatin looping and its cause-effect relationship with transcriptional activation are unclear. Here, we examined whether Ldb1 is an effector of GATA-1 during loop formation. Specifically, we tested whether artificial tethering of Ldb1 to the endogenous β-globin promoter and LCR can substitute for GATA-1 function. Ldb1 was fused to artificial zinc finger proteins (ZFP) designed to bind to the LCR and β-globin promoter. Ldb1-ZFPs were introduced pairwise into murine GATA-1 null erythroid cells in which the β-globin locus is relaxed and transcriptionally silent. In vivo binding of the Ldb1-ZFPs to their targets was verified by ChIP assay. Strikingly, expression of Ldb1-ZFPs but not Ldb1 or ZFPs alone led to substantial activation of β-globin transcription in the absence of GATA-1. Moreover, chromosome conformation capture experiments showed that Ldb1-ZFPs triggered physical association between the LCR and the β-globin promoter. Recruitment of RNA polymerase II (Pol II) and its phosphorylation at serine 5 are critical LCR-dependent regulatory steps in β-globin transcription. We found that Ldb1-ZFP expression facilitated Pol II recruitment at the β-globin promoter and serine 5 phosphorylation to the same level as GATA-1-expressing erythroid cells. This is consistent with an Ldb1-ZFP-induced LCR-β-globin promoter chromatin loop. In concert, these results indicate that Ldb1 is a critical effector for GATA-1 by mediating enhancer-promoter loops. In broader terms, our results suggest that chromatin loop formation can be sufficient for gene activation in the absence of an essential transcription factor. We are currently in the process of examining whether targeting of the LCR to embryonic and fetal globin genes can be used to activate them in adult cells. Targeted chromatin loop formation may provide a method to activate fetal or adult hemoglobin expression in individuals with β-thalassemia or sickle cell anemia. Disclosures: No relevant conflicts of interest to declare.

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

1991 ◽  
Vol 11 (5) ◽  
pp. 2558-2566 ◽  
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.

Methylation of α-type embryonic globin gene απrepresses transcription in primary erythroid cells

Blood ◽  
2002 ◽  
Vol 100 (12) ◽  
pp. 4217-4222 ◽  
Author(s):  
Rakesh Singal ◽  
Jane M. vanWert ◽  
Larry Ferdinand
Keyword(s):  
Globin Gene ◽  
Gene Promoter ◽  
Luciferase Reporter ◽  
Globin Genes ◽  
Erythroid Cells ◽  
Histone H4 ◽  
Luciferase Reporter Gene ◽  
Cpg Dinucleotides ◽  
Expression Construct ◽  
Globin Promoter

The inverse relationship between expression and methylation of β-type globin genes is well established. However, little is known about the relationship between expression and methylation of avian α-type globin genes. The embryonicαπ-globin promoter was unmethylated, andαπ-globin RNA was easily detected in 5-day chicken erythroid cells. A progressive methylation of the CpG dinucleotides in the απ promoter associated with loss of expression of απ-globin gene was seen during development in primary erythroid cells. A 315-bpαπ-globin promoter region was cloned in an expression construct (απpGL3E) containing a luciferase reporter gene and SV40 enhancer. The απpGL3E construct was transfected into primary erythroid cells derived from 5-day-old chicken embryos. Methylation of απpGL3E plasmid andαπ-globin promoter alone resulted in a 20-fold and 7-fold inhibition of expression, respectively. The fully methylated but not the unmethylated 315-bpαπ-globin gene promoter fragment formed amethyl cytosine-binding proteincomplex (MeCPC). Chromatin immunoprecipitation assays were combined with quantitative real-time polymerase chain reaction to assess histone acetylation associated with theαπ-globin gene promoter. Slight hyperacetylation of histone H3 but a marked hyperacetylation of histone H4 was seen in 5-day when compared with 14-day erythroid cells. These results demonstrate that methylation can silence transcription of an avian α-type embryonic globin gene in homologous primary erythroid cells, possibly by interacting with an MeCPC and histone deacetylase complex.

A Facilitated Tracking and Transcription Mechanism of the HS2 Enhancer in the β-Globin Locus Control Region.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1596-1596
Author(s):  
Dorothy Tuan ◽  
Xingguo Zhu ◽  
Jianhua Ling ◽  
Ling Zhang ◽  
Wenhu Pi ◽  
...  
Keyword(s):  
Long Range ◽  
Control Region ◽  
Gene Locus ◽  
Globin Gene ◽  
Locus Control Region ◽  
Globin Genes ◽  
Long Distance ◽  
Pol Ii ◽  
Enhancer Function ◽  
Globin Promoter

Abstract The 100 kb human β-globin gene locus contains the embryonic ε-, the fetal Gγ- and Aγ-, and the adult δ- and β-globin genes arranged in the transcriptional order of 5′ ε-Gγ-Aγ-δ-β 3′. The β-globin Locus Control Region (LCR), defined by DNase I hypersensitive sites HS1, 2, 3, 4 and 5 located far upstream of the globin genes, is absolutely required for transcriptional activation of the globin genes in erythroid cells. The HS2 site, located respectively 11 and 55 kb upstream of the ε- and β-globin genes, possesses prominent enhancer activity and is able to activate transcription of the globin genes over the long distance. How the HS2 enhancer acts over the long distance is not fully understood. Three mechanisms have been proposed to explain long-range enhancer function: The looping model, the protein tracking/linking model and the facilitated tracking model. We have shown earlier that a transcription mechanism of the RNA polymerase II (pol II)-complex assembled by the HS2 enhancer mediates long-range HS2 enhancer function. In the present study, we used the chicken HS4 insulator to further study the mechanism of long-range HS2 enhancer function. We created the LCR(+I) and LCR(−I) lines in human erythroid K562 cells using Cre-loxP mediated in situ recombination and in zebrafish. The integrated LCR(+I) and LCR(−I) plasmids spanned the natural 11 kb human ε-globin gene locus either with or without the chicken HS4 insulator inserted between the LCR HS2 enhancer and the ε-globin promoter. Analysis of enhancer function in the integrated plasmids and the endogenous ε-globin gene locus provided evidence for a novel facilitated tracking and transcription mechanism of long-range enhancer function: The HS2 enhancer complex containing both the enhancer DNA and the associated pol II tracked and transcribed through the 10 kb intervening DNA to loop with and activate the ε-globin promoter. The interposed insulator interrupted this facilitated tracking and transcription mechanism of the enhancer complex through the intervening DNA, thereby blocking long-range enhancer function.

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

1991 ◽  
Vol 11 (5) ◽  
pp. 2558-2566
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.

scholarly journals Defective Erythropoiesis in Transgenic Mice Expressing Dominant-Negative Upstream Stimulatory Factor

2009 ◽  
Vol 29 (21) ◽  
pp. 5900-5910 ◽  
Author(s):  
Shermi Y. Liang ◽  
Babak Moghimi ◽  
Valerie J. Crusselle-Davis ◽  
I-Ju Lin ◽  
Michael H. Rosenberg ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Transgenic Mice ◽  
Rna Polymerase Ii ◽  
Globin Gene ◽  
Dominant Negative ◽  
Erythroid Cell ◽  
Dominant Negative Mutant ◽  
Globin Genes ◽  
Erythroid Cells ◽  
Transcription Complexes

ABSTRACT Transcription factor USF is a ubiquitously expressed member of the helix-loop-helix family of proteins. It binds with high affinity to E-box elements and, through interaction with coactivators, aids in the formation of transcription complexes. Previous work demonstrated that USF regulates genes during erythroid differentiation, including HoxB4 and β-globin. Here, we show that the erythroid cell-specific expression of a dominant-negative mutant of USF, A-USF, in transgenic mice reduces the expression of all β-type globin genes and leads to the diminished association of RNA polymerase II with locus control region element HS2 and with the β-globin gene promoter. We further show that the expression of A-USF reduces the expression of several key erythroid cell-specific transcription factors, including EKLF and Tal-1. We provide evidence demonstrating that USF interacts with known regulatory DNA elements in the EKLF and Tal-1 gene loci in erythroid cells. Furthermore, A-USF-expressing transgenic mice exhibit a defect in the formation of CD71+ progenitor and Ter-119+ mature erythroid cells. In summary, the data demonstrate that USF regulates globin gene expression indirectly by enhancing the expression of erythroid transcription factors and directly by mediating the recruitment of transcription complexes to the globin gene locus.

Human globin gene promoter sequences are sufficient for specific expression of a hybrid gene transfected into tissue culture cells

1987 ◽  
Vol 7 (1) ◽  
pp. 398-402
Author(s):  
T Rutherford ◽  
A W Nienhuis
Keyword(s):  
Gene Expression ◽  
Globin Gene ◽  
Gene Promoter ◽  
Gene Promoters ◽  
Globin Genes ◽  
Specific Expression ◽  
Tissue Specific ◽  
Promoter Sequences ◽  
Erythroleukemia Cells ◽  
Murine Erythroleukemia

The contribution of the human globin gene promoters to tissue-specific transcription was studied by using globin promoters to transcribe the neo (G418 resistance) gene. After transfection into different cell types, neo gene expression was assayed by scoring colony formation in the presence of G418. In K562 human erythroleukemia cells, which express fetal and embryonic globin genes but not the adult beta-globin gene, the neo gene was expressed strongly from a fetal gamma- or embryonic zeta-globin gene promoter but only weakly from the beta promoter. In murine erythroleukemia cells which express the endogenous mouse beta genes, the neo gene was strongly expressed from both beta and gamma promoters. In two nonerythroid cell lines, human HeLa cells and mouse 3T3 fibroblasts, the globin gene promoters did not allow neo gene expression. Globin-neo genes were integrated in the erythroleukemia cell genomes mostly as a single copy per cell and were transcribed from the appropriate globin gene cap site. We conclude that globin gene promoter sequences extending from -373 to +48 base pairs (bp) (relative to the cap site) for the beta gene, -385 to +34 bp for the gamma gene, and -555 to +38 bp for the zeta gene are sufficient for tissue-specific and perhaps developmentally specific transcription.

scholarly journals Sardinian G gamma-HPFH: a T----C substitution in a conserved "octamer" sequence in the G gamma-globin promoter

Blood ◽  
1988 ◽  
Vol 71 (3) ◽  
pp. 815-817 ◽  
Author(s):  
S Ottolenghi ◽  
S Nicolis ◽  
R Taramelli ◽  
N Malgaretti ◽  
R Mantovani ◽  
...  
Keyword(s):  
Globin Gene ◽  
Single Point ◽  
Point Mutations ◽  
Fetal Hemoglobin ◽  
Regulatory Elements ◽  
Globin Genes ◽  
Gamma Globin ◽  
New Type ◽  
Hereditary Persistence ◽  
Globin Promoter

Abstract A survey of hemoglobinopathies in Northern Sardinia allowed the identification of two subjects heterozygous for a new type of G gamma hereditary persistence of fetal hemoglobin (HPFH). The G gamma-globin gene from the HPFH chromosome shows the presence of a T----C substitution 175 nucleotides upstream of the CAP site, adding a new example of single-point mutations occurring in the promoter region of the gamma-globin genes and linked to HPFH phenotypes. In this case the mutation affects the 3′ end nucleotide of a conserved octamer sequence known to be present in other regulatory elements of several genes.

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