HDAC1 is required for GATA-1 transcription activity, global chromatin occupancy and hematopoiesis.

The activity of hematopoietic factor GATA-1 is modulated through p300/CBP-mediated acetylation and FOG-1 mediated indirect interaction with HDAC1/2 containing NuRD complex. Although GATA-1 acetylation is implicated in GATA-1 activation, the role of deacetylation is not studied. Here, we found that the FOG-1/NuRD does not deacetylate GATA-1. However, HDAC1/2 can directly bind and deacetylate GATA-1. Two arginine residues within the GATA-1 linker region mediates direct interaction with HDAC1. The arginine to alanine mutation (2RA) blocks GATA-1 deacetylation and fails to induce erythroid differentiation. Gene expression profiling and ChIP-seq analysis further demonstrate the importance of GATA-1 deacetylation for gene activation and chromatin recruitment. GATA-12RA knock-in (KI) mice suffer mild anemia and thrombocytopenia with accumulation of immature erythrocytes and megakaryocytes in bone marrow and spleen. Single cell RNA-seq analysis of Lin- cKit+ (LK) cells further reveal a profound change in cell subpopulations and signature gene expression patterns in HSC, myeloid progenitors, and erythroid/megakaryocyte clusters in KI mice. Thus, GATA-1 deacetylation and its interaction with HDAC1 modulates GATA-1 chromatin binding and transcriptional activity that control erythroid/megakaryocyte commitment and differentiation.

Diamond-Blackfan anemia.

Diamond-Blackfan anemia (DBA) was the first ribosomopathy described and is a constitutional inherited bone marrow failure syndrome. Erythroblastopenia is the major characteristic of the disease, which is a model for ribosomal diseases, related to a heterozygous allelic variation in 1 of the 20 ribosomal protein genes of either the small or large ribosomal subunit. The salient feature of classical DBA is a defect in ribosomal RNA maturation that generates nucleolar stress, leading to stabilization of p53 and activation of its targets, resulting in cell-cycle arrest and apoptosis. Although activation of p53 may not explain all aspects of DBA erythroid tropism, involvement of GATA1/HSP70 and globin/heme imbalance, with an excess of the toxic free heme leading to reactive oxygen species production, account for defective erythropoiesis in DBA. Despite significant progress in defining the molecular basis of DBA and increased understanding of the mechanistic basis for DBA pathophysiology, progress in developing new therapeutic options has been limited. However, recent advances in gene therapy, better outcomes with stem cell transplantation, and discoveries of putative new drugs through systematic drug screening using large chemical libraries provide hope for improvement.

Novel targets to cure primary myelofibrosis from studies on Gata1low mice.

In 2002, we discovered that mice carrying the hypomorphic Gata1low mutation that reduces expression of the transcription factor GATA1 in megakaryocytes (Gata1low mice) develop myelofibrosis, a phenotype that recapitulates the features of primary myelofibrosis (PMF), the most severe of the Philadelphia-negative myeloproliferative neoplasms (MPNs). At that time, this discovery had a great impact on the field because mutations driving the development of PMF had yet to be discovered. Later studies identified that PMF, as the others MPNs, is associated with mutations activating the thrombopoietin/JAK2 axis raising great hope that JAK inhibitors may be effective to treat the disease. Unfortunately, ruxolitinib, the JAK1/2 inhibitor approved by FDA and EMEA for PMF, ameliorates symptoms but does not improve the natural course of the disease, and the cure of PMF is still an unmet clinical need. Although GATA1 is not mutated in PMF, reduced GATA1 content in megakaryocytes as a consequence of ribosomal deficiency is a hallmark of myelofibrosis (both in humans and mouse models) and, in fact, a driving event in the disease. Conversely, mice carrying the hypomorphic Gata1low mutation express an activated TPO/JAK2 pathway and partially respond to JAK inhibitors in a fashion similar to PMF patients (reduction of spleen size but limited improvement of the natural history of the disease). These observations cross-validated Gata1low mice as a bona fide animal model for PMF and prompted the use of this model to identify abnormalities that might be targeted to cure the disease. We will summarize here data generated in Gata1low mice indicating that the TGF-ß/P-selectin axis is abnormal in PMF and represents a novel target for its treatment.

The expression of CKLFSF2B is regulated by GATA1 and CREB in the Leydig cells, which modulates testicular steroidogenesis.

CKLFSF is a protein family that serves as a functional bridge between chemokines and members of the transmembrane 4 superfamily (TM4SF). In the course of evolution, CKLFSF2 has evolved as two isoforms, namely CKLFSF2A and CKLFSF2B, in mice. CKLFSF2A, also known as CMTM2A and ARR19, is expressed in the testis and is important for testicular steroidogenesis. CKLFSF2B is also known to be highly expressed in the testis. In the prepubertal stage, CKLFSF2B is expressed only in Leydig cells, but it is highly expressed in haploid germ cells and Leydig cells in adult testis. CKLFSF2B is naturally processed inside the cell at its C-terminus to yield smaller proteins compared to its theoretical size of ≈25 kDa. The Cklfsf2b gene is regulated by GATA-1 and CREB protein, binding to their respective binding elements present in the 2-kb upstream promoter sequence. In addition, the overexpression of CKLFSF2B inhibited the activity of the Nur77 promoter, which consequently represses the promoter activity of Nur77-target steroidogenic genes such as P450c17, 3ß-HSD, and StAR in MA-10 Leydig cells. Adenovirus-mediated overexpression of CKLFSF2B in primary Leydig cells isolated from adult mice shows a repression of steroidogenic gene expression and consequently testosterone production. Moreover, intratesticular injection of CKLFSF2B-expressing adenovirus in adult mice clearly had a repressive effect compared to the control injected with only GFP-expressing adenovirus. Altogether, these findings suggest that CKLFSF2B might be involved in the development and function of Leydig cells and regulate testicular testosterone production by fine-tuning the expression of steroidogenic genes.

Perturbed MafB/GATA1 axis after burn trauma bares the potential mechanism for immune suppression and anemia of critical illness.

Patients who survive initial burn injury are susceptible to nosocomial infections. Anemia of critical illness is a compounding factor in burn patients that necessitates repeated transfusions, which further increase their susceptibility to infections and sepsis. Robust host response is dependent on an adequate number and function of monocytes/macrophages and dendritic cells. In addition to impaired RBC production, burn patients are prone to depletion of dendritic cells and an increase in deactivated monocytes. In steady-state hematopoiesis, RBCs, macrophages, and dendritic cells are all generated from a common myeloid progenitor within the bone marrow. We hypothesized in a mouse model of burn injury that an increase in myeloid-specific transcription factor V-maf musculoaponeurotic fibrosarcoma oncogene homolog B at the common myeloid progenitor stage steers their lineage potential away from the megakaryocyte erythrocyte progenitor production and drives the terminal fate of common myeloid progenitors to form macrophages vs. dendritic cells, with the consequences being anemia, monocytosis, and dendritic cell deficits. Results indicate that, even though burn injury stimulated bone marrow hematopoiesis by increasing multipotential stem cell production (LinnegSca1poscKitpos), the bone marrow commitment is shifted away from the megakaryocyte erythrocyte progenitor and toward granulocyte monocyte progenitors with corresponding alterations in peripheral blood components, such as hemoglobin, hematocrit, RBCs, monocytes, and granulocytes. Furthermore, burn-induced V-maf musculoaponeurotic fibrosarcoma oncogene homolog B in common myeloid progenitors acts as a transcriptional activator of M-CSFR and a repressor of transferrin receptors, promoting macrophages and inhibiting erythroid differentiations while dictating a plasmacytoid dendritic cell phenotype. Results from small interfering RNA and gain-of-function (gfp-globin transcription factor 1 retrovirus) studies indicate that targeted interventions to restore V-maf musculoaponeurotic fibrosarcoma oncogene homolog B/globin transcription factor 1 balance can mitigate both immune imbalance and anemia of critical illness.

Pluripotent stem cells reveal erythroid-specific activities of the GATA1 N-terminus.

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.

Hematopoietic transcriptional mechanisms: from locus-specific to genome-wide vantage points.

Hematopoiesis is an exquisitely regulated process in which stem cells in the developing embryo and the adult generate progenitor cells that give rise to all blood lineages. Master regulatory transcription factors control hematopoiesis by integrating signals from the microenvironment and dynamically establishing and maintaining genetic networks. One of the most rudimentary aspects of cell type-specific transcription factor function, how they occupy a highly restricted cohort of cis-elements in chromatin, remains poorly understood. Transformative technologic advances involving the coupling of next-generation DNA sequencing technology with the chromatin immunoprecipitation assay (ChIP-seq) have enabled genome-wide mapping of factor occupancy patterns. However, formidable problems remain; notably, ChIP-seq analysis yields hundreds to thousands of chromatin sites occupied by a given transcription factor, and only a fraction of the sites appear to be endowed with critical, non-redundant function. It has become en vogue to map transcription factor occupancy patterns genome-wide, while using powerful statistical tools to establish correlations to inform biology and mechanisms. With the advent of revolutionary genome editing technologies, one can now reach beyond correlations to conduct definitive hypothesis testing. This review focuses on key discoveries that have emerged during the path from single loci to genome-wide analyses, specifically in the context of hematopoietic transcriptional mechanisms.

Transcription factor GATA1 is dispensable for mast cell differentiation in adult mice.

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(+) FcεRIα(+) 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.

Preclinical models for drug selection in myeloproliferative neoplasms.

The discovery that an abnormally activated JAK-STAT signaling pathway is central to the pathogenesis of myeloproliferative neoplasms has promoted the clinical development of small-molecule JAK2 inhibitors. These agents have shown remarkable efficacy in disease control, but do not induce molecular remission; on the other hand, interferon holds the promise to target the putative hematopoietic progenitor cell initiating the disease. The presence of additional molecular abnormalities indicates a high molecular complexity of myeloproliferative neoplasms, and the need for simultaneously targeting different targets. Several drugs are currently under study as single agents and in combination. This review briefly describes the several in vitro and in vivo models of myeloproliferative neoplasms that are being used as preclinical models for drug development.

The Gata1 5' region harbors distinct cis-regulatory modules that direct gene activation in erythroid cells and gene inactivation in HSCs.

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.

Characterization of the TGF-ß1 signaling abnormalities in the Gata1low mouse model of myelofibrosis.

Primary myelofibrosis (PMF) is characterized by fibrosis, ineffective hematopoiesis in marrow, and hematopoiesis in extramedullary sites and is associated with abnormal megakaryocyte (MK) development and increased transforming growth factor (TGF)-ß1 release. To clarify the role of TGF-ß1 in the pathogenesis of this disease, the TGF-ß1 signaling pathway of marrow and spleen of the Gata1(low) mouse model of myelofibrosis (MF) was profiled and the consequences of inhibition of TGF-ß1 signaling on disease manifestations determined. The expression of 20 genes in marrow and 36 genes in spleen of Gata1(low) mice was altered. David-pathway analyses identified alterations of TGF-ß1, Hedgehog, and p53 signaling in marrow and spleen and of mammalian target of rapamycin (mTOR) in spleen only and predicted that these alterations would induce consequences consistent with the Gata1(low) phenotype (increased apoptosis and G1 arrest both in marrow and spleen and increased osteoblast differentiation and reduced ubiquitin-mediated proteolysis in marrow only). Inhibition of TGF-ß1 signaling normalized the expression of p53-related genes, restoring hematopoiesis and MK development and reducing fibrosis, neovascularization, and osteogenesis in marrow. It also normalized p53/mTOR/Hedgehog-related genes in spleen, reducing extramedullary hematopoiesis. These data identify altered expression signatures of TGF-ß1 signaling that may be responsible for MF in Gata1(low) mice and may represent additional targets for therapeutic intervention in PMF.

Identification of biologically relevant enhancers in human erythroid cells.

Identification of cell type-specific enhancers is important for understanding the regulation of programs controlling cellular development and differentiation. Enhancers are typically marked by the co-transcriptional activator protein p300 or by groups of cell-expressed transcription factors. We hypothesized that a unique set of enhancers regulates gene expression in human erythroid cells, a highly specialized cell type evolved to provide adequate amounts of oxygen throughout the body. Using chromatin immunoprecipitation followed by massively parallel sequencing, genome-wide maps of candidate enhancers were constructed for p300 and four transcription factors, GATA1, NF-E2, KLF1, and SCL, using primary human erythroid cells. These data were combined with gene expression analyses, and candidate enhancers were identified. Consistent with their predicted function as candidate enhancers, there was statistically significant enrichment of p300 and combinations of co-localizing erythroid transcription factors within 1-50 kb of the transcriptional start site (TSS) of genes highly expressed in erythroid cells. Candidate enhancers were also enriched near genes with known erythroid cell function or phenotype. Candidate enhancers exhibited moderate conservation with mouse and minimal conservation with nonplacental vertebrates. Candidate enhancers were mapped to a set of erythroid-associated, biologically relevant, SNPs from the genome-wide association studies (GWAS) catalogue of NHGRI, National Institutes of Health. Fourteen candidate enhancers, representing 10 genetic loci, mapped to sites associated with biologically relevant erythroid traits. Fragments from these loci directed statistically significant expression in reporter gene assays. Identification of enhancers in human erythroid cells will allow a better understanding of erythroid cell development, differentiation, structure, and function and provide insights into inherited and acquired hematologic disease.

NF-Y recruits both transcription activator and repressor to modulate tissue- and developmental stage-specific expression of human γ-globin gene.

The human embryonic, fetal and adult ß-like globin genes provide a paradigm for tissue- and developmental stage-specific gene regulation. The fetal γ-globin gene is expressed in fetal erythroid cells but is repressed in adult erythroid cells. The molecular mechanism underlying this transcriptional switch during erythroid development is not completely understood. Here, we used a combination of in vitro and in vivo assays to dissect the molecular assemblies of the active and the repressed proximal γ-globin promoter complexes in K562 human erythroleukemia cell line and primary human fetal and adult erythroid cells. We found that the proximal γ-globin promoter complex is assembled by a developmentally regulated, general transcription activator NF-Y bound strongly at the tandem CCAAT motifs near the TATA box. NF-Y recruits to neighboring DNA motifs the developmentally regulated, erythroid transcription activator GATA-2 and general repressor BCL11A, which in turn recruit erythroid repressor GATA-1 and general repressor COUP-TFII to form respectively the NF-Y/GATA-2 transcription activator hub and the BCL11A/COUP-TFII/GATA-1 transcription repressor hub. Both the activator and the repressor hubs are present in both the active and the repressed γ-globin promoter complexes in fetal and adult erythroid cells. Through changes in their levels and respective interactions with the co-activators and co-repressors during erythroid development, the activator and the repressor hubs modulate erythroid- and developmental stage-specific transcription of γ-globin gene.

Protein kinase D-HDAC5 signaling regulates erythropoiesis and contributes to erythropoietin cross-talk with GATA1.

In red cell development, the differentiation program directed by the transcriptional regulator GATA1 requires signaling by the cytokine erythropoietin, but the mechanistic basis for this signaling requirement has remained unknown. Here we show that erythropoietin regulates GATA1 through protein kinase D activation, promoting histone deacetylase 5 (HDAC5) dissociation from GATA1, and subsequent GATA1 acetylation. Mice deficient for HDAC5 show resistance to anemic challenge and altered marrow responsiveness to erythropoietin injections. In ex vivo studies, HDAC5(-/-) progenitors display enhanced entry into and passage through the erythroid lineage, as well as evidence of erythropoietin-independent differentiation. These results reveal a molecular pathway that contributes to cytokine regulation of hematopoietic differentiation and offer a potential mechanism for fine tuning of lineage-restricted transcription factors by lineage-specific cytokines.

High MafB expression following burn augments monocyte commitment and inhibits DC differentiation in hemopoietic progenitors.

We have previously shown that perturbed bone marrow progenitor development promotes hyporesponsive monocytes following experimental burn sepsis. Clinical and experimental sepsis is associated with monocyte deactivation and depletion of mDCs. Decrease in circulating DCs is reported in burn patients who develop sepsis. In our 15% TBSA scald burn model, we demonstrate a significant reduction in the circulating MHC-II(+) population and mDCs (Gr1(neg)CD11b(+)CD11c(+)) with a corresponding decrease in bone marrow MHC-II(+) cells and mDCs for up to 14 days following burn. We explored the underlying mechanism(s) that regulate bone marrow development of monocytes and DCs following burn injury. We found a robust bone marrow response with a significant increase in multipotential HSCs (LSK) and bipotential GMPs following burn injury. GMPs from burn mice exhibit a significant reduction in GATA-1, which is essential for DC development, but express high levels of MafB and M-CSFRs, both associated with monocyte production. GMPs obtained from burn mice differentiated 1.7 times more into Mϕ and 1.6-fold less into DCs compared with sham. Monocytes and DCs expressed 50% less MHC-II in burn versus sham. Increased monocyte commitment in burn GMPs was a result of high MafB and M-CSFR expressions. Transient silencing of MafB (siRNA) in GMP-derived monocytes from burn mice partially restored DC differentiation deficits and increased GATA-1 expression. We provide evidence that high MafB following burn plays an inhibitory role in monocyte-derived DC differentiation by regulating M-CSFR and GATA-1 expressions.

FOG-1 and GATA-1 act sequentially to specify definitive megakaryocytic and erythroid progenitors.

The transcription factors that control lineage specification of haematopoietic stem cells (HSCs) have been well described for the myeloid and lymphoid lineages, whereas transcriptional control of erythroid (E) and megakaryocytic (Mk) fate is less understood. We here use conditional removal of the GATA-1 and FOG-1 transcription factors to identify FOG-1 as required for the formation of all committed Mk- and E-lineage progenitors, whereas GATA-1 was observed to be specifically required for E-lineage commitment. FOG-1-deficient HSCs and preMegEs, the latter normally bipotent for the Mk and E lineages, underwent myeloid transcriptional reprogramming, and formed myeloid, but not erythroid and megakaryocytic cells in vitro. These results identify FOG-1 and GATA-1 as required for formation of bipotent Mk/E progenitors and their E-lineage commitment, respectively, and show that FOG-1 mediates transcriptional Mk/E programming of HSCs as well as their subsequent Mk/E-lineage commitment. Finally, C/EBPs and FOG-1 exhibited transcriptional cross-regulation in early myelo-erythroid progenitors making their functional antagonism a potential mechanism for separation of the myeloid and Mk/E lineages.

NUP98-HOXA9-transgenic zebrafish develop a myeloproliferative neoplasm and provide new insight into mechanisms of myeloid leukaemogenesis.

NUP98-HOXA9 [t(7;11) (p15;p15)] is associated with inferior prognosis in de novo and treatment-related acute myeloid leukaemia (AML) and contributes to blast crisis in chronic myeloid leukaemia (CML). We have engineered an inducible transgenic zebrafish harbouring human NUP98-HOXA9 under the zebrafish spi1(pu.1) promoter. NUP98-HOXA9 perturbed zebrafish embryonic haematopoiesis, with upregulated spi1 expression at the expense of gata1a. Markers associated with more differentiated myeloid cells, lcp1, lyz, and mpx were also elevated, but to a lesser extent than spi1, suggesting differentiation of early myeloid progenitors may be impaired by NUP98-HOXA9. Following irradiation, NUP98-HOXA9-expressing embryos showed increased numbers of cells in G2-M transition compared to controls and absence of a normal apoptotic response, which may result from an upregulation of bcl2. These data suggest NUP98-HOXA9-induced oncogenesis may result from a combination of defects in haematopoiesis and an aberrant response to DNA damage. Importantly, 23% of adult NUP98-HOXA9-transgenic fish developed a myeloproliferative neoplasm (MPN) at 19-23 months of age. In summary, we have identified an embryonic haematopoietic phenotype in a transgenic zebrafish line that subsequently develops MPN. This tool provides a unique opportunity for high-throughput in vivo chemical modifier screens to identify novel therapeutic agents in high risk AML.

Networking erythropoiesis.

A relatively small cadre of lineage-restricted transcription factors largely orchestrates erythropoiesis, but how these nuclear factors interact to regulate this complex biology is still largely unknown. However, recent technological advances, such as chromatin immunoprecipitation (ChIP) paired with massively parallel sequencing (ChIP-seq), gene expression profiling, and comprehensive bioinformatic analyses, offer new insights into the intricacies of red cell molecular circuits.

The transcription factor GATA-1 is overexpressed in breast carcinomas and contributes to survivin upregulation via a promoter polymorphism.

Expression of survivin, a member of the inhibitor of apoptosis protein family, is elevated in human cancers and considered as a new therapeutic target. Mechanism upregulating survivin expression in tumour cells is poorly understood. In this study, we show that breast cancer patients harbouring a polymorphism G235A in the survivin promoter present a higher level of survivin expression. This polymorphism creates a binding site for the transcription factor GATA-1 inducing a second GATA-1-binding site in survivin promoter. At the mRNA level, GATA-1 was present in breast carcinomas and adjacent normal tissues, whereas the protein was only detected in carcinomas by western blot and immunohistochemistry. Transfection of wild-type and different constitutively active GATA-1 mutants (serine 26, 178 or 310) showed that only phospho-serine 26 GATA-1 was able to increase survivin expression. This increase was higher in G235A than in G235G cell lines. Phospho-serine 26 GATA-1 bound directly survivin promoter, with a stronger interaction in G235A than in G235G polymorphism indicating that both GATA-1-binding sites are functional. These data identify GATA-1 as a key feature in tumour aggressiveness by enhancing survivin expression and delineate its targeting as a possible new therapeutic strategy in breast carcinomas.