-
Jian Xu,
Zhen Shao,
Kimberly Glass,
Daniel E Bauer,
Luca Pinello,
Ben Van Handel,
Serena Hou,
John A Stamatoyannopoulos,
Hanna K A Mikkola,
Guo-Cheng Yuan,
Stuart H Orkin
[show abstract]
[hide abstract]
ABSTRACT: Gene-distal enhancers are critical for tissue-specific gene expression, but their genomic determinants within a specific lineage at different stages of development are unknown. Here we profile chromatin state maps, transcription factor occupancy, and gene expression profiles during human erythroid development at fetal and adult stages. Comparative analyses of human erythropoiesis identify developmental stage-specific enhancers as primary determinants of stage-specific gene expression programs. We find that erythroid master regulators GATA1 and TAL1 act cooperatively within active enhancers but confer little predictive value for stage specificity. Instead, a set of stage-specific coregulators collaborates with master regulators and contributes to differential gene expression. We further identify and validate IRF2, IRF6, and MYB as effectors of an adult-stage expression program. Thus, the combinatorial assembly of lineage-specific master regulators and transcriptional coregulators within developmental stage-specific enhancers determines gene expression programs and temporal regulation of transcriptional networks in a mammalian genome.
Developmental cell 10/2012; 23(4):796-811. · 13.36 Impact Factor
-
Jian Xu,
Cong Peng,
Vijay G Sankaran,
Zhen Shao,
Erica B Esrick,
Bryan G Chong,
Gregory C Ippolito,
Yuko Fujiwara,
Benjamin L Ebert,
Philip W Tucker,
Stuart H Orkin
[show abstract]
[hide abstract]
ABSTRACT: Persistence of human fetal hemoglobin (HbF, α(2)γ(2)) in adults lessens the severity of sickle cell disease (SCD) and the β-thalassemias. Here, we show that the repressor BCL11A is required in vivo for silencing of γ-globin expression in adult animals, yet dispensable for red cell production. BCL11A serves as a barrier to HbF reactivation by known HbF inducing agents. In a proof-of-principle test of BCL11A as a potential therapeutic target, we demonstrate that inactivation of BCL11A in SCD transgenic mice corrects the hematologic and pathologic defects associated with SCD through high-level pancellular HbF induction. Thus, interference with HbF silencing by manipulation of a single target protein is sufficient to reverse SCD.
Science 11/2011; 334(6058):993-6. · 31.20 Impact Factor
-
Vijay G Sankaran, Jian Xu,
Rachel Byron,
Harvey A Greisman,
Chris Fisher,
David J Weatherall,
Daniel E Sabath,
Mark Groudine,
Stuart H Orkin,
Anuja Premawardhena,
M A Bender
[show abstract]
[hide abstract]
ABSTRACT: An improved understanding of the regulation of the fetal hemoglobin genes holds promise for the development of targeted therapeutic approaches for fetal hemoglobin induction in the β-hemoglobinopathies. Although recent studies have uncovered trans-acting factors necessary for this regulation, limited insight has been gained into the cis-regulatory elements involved.
We identified three families with unusual patterns of hemoglobin expression, suggestive of deletions in the locus of the β-globin gene (β-globin locus). We performed array comparative genomic hybridization to map these deletions and confirmed breakpoints by means of polymerase-chain-reaction assays and DNA sequencing. We compared these deletions, along with previously mapped deletions, and studied the trans-acting factors binding to these sites in the β-globin locus by using chromatin immunoprecipitation.
We found a new (δβ)(0)-thalassemia deletion and a rare hereditary persistence of fetal hemoglobin deletion with identical downstream breakpoints. Comparison of the two deletions resulted in the identification of a small intergenic region required for γ-globin (fetal hemoglobin) gene silencing. We mapped a Kurdish β(0)-thalassemia deletion, which retains the required intergenic region, deletes other surrounding sequences, and maintains fetal hemoglobin silencing. By comparing these deletions and other previously mapped deletions, we elucidated a 3.5-kb intergenic region near the 5' end of the δ-globin gene that is necessary for γ-globin silencing. We found that a critical fetal hemoglobin silencing factor, BCL11A, and its partners bind within this region in the chromatin of adult erythroid cells.
By studying three families with unusual deletions in the β-globin locus, we identified an intergenic region near the δ-globin gene that is necessary for fetal hemoglobin silencing. (Funded by the National Institutes of Health and others.).
New England Journal of Medicine 09/2011; 365(9):807-14. · 53.30 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The molecular mechanisms underlying erythroid-specific gene regulation remain incompletely understood. Closely spaced binding sites for GATA, NF-E2/maf, and CACCC interacting transcription factors play functionally important roles in globin and other erythroid-specific gene expression. We and others recently identified the CACCC-binding transcription factor ZBP-89 as a novel GATA-1 and NF-E2/mafK interacting partner. Here, we examined the role of ZBP-89 in human globin gene regulation and erythroid maturation using a primary CD34(+) cell ex vivo differentiation system. We show that ZBP-89 protein levels rise dramatically during human erythroid differentiation and that ZBP-89 occupies key cis-regulatory elements within the globin and other erythroid gene loci. ZBP-89 binding correlates strongly with RNA Pol II occupancy, active histone marks, and high-level gene expression. ZBP-89 physically associates with the histone acetyltransferases p300 and Gcn5/Trrap, and occupies common sites with Gcn5 within the human globin loci. Lentiviral short hairpin RNAs knockdown of ZBP-89 results in reduced Gcn5 occupancy, decreased acetylated histone 3 levels, lower globin and erythroid-specific gene expression, and impaired erythroid maturation. Addition of the histone deacetylase inhibitor valproic acid partially reverses the reduced globin gene expression. These findings reveal an activating role for ZBP-89 in human globin gene regulation and erythroid differentiation.
Blood 08/2011; 118(13):3684-93. · 9.90 Impact Factor
-
The EMBO Journal 03/2011; 30(6):983-5. · 9.20 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The beta-thalassemia syndromes are a major global health problem. Increased levels of fetal hemoglobin (HbF) ameliorate the clinical symptoms seen in this disease. By taking advantage of the natural variation in the level of HbF in various populations, we and others identified several common genetic variants in three major loci that regulate HbF levels. One of these variants resides in the gene BCL11A. We have studied the role of this gene product and established that BCL11A maintains silencing of gamma-globin expression in adult erythroid cells and functions as a direct transcriptional regulator of the fetal to adult hemoglobin switch in humans. Moreover, we found that BCL11A plays a central role in the evolutionarily divergent globin gene switches of mammals. As a factor critical for gamma-globin gene silencing, BCL11A should be considered as a therapeutic target to increase HbF in a directed manner in beta-thalassemia patients.
Annals of the New York Academy of Sciences 08/2010; 1202:64-8. · 3.15 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The developmental switch from human fetal (gamma) to adult (beta) hemoglobin represents a clinically important example of developmental gene regulation. The transcription factor BCL11A is a central mediator of gamma-globin silencing and hemoglobin switching. Here we determine chromatin occupancy of BCL11A at the human beta-globin locus and other genomic regions in vivo by high-resolution chromatin immunoprecipitation (ChIP)-chip analysis. BCL11A binds the upstream locus control region (LCR), epsilon-globin, and the intergenic regions between gamma-globin and delta-globin genes. A chromosome conformation capture (3C) assay shows that BCL11A reconfigures the beta-globin cluster by modulating chromosomal loop formation. We also show that BCL11A and the HMG-box-containing transcription factor SOX6 interact physically and functionally during erythroid maturation. BCL11A and SOX6 co-occupy the human beta-globin cluster along with GATA1, and cooperate in silencing gamma-globin transcription in adult human erythroid progenitors. These findings collectively demonstrate that transcriptional silencing of gamma-globin genes by BCL11A involves long-range interactions and cooperation with SOX6. Our findings provide insight into the mechanism of BCL11A action and new clues for the developmental gene regulatory programs that function at the beta-globin locus.
Genes & development 04/2010; 24(8):783-98. · 12.08 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The study of haemoglobin switching has represented a focus in haematology due in large part to the clinical relevance of the fetal to adult haemoglobin switch for developing targeted approaches to ameliorate the severity of the beta-haemoglobinopathies. Additionally, the process by which this switch occurs represents an important paradigm for developmental gene regulation. In this review, we provide an overview of both the embryonic primitive to definitive switch in haemoglobin expression, as well as the fetal to adult switch that is unique to humans and old world monkeys. We discuss the nature of these switches and models of their regulation. The factors that have been suggested to regulate this process are then discussed. With the increased understanding and discovery of molecular regulators of haemoglobin switching, such as BCL11A, new avenues of research may lead ultimately to novel therapeutic, mechanism-based approaches to fetal haemoglobin reactivation in patients.
British Journal of Haematology 03/2010; 149(2):181-94. · 4.94 Impact Factor
-
Vijay G Sankaran, Jian Xu,
Tobias Ragoczy,
Gregory C Ippolito,
Carl R Walkley,
Shanna D Maika,
Yuko Fujiwara,
Masafumi Ito,
Mark Groudine,
M A Bender,
Philip W Tucker,
Stuart H Orkin
[show abstract]
[hide abstract]
ABSTRACT: The contribution of changes in cis-regulatory elements or trans-acting factors to interspecies differences in gene expression is not well understood. The mammalian beta-globin loci have served as a model for gene regulation during development. Transgenic mice containing the human beta-globin locus, consisting of the linked embryonic (epsilon), fetal (gamma) and adult (beta) genes, have been used as a system to investigate the temporal switch from fetal to adult haemoglobin, as occurs in humans. Here we show that the human gamma-globin (HBG) genes in these mice behave as murine embryonic globin genes, revealing a limitation of the model and demonstrating that critical differences in the trans-acting milieu have arisen during mammalian evolution. We show that the expression of BCL11A, a repressor of human gamma-globin expression identified by genome-wide association studies, differs between mouse and human. Developmental silencing of the mouse embryonic globin and human gamma-globin genes fails to occur in mice in the absence of BCL11A. Thus, BCL11A is a critical mediator of species-divergent globin switching. By comparing the ontogeny of beta-globin gene regulation in mice and humans, we have shown that alterations in the expression of a trans-acting factor constitute a critical driver of gene expression changes during evolution.
Nature 09/2009; 460(7259):1093-7. · 36.28 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Differences in the amount of fetal hemoglobin (HbF) that persists into adulthood affect the severity of sickle cell disease and the beta-thalassemia syndromes. Genetic association studies have identified sequence variants in the gene BCL11A that influence HbF levels. Here, we examine BCL11A as a potential regulator of HbF expression. The high-HbF BCL11A genotype is associated with reduced BCL11A expression. Moreover, abundant expression of full-length forms of BCL11A is developmentally restricted to adult erythroid cells. Down-regulation of BCL11A expression in primary adult erythroid cells leads to robust HbF expression. Consistent with a direct role of BCL11A in globin gene regulation, we find that BCL11A occupies several discrete sites in the beta-globin gene cluster. BCL11A emerges as a therapeutic target for reactivation of HbF in beta-hemoglobin disorders.
Science 01/2009; 322(5909):1839-42. · 31.20 Impact Factor
