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Suming Huang - One of the best experts on this subject based on the ideXlab platform.
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The TAL1 Short Isoform Generated By PRMT1-Mediated Alternative RNA Splicing Promotes Erythroid Differentiation
Blood, 2016Co-Authors: Ngoc-tung Tran, Hairui Su, Suming Huang, Xinyang ZhaoAbstract:The isoforms of key transcription factors in hematopoiesis such as TAL1, GATA1 and RUNX1 are generated through alternative RNA splicing regulated by the PRMT1-RBM15 axis (Zhang et al. 2015). The functions of short isoforms of GATA1 (GATA1s) and RUNX1 (RUNX1a) are well characterized, yet it is unknown how the short isoform of TAL1 (TAL1s) regulates hematopoiesis. In this presentation, we report that the short isoform of TAL1, i.e. TAL1s, is generated via alternative RNA splicing as detected by isoform specific real-time PCR reactions using RNA isolated from leukemia cell lines and primary human cord blood cells. RBM15, an RNA binding protein, which is involved in chromosome translocation to produce RBM15-MKL1 fusion protein in acute megakaryocytic leukemia, regulates the alternative RNA splicing of TAL1. RBM15 promotes the production of full-length TAL1 mRNA, while reduction of RBM15 protein level via PRMT1-mediated degradation pathway favors the production of TAL1s. RBM15 directly binds to intronic regions on TAL1 pre-mRNA. Binding of RBM15 is responsible for recruiting SF3B1-associated RNA splicing complex. Given that PRMT1 senses the hypoxia status of hematopoietic cells, the changing of TAL1s/TAL1fl ratio by PRMT1 activity may be an adaptive response of hematopoietic cells to hypoxia status. The short form TAL1s still contains the helix-loop-helix DNA binding domain but not the N terminal regions upstream of the DNA binding domain. Thus, the TAL1s may act as a dominant negative mutant of the full-length TAL1fl to block TAL1fl-regulated transcription. We demonstrated that overexpression of TAL1s not the full-length TAL1promotes the erythroid differentiation of K562 cells. Although TAL1 gene is required for both erythroid and megakaryocyte differentiation at early stage of hematopoiesis, TAL1s does not promote megakaryocyte differentiation. Therefore, fine-tuning the TAL1 isoforms by the PRMT1-RBM15 axis determine the cell fate of a MEP progenitor cell. Using immunoprecipitation assays and mass spectrometry analysis, we identified proteins specifically associated with the N terminal region of TAL1. How unique TAL1s-associated transcriptional regulatory complex plays in erythroid differentiation will be discussed in the presentation in comparison with the TAL1fl-asociated protein complex. In summary, our findings stratify another new layer of regulation by PRMT1, which relays extracellular signals (such as hypoxia signal) to transcriptional regulatory program. Given that PRMT1 is often constitutively highly expressed in leukemia cells, how overproduction of short form TAL1 interferes with normal hematopoiesis may help to explain the molecular mechanisms of many hematological malignancies associated with dysregulation of TAL1 expression. Disclosures No relevant conflicts of interest to declare.
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dynamic interaction between TAL1 oncoprotein and lsd1 regulates TAL1 function in hematopoiesis and leukemogenesis
Oncogene, 2012Co-Authors: Changwang Deng, Yi Qiu, Keji Zhao, Bhavita Patel, Marjorie Brand, Suming HuangAbstract:TAL1/SCL is a hematopoietic-specific oncogene and its activity is regulated by associated transcriptional co-activators and corepressors. Dysregulation of TAL1 activity has been associated with T-cell leukemogenesis. However, it remains unclear how the interactions between TAL1 and corepressors versus co-activators are properly regulated. Here, we reported that protein kinase A (PKA)-mediated phosphorylation regulates TAL1 interaction with the lysine-specific demethylase (LSD1) that removes methyl group from methylated Lys 4 on histone H3 tails. Phosphorylation of serine 172 in TAL1 specifically destabilizes the TAL1-LSD1 interaction leading to promoter H3K4 hypermethylation and activation of target genes that have been suppressed in normal and malignant hematopoiesis. Knockdown of TAL1 or LSD1 led to a derepression of the TAL1 target genes in T-cell acute lymphoblast leukemia (T-ALL) Jurkat cells, which is accompanied by elevating promoter H3K4 methylation. Similarly, treatment of PKA activator forskolin resulted in derepression of target genes by reducing its interaction with LSD1 while PKA inhibitor H89 represses them by suppressing H3K4 methylation levels. Consistent with the dual roles of TAL1 in transcription, TAL1-associated LSD1 is decreased while recruitment of hSET1 is increased at the TAL1 targets during erythroid differentiation. This process is accompanied by a dramatic increase in H3K4 methylation. Thus, our data revealed a novel interplay between PKA phosphorylation and TAL1-mediated epigenetic regulation that regulates hematopoietic transcription and differentiation programs during hematopoiesis and leukemogenesis.
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ctcf mediated enhancer and promoter interaction regulates differential expression of TAL1 oncogene in normal and malignant hematopoiesis
Blood, 2012Co-Authors: Bhavita Patel, Yi Qiu, Keji Zhao, Changwang Deng, Michael D Litt, Mariana St Just Riberio, Kairong Cui, Yuanyuan Kang, Suming HuangAbstract:Abstract Abstract 281 The key hematopoietic transcription factor TAL1/SCL is essential for the specification of hematopoietic stem cells (HSCs) and differentiation along myeloid and erythroid lineages. However, ectopic activation of TAL1 gene is the most frequent gain-of-function mutation associated with childhood T-cell acute lymphoblastic leukemia (T-ALL) in approximately 60% of patients. To understand underlying epigenetic mechanisms of TAL1 activation in normal and in malignant hematopoiesis, we performed ChIP, ChIP-seq, and chromatin conformation capture assays to investigate chromatin structure profile which correlates with transcriptional activation, at the TAL1 locus comparing erythroid progenitors, K562 and CD36+ cells, with T-ALL cells, Jurkat and Rex cells. We found distinct epigenetic landscape and chromatin organization across the TAL1 locus in these two different types of hematopoietic cells. Although H3K4me3 is enriched at the TAL1 promoter in both erythroid progenitors and in T-ALL cells, the erythroid specific +51 enhancer is marked by active histone modifications only in erythroid progenitors, but not in T-ALL cells. Furthermore, we demonstrated that +51 enhancer interacts with the TAL1 promoter 1a via a long range chromatin loop in vivo in erythroid CD36+ and K562 cells. The recruitment of hSET1 HMT complex facilitates this enhancer/promoter chromatin interaction and depletion of hSET1 leads to loss of H3K4 methylation, enhancer/promoter interaction, RNA PolII loading, and TAL1 transcription at both the TAL1 promoter 1a and the +51 enhancer. In addition, loss of SET1 in CD34+ HSCs exhibited a significant reduction in the formation of CFU-E and BFU-E colonies. In contrast, neither H3K4 methylation nor enhancer/promoter interaction was detected at the +51 enhancer in T-ALL cells. Finally, we investigated the role of insulator protein CTCF in the regulation of TAL1 expression in normal and in malignant cells. We found that regardless of similar CTCF binding patterns at the TAL1 locus in erythroid progenitors and T-ALL cells, CTCF elements flanking the TAL1 locus mediate a long-range loop that encloses TAL1 promoter and enhancer in the same chromatin domain in erythroid progenitors, whereas in T-ALL Jurkat cells the CTCF site located between the promoter 1a and the +51 enhancer forms a repressive loop that expels the +51 enhancer from the same domain with the promoter 1a. Overall, our study demonstrates that a novel long range intrachromosomal interaction between the TAL1 promoter 1a and the +51 enhancer regulated and organized by hSET1 histone methyltransferase and insulator protein CTCF controls TAL1 promoter activity in normal versus malignant hematopoiesis. Disclosures: No relevant conflicts of interest to declare.
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a t cell specific element activates the TAL1 oncogene via an interchromosomal interaction during leukemogenesis
Blood, 2012Co-Authors: Yuanyuan Kang, Yi Qiu, Keji Zhao, Bhavita Patel, Kairong Cui, Suming HuangAbstract:Abstract 3507 T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignant disease of thymocytes that mainly affects children and has very poor prognosis with high rates of relapse. A prominent feature observed in 60% of T-ALL childhood patients is the ectopic expression of a key hematopoietic transcription factor TAL1/SCL. Although several enhancers has been identified to play an important role in normal hematopoietic differentiation, the histone modification patterns and chromatin organization over the whole TAL1 locus reveled that none of them is active in T-ALL cell lines such as Jurkat and Rex cells. It remains currently unknown how TAL1 is activated in the majority of T-ALL patients lacking the TAL1 locus rearrangements. To understand the molecular mechanism underlying regulation of the TAL1 oncogene in leukemic T-cells, we employed circularized chromosome conformation capture (4C) methodology to identify new regulatory elements that activate TAL1 specifically in T-ALL leukemia. Using the TAL1 promoter 1a as the bait, we discovered that the TAL1 promoter 1a interacts with the TIL16 element (TAL1 interacting locus in chromosome 16) that is located at ∼15 Kb downstream of T-cell specific CD2BP2 gene in T-ALL cell line Jurkat, but not in erythroid progenitor K562 cells. The CD2BP2 protein is a cellular adapter protein that was originally identified as a binding partner of the T cell adhesion protein CD2 in the context of T cell signaling. The TIL16 element contains the bind sites for several transcription factors that are important for hematopoiesis such as C-Maf, Pax5, HoxA7 and USF2. The inter-chromosomal interaction between the TIL16 and the TAL1 promoter 1a was further confirmed by chromosome conformation capture (3C) assay in three TAL1 over-expressing T-ALL cell lines, Jurkat, REX and Molt4, but not in K562 cells. Recent genome wide study has correlates H3K4 mono- or dimethyl marks with distal enhancers while trimethyl H3K4 is enriched in promoters of active genes. To further test if the TIL16 acts as T-cell specific enhancer for TAL1 activation in T-ALL cells, we carried out ChIP-seq and ChIP analysis in CD4 T cells, Jurkat, and K562 cells. We found that the TIL16 element is specifically marked by H3K4me1 in Jurkat and CD4+ T-cells but not in K562 cells. The enrichment of H3K4me1 is correlated with the binding of c-Maf, a T-cell specific transcription factor. To further test whether the TIL16 element contributes to transcription activity, a DNA fragments containing the TIL16 element were cloneed into SV40 minimal promoter driven luciferase reporter and introduced into K562 and several T-ALL cell lines. Compared to the pGL3-SV40 vector that showed only minimal luciferase activity, the 1 Kb TIL element specifically activated transcription of the luciferase reporter in T-ALL cells, but not in erythroid progenitor K562 cells suggesting that the TIL16 element functions as a T-cell specific TAL1 enhancer. Thus, our data revealed a novel epigenetic mechanism by which the TAL1 oncogene is ectopically activated in T-cell leukemia. Disclosures: No relevant conflicts of interest to declare.
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t cell acute leukemia 1 TAL1 regulation of erythropoietin receptor and association with excessive erythrocytosis
Journal of Biological Chemistry, 2012Co-Authors: Heather Rogers, Suming Huang, Keji Zhao, Kairong Cui, Li Wang, Mawadda Alnaeeli, James J Bieker, Josef T Prchal, Babette B Weksler, Constance Tom NoguchiAbstract:During erythropoiesis, erythropoietin stimulates induction of erythroid transcription factors that activate expression of erythroid genes including the erythropoietin receptor (EPO-R) that results in increased sensitivity to erythropoietin. DNA binding of the basic helix-loop-helix transcription factor, TAL1/SCL, is required for normal erythropoiesis. A link between elevated TAL1 and excessive erythrocytosis is suggested by erythroid progenitor cells from a patient that exhibits unusually high sensitivity to erythropoietin with concomitantly elevated TAL1 and EPO-R expression. We found that TAL1 regulates EPO-R expression mediated via three conserved E-box binding motifs (CAGCTG) in the EPO-R 5′ untranslated transcribed region. TAL1 increases association of the GATA-1·TAL1·LMO2·LDB1 transcription activation complex to the region that includes the transcription start site and the 5′ GATA and 3′ E-box motifs flanking the EPO-R transcription start site suggesting that TAL1 promotes accessibility of this region. Nucleosome shifting has been demonstrated to facilitate TAL1 but not GATA-1 binding to regulate target gene expression. Accordingly, we observed that with induced expression of EPO-R in hemotopoietic progenitor cells, nucleosome phasing shifts to increase the linker region containing the EPO-R transcription start site and TAL1 binds to the flanking 5′ GATA and 3′ E-box regions of the promoter. These data suggest that TAL1 binds to the EPO-R promoter to activate EPO-R expression and provides a potential link to elevated EPO-R expression leading to hypersensitivity to erythropoietin and the resultant excessive erythrocytosis.
Richard Baer - One of the best experts on this subject based on the ideXlab platform.
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does activation of the TAL1 gene occur in a majority of patients with t cell acute lymphoblastic leukemia a pediatric oncology group study
Blood, 1995Co-Authors: Robert O Bash, S Hall, Charles F Timmons, W M Crist, Michael D Amylon, R G Smith, Richard BaerAbstract:Almost 25% of patients with T-cell acute lymphoblastic leukemia (T-ALL) have tumor-specific rearrangements of the TAL1 gene. Although TAL1 expression has not been observed in normal lymphocytes, TAL1 gene products are readily detected in leukemic cells that harbor a rearranged TAL1 allele. Hence, it has been proposed that ectopic expression of TAL1 promotes the development of T-ALL. In this report, we show that TAL1 is expressed in the leukemic cells of most patients with T-ALL, including many that do not display an apparent TAL1 gene alteration. A polymorphic dinucleotide repeat in the transcribed sequences of TAL1 was used to determine the allele specificity of TAL1 transcription in primary T-ALL cells. Monoallelic expression of TAL1 was observed in the leukemic cells of all patients (8 of 8) bearing a TAL1 gene rearrangement. In the leukemic cells of patients without detectable TAL1 rearrangements, TAL1 transcription occurred in either a monoallelic (3 of 7 patients) or a biallelic (4 of 7 patients) fashion. Thus, TAL1 activation in these patients may result from subtle alterations in cis-acting regulatory sequences (affecting expression of a single TAL1 allele) or changes in trans-acting factors that control TAL1 transcription (affecting expression of both TAL1 alleles).
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positive and negative transcriptional control by the TAL1 helix loop helix protein
Proceedings of the National Academy of Sciences of the United States of America, 1994Co-Authors: Hailing Hsu, Isobel Wadman, Julia Tsou Tsan, Richard BaerAbstract:Tumor-specific activation of the TAL1 gene is the most common genetic defect associated with T-cell acute lymphoblastic leukemia. The TAL1 gene products possess a basic helix-loop-helix (bHLH) motif, a protein-dimerization and DNA-binding domain found in several transcription factors. TAL1 polypeptides interact, in vitro and in vivo, with class A bHLH proteins (e.g., E47) to form heterodimers with sequence-specific DNA-binding activity. In this study, we show that TAL1 can regulate the transcription of an artificial reporter gene that contains binding sites for bHLH heterodimers involving TAL1. Transcription of the reporter is strongly induced by E47-E47 homodimers and moderately induced by TAL1-E47 heterodimers. Thus, in a cellular environment that allows formation of E47-E47 homodimers (e.g., in the absence of Id regulatory proteins) TAL1 can repress transcription by recruiting E47 into bHLH complexes with less transcriptional activity (i.e., TAL1-E47 heterodimers). However, in other settings TAL1 can activate transcription because TAL1-E47 heterodimers are more resistant to negative regulation by Id proteins. Hence, TAL1 can potentially regulate transcription in either a positive or negative fashion.
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formation of in vivo complexes between the TAL1 and e2a polypeptides of leukemic t cells
Proceedings of the National Academy of Sciences of the United States of America, 1994Co-Authors: Hailing Hsu, Isobel Wadman, Richard BaerAbstract:Abstract Tumor-specific activation of the TAL1 gene occurs in approximately 25% of patients with T-cell acute lymphoblastic leukemia (T-ALL). The TAL1 gene products possess a basic helix-loop-helix (bHLH) domain that interacts in vitro with the bHLH proteins (E12 and E47) encoded by the E2A locus. We have now applied two independent methods, the two-hybrid procedure and co-immunoprecipitation analysis, to demonstrate that TAL1 and E2A polypeptides also associate in vivo. These studies show that the bHLH domain of TAL1 selectively interacts with the bHLH domains of E12 and E47, but not with the Id1 helix-loop-helix protein. TAL1 does not self-associate to form homodimeric complexes, implying that the in vivo functions of TAL1 depend on heterologous interaction with other bHLH proteins such as E12 and E47. Co-immunoprecipitation analysis revealed the presence of endogenous TAL1/E2A complexes in Jurkat cells, a leukemic line derived from a T-ALL patient. Thus, the malignant properties of TAL1 may be due to obligate interaction with the E2A polypeptides.
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preferred sequences for dna recognition by the TAL1 helix loop helix proteins
Molecular and Cellular Biology, 1994Co-Authors: Hailing Hsu, Julia Tsou Tsan, Lan Huang, Walter Funk, Woodring E Wright, Robert E Kingston, Richard BaerAbstract:Tumor-specific activation of the TAL1 gene is the most common genetic alteration seen in patients with T-cell acute lymphoblastic leukemia. The TAL1 gene products contain the basic helix-loop-helix (bHLH) domain, a protein dimerization and DNA-binding motif common to several known transcription factors. A binding-site selection procedure has now been used to evaluate the DNA recognition properties of TAL1. These studies demonstrate that TAL1 polypeptides do not have intrinsic DNA-binding activity, presumably because of their inability to form bHLH homodimers. However, TAL1 readily interacts with any of the known class A bHLH proteins (E12, E47, E2-2, and HEB) to form heterodimers that bind DNA in a sequence-specific manner. The TAL1 heterodimers preferentially recognize a subset of E-box elements (CANNTG) that can be represented by the consensus sequence AACAGATGGT. This consensus is composed of half-sites for recognition by the participating class A bHLH polypeptide (AACAG) and the TAL1 polypeptide (ATGGT). TAL1 heterodimers with DNA-binding activity are readily detected in nuclear extracts of Jurkat, a leukemic cell line derived from a patient with T-cell acute lymphoblastic leukemia. Hence, TAL1 is likely to bind and regulate the transcription of a unique subset of subordinate target genes, some of which may mediate the malignant function of TAL1 during T-cell leukemogenesis.
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expression of the TAL1 proto oncogene in cultured endothelial cells and blood vessels of the spleen
Oncogene, 1993Co-Authors: Larnyuan Hwang, Richard Baer, Mark Siegelman, Laurie S Davis, Nancy OppenheimermarksAbstract:The TAL1 proto-oncogene encodes a basic helix-loop-helix (bHLH) protein that has been implicated in the pathogenesis of T-cell acute lymphoblastic leukemia. Normal expression of TAL1 is observed in erythrocytic, megakaryocytic and mastocytic cells of the hematopoietic lineage. We now report that both RNA transcripts and polypeptide products of TAL1 are present in human umbilical vein endothelial cells cultured in vitro. Moreover, in situ hybridization revealed a restricted pattern of TAL1 expression in endothelial cells in vivo, including vessels within the white pulp and follicles of the spleen. In view of its presumptive role as a transcriptional factor, the TAL1 gene product may serve during normal development as a regulator of endothelial cell growth or differentiation.
Constance Tom Noguchi - One of the best experts on this subject based on the ideXlab platform.
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t cell acute leukemia 1 TAL1 regulation of erythropoietin receptor and association with excessive erythrocytosis
Journal of Biological Chemistry, 2012Co-Authors: Heather Rogers, Suming Huang, Keji Zhao, Kairong Cui, Li Wang, Mawadda Alnaeeli, James J Bieker, Josef T Prchal, Babette B Weksler, Constance Tom NoguchiAbstract:During erythropoiesis, erythropoietin stimulates induction of erythroid transcription factors that activate expression of erythroid genes including the erythropoietin receptor (EPO-R) that results in increased sensitivity to erythropoietin. DNA binding of the basic helix-loop-helix transcription factor, TAL1/SCL, is required for normal erythropoiesis. A link between elevated TAL1 and excessive erythrocytosis is suggested by erythroid progenitor cells from a patient that exhibits unusually high sensitivity to erythropoietin with concomitantly elevated TAL1 and EPO-R expression. We found that TAL1 regulates EPO-R expression mediated via three conserved E-box binding motifs (CAGCTG) in the EPO-R 5′ untranslated transcribed region. TAL1 increases association of the GATA-1·TAL1·LMO2·LDB1 transcription activation complex to the region that includes the transcription start site and the 5′ GATA and 3′ E-box motifs flanking the EPO-R transcription start site suggesting that TAL1 promotes accessibility of this region. Nucleosome shifting has been demonstrated to facilitate TAL1 but not GATA-1 binding to regulate target gene expression. Accordingly, we observed that with induced expression of EPO-R in hemotopoietic progenitor cells, nucleosome phasing shifts to increase the linker region containing the EPO-R transcription start site and TAL1 binds to the flanking 5′ GATA and 3′ E-box regions of the promoter. These data suggest that TAL1 binds to the EPO-R promoter to activate EPO-R expression and provides a potential link to elevated EPO-R expression leading to hypersensitivity to erythropoietin and the resultant excessive erythrocytosis.
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gata binding protein 4 gata 4 and t cell acute leukemia 1 TAL1 regulate myogenic differentiation and erythropoietin response via cross talk with sirtuin1 sirt1
Journal of Biological Chemistry, 2012Co-Authors: Li Wang, Suming Huang, Heather Rogers, Yi Jia, Constance Tom NoguchiAbstract:Erythropoietin (EPO), the cytokine required for erythrocyte production, contributes to muscle progenitor cell proliferation and delay myogenic differentiation. However, the underlying mechanism is not yet fully understood. Here, we report that EPO changes the skeletal myogenic regulatory factor expression program and delays differentiation via induction of GATA-4 and the basic helix-loop-helix TAL1 and that knockdown of both factors promotes differentiation. EPO increases the Sirt1 level, a NAD+-dependent deacetylase, and also induces the NAD+/NADH ratio that further increases Sirt1 activity. Sirt1 knockdown reduced GATA-4 and TAL1 expression, impaired EPO effect on delayed myogenic differentiation, and the Sirt1 knockdown effect was abrogated when combined with overexpression of GATA-4 or TAL1. GATA-4 interacts with Sirt1 and targets Sirt1 to the myogenin promoter and represses myogenin expression, whereas TAL1 inhibits myogenin expression by decreasing MyoD binding to and activation of the myogenin promoter. Sirt1 was found to bind to the GATA-4 promoter to directly regulate GATA-4 expression and GATA-4 binds to the TAL1 promoter to regulate TAL1 expression positively. These data suggest that GATA-4, TAL1, and Sirt1 cross-talk each other to regulate myogenic differentiation and mediate EPO activity during myogenic differentiation with Sirt1 playing a role upstream of GATA-4 and TAL1. Taken together, our findings reveal a novel role for GATA-4 and TAL1 to affect skeletal myogenic differentiation and EPO response via cross-talk with Sirt1.
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LSD1-mediated epigenetic modification is required for TAL1 function and hematopoiesis
Proceedings of the National Academy of Sciences of the United States of America, 2009Co-Authors: Kristell Valverde, Yi Qiu, Constance Tom Noguchi, Suming HuangAbstract:TAL1 is a critical transcription factor required for hematopoiesis. However, perturbation of its activity often leads to T cell leukemia. Whether and how its transcriptional activities are regulated during hematopoiesis remains to be addressed. Here, we show that TAL1 is associated with histone demethylase complexes containing lysine-specific demethylase 1 (LSD1), RE1 silencing transcription factor corepressor (CoREST), histone deacetylase 1 (HDAC1), and histone deacetylase 2 in erythroleukemia and T cell leukemia cells. The enzymatic domain of LSD1 plays an important role in repressing the TAL1-directed transcription of GAL4 reporter linked to a thymidine kniase minimal promoter. Furthermore, we demonstrate that the TAL1-associated LSD1, HDAC1, and their enzymatic activities are coordinately down-regulated during the early phases of erythroid differentiation. Consistent with the rapid changes of TAL1-corepressor complex during differentiation, TAL1 recruits LSD1 to the silenced p4.2 promoter in undifferentiated, but not in differentiated, murine erythroleukemia (MEL) cells. Finally, shRNA-mediated knockdown of LSD1 in MEL cells resulted in derepression of the TAL1 target gene accompanied by increasing dimeH3K4 at the promoter region. Thus, our data revealed that histone lysine demethylase LSD1 may negatively regulate TAL1-mediated transcription and suggest that the dynamic regulation of TAL1-associated LSD1/HDAC1 complex may determine the onset of erythroid differentiation programs.
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erythropoietin receptor expression a role for the TAL1 scl and gata 1 complex
Blood, 2008Co-Authors: Heather Rogers, Li Wang, Constance Tom NoguchiAbstract:TAL1/SCL is required for hematopoietic stem cell development and is essential for normal erythropoiesis. Although TAL1-DNA binding is not required for hematopoietic development, TAL1 participates in a complex with other binding partners GATA-1, LMO2, Ldb1 and possibly Sp1. During erythropoiesis, TAL1 can activate expression via direct association with E-box-GATA DNA sequence motifs for lineage specific genes such as Protein 4.2. In mice, loss of TAL1 in adult erythropoiesis also affects TER119 expression and BFU-E growth resulting in anemia. We found that knock down of TAL1 expression in primary erythropoietin (EPO) stimulated hematopoietic progenitor cells in culture inhibited EPO receptor (EPOR) expression and erythroid differentiation, consistent with the anemia observed in mice with targeted deletion of adult TAL1 expression. In contrast, overexpression of TAL1 in erythroid cells increased both EPOR expression and erythroid differentiation. In fact, overexpression of EPOR was sufficient to increase differentiation in erythroid progenitor cell cultures. Increased EPOR expression by TAL1 was mediated by 3 conserved E-boxes in the 5′ UTR. In reporter gene assays in K562 erythroid cells, EPOR promoter activity was lost with the mutation of these E-boxes, which are located 75 bp downstream of the required GATA-1 binding motif in the human EPOR proximal promoter. TAL1/E2A dimer binding to the E-box region was demonstrated by gel mobility shift assay. TAL1 transactivated EPOR mRNA and chromatin immunoprecipitation (ChIP) analysis confirmed that TAL1 bound directly to the E-box region in intact erythroid cells. We previously showed that GATA-1 also transactivates EPOR gene expression and is required for high level of EPOR transcription activity. In ChIP assays, an antibody to TAL1 also pulled down chromatin containing the GATA-1 binding site as well as the E-box region and conversely, antibodies to GATA-1 pulled down chromatin corresponding to the GATA-1 binding site and the E-box region. These data show that the complex containing TAL1 and GATA-1 bound to the E-box region and to the GATA-1 site in the EPOR proximal promoter and that the complex has greater occupancy at the GATA-1 site in the proximal promoter. High-level TAL1 expression increased chromatin containing TAL1 or GATA-1 associated with the GATA-1 binding site as well as the down stream E-box region, with a greater proportionate increase in the E-box region. Hence, in addition to the GATA-1 binding site, the downstream E-box region is necessary for high-level EPOR expression. These data suggest that transactivation of the EPOR promoter via these sites is mediated by a TAL1 and GATA-1 containing complex and that during erythropoiesis EPO induction of both TAL1 and GATA-1 regulates the high expression of its own receptor. We hypothesize that the high binding of GATA-1 and TAL1 at the GATA site opens the upstream chromatin structure. The over-expression of TAL1 promotes the open status and facilitates the spreading of open chromatin along the chromatin to the downstream E-box region. Subsequent E-box binding by TAL1 and its interaction with GATA-1 maintains the open chromatin and activates transcription.
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excessive isolated erythrocytosis linked to induction of scl TAL1 and erythropoietin receptor expression
Blood, 2007Co-Authors: Heather Rogers, Josef T Prchal, Babette B Weksler, Constance Tom NoguchiAbstract:The JAK2 V617F mutation has been detected in more than 90% polycythemia vera patients and is associated with increased erythropoietin (EPO) sensitivity or EPO independence of erythroid progenitor cells during early erythropoiesis. To determine if other molecular lesions give rise to increased erythrocytosis and realizing that the JAK2 V617F mutation may not be the initiating event in polycythemia vera, we identified an individual with isolated, excessive erythrocytosis who showed erythroid precursor hypersensitivity to EPO; no mutations on sequencing genes for erythropoietin receptor (EPO-R), VHL, or HIF-1 IRE; normal serum EPO level; normal hemoglobin function and p50; normal cardiac, hepatic and pulmonary function; no JAK2 V617F mutation and no EPO independent BFU-E. While it was not certain that this individual’s erythrocytosis was lifelong, other relatives had increased hemoglobin levels. The cause of this individual’s increased erythropoiesis was investigated using EPO-stimulated cultures of hematopoietic progenitor cells isolated from peripheral blood. As with normal controls, cell numbers were not increased after 12 days of incubation with low levels of EPO ( JAK2 V617F positive erythroid progenitor cells. Analyses of transcription factor expression revealed that induction of GATA-1 and down regulation of GATA-2 were similar to control. However, SCL/TAL1 and EKLF markedly increased beginning at day 8 to 10 and continued to rise during late erythropoiesis in contrast to control and JAK2 V617F positive cultures in which SCL/TAL1 and EKLF peaked at day 10 and decreased with late erythroid differentiation. Since increased beta-globin expression is concomitant with induction of EKLF, the rise in EKLF may account for the marked increase in benzidine positive cells from our subject. Moreover, expression of EPO-R followed the continuing rise in SCL/TAL1, increasing by 3.5 fold at day 12 compared with control cultures. This high EPO-R expression is consistent with the dramatic increase in cell proliferation during late erythropoiesis. Using forced expression of SCL/TAL1, reporter gene assay and gel mobility shift analysis in erythroid cells, we determined that SCL/TAL1 is able to bind to E-boxes located 3′ of the EPO-R proximal promoter and to activate transcription. Although EPO stimulation of erythroid progenitor cells activates EPO-R, we found that forced expression of EPO-R in primary erythroid progenitor cells in the presence of EPO increases expression of GATA-1 as well as SCL/TAL1 and EKLF, providing further evidence that specific increase in SCL/TAL1 but not GATA-1 must precede induction of EPO-R in this case of excessive erythrocytosis. These data demonstrate a link between high level induction of SCL/TAL1 expression and increased erythrocytosis and suggest a mechanism that contributes to increased erythropoietin sensitivity during late erythropoiesis.
Terence H. Rabbitts - One of the best experts on this subject based on the ideXlab platform.
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lmo2 and scl TAL1 convert non axial mesoderm into haemangioblasts which differentiate into endothelial cells in the absence of gata1
Development, 2003Co-Authors: Martin Gering, Terence H. Rabbitts, Yoshihiro Yamada, Roger PatientAbstract:The LIM domain protein Lmo2 and the basic helix-loop-helix transcription factor Scl/TAL1 are expressed in early haematopoietic and endothelial progenitors and interact with each other in haematopoietic cells. While loss-of-function studies have shown that Lmo2 and Scl/TAL1 are essential for haematopoiesis and angiogenic remodelling of the vasculature, gain-of-function studies have suggested an earlier role for Scl/TAL1 in the specification of haemangioblasts, putative bipotential precursors of blood and endothelium. In zebrafish embryos, Scl/TAL1 can induce these progenitors from early mesoderm mainly at the expense of the somitic paraxial mesoderm. We show that this restriction to the somitic paraxial mesoderm correlates well with the ability of Scl/TAL1 to induce ectopic expression of its interaction partner Lmo2. Co-injection of lmo2 mRNA with scl/TAL1 dramatically extends its effect to head, heart, pronephros and pronephric duct mesoderm inducing early blood and endothelial genes all along the anteroposterior axis. Erythroid development, however, is expanded only into pronephric mesoderm, remaining excluded from head, heart and somitic paraxial mesoderm territories. This restriction correlates well with activation of gata1 transcription and co-injection of gata1 mRNA along with scl/TAL1 and lmo2 induces erythropoiesis more broadly without ventralising or posteriorising the embryo. While no ectopic myeloid development from the Scl/TAL1-Lmo2-induced haemangioblasts was observed, a dramatic increase in the number of endothelial cells was found. These results suggest that, in the absence of inducers of erythroid or myeloid haematopoiesis, Scl/TAL1-Lmo2-induced haemangioblasts differentiate into endothelial cells.
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protein dimerization between lmo2 rbtn2 and TAL1 alters thymocyte development and potentiates t cell tumorigenesis in transgenic mice
The EMBO Journal, 1996Co-Authors: R C Larson, R Baer, Alan J. Warren, Isobel Wadman, Isabelle Lavenir, T A Larson, K Nottage, Terence H. RabbittsAbstract:Abstract The LMO2 and TAL1 genes were first identified via chromosomal translocations and later found to encode proteins that interact during normal erythroid development. Some T cell leukaemia patients have chromosomal abnormalities involving both genes, implying that LMO2 and TAL1 act synergistically to promote tumorigenesis after their inappropriate co-expression. To test this hypothesis, transgenic mice were made which co-express Lmo2 and TAL1 genes in T cells. Dimers of Lmo2 and TAL1 proteins were formed in thymocytes of double but not single transgenic mice. Furthermore, thymuses of double transgenic mice were almost completely populated by immature T cells from birth, and these mice develop T cell tumours approximately 3 months earlier than those with only the Lmo2 transgene. Thus interaction between these two proteins can alter T cell development and potentiate tumorigenesis. The data also provide formal proof that TAL1 is an oncogene, apparently acting as a tumour promoter in this system.
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association of erythroid transcription factors complexes involving the lim protein rbtn2 and the zinc finger protein gata1
Proceedings of the National Academy of Sciences of the United States of America, 1995Co-Authors: Hirotaka Osada, Hakan Axelson, Gerald Grutz, A Forster, Terence H. RabbittsAbstract:The RBTN2 LIM-domain protein, originally identified as an oncogenic protein in human T-cell leukemia, is essential for erythropoiesis. A possible role for RBTN2 in transcription during erythropoiesis has been investigated. Direct interaction of the RBTN2 protein was observed in vivo and in vitro with the GATA1 or -2 zinc-finger transcription factors, as well as with the basic helix-loop-helix protein TAL1. By using mammalian two-hybrid analysis, complexes involving RBTN2, TAL1, and GATA1, together with E47, the basic helix-loop-helix heterodimerization partner of TAL1, could be demonstrated. Thus, a molecular link exists between three proteins crucial for erythropoiesis, and the data suggest that variations in amounts of complexes involving RBTN2, TAL1, and GATA1 could be important for erythroid differentiation.
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The LIM protein RBTN2 and the basic helix-loop-helix protein TAL1 are present in a complex in erythroid cells
Proceedings of the National Academy of Sciences of the United States of America, 1994Co-Authors: V. Valge-archer, R Baer, Hirotaka Osada, Alan J. Warren, Alan Forster, Terence H. RabbittsAbstract:Abstract Chromosomal translocations in T-cell acute leukemias can activate genes encoding putative transcription factors such as the LIM proteins RBTN1 and RBTN2 and the DNA-binding basic helix-loop-helix transcription factor TAL1 associated with T-cell acute lymphocytic leukemia. While not expressed in normal T cells, RBTN2 and TAL1 are coexpressed in erythroid cells and are both important for erythroid differentiation. We demonstrate, using anti-RBTN2 and anti-TAL1 antisera, that the LIM protein RBTN2 is not phosphorylated and is complexed with the TAL1 phosphoprotein in the nucleus of erythroid cells. A complex containing both RBTN1 and TAL1 also occurs in a T-cell acute leukemia cell line. Since both RBTN2 and TAL1 are crucial for normal erythropoiesis, these data have important implications for transcription networks therein. Further, since both proteins can be involved in leukemogenesis, these data provide a direct link between proteins activated by chromosomal translocations in T-cell acute leukemia.
Yi Qiu - One of the best experts on this subject based on the ideXlab platform.
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aberrant TAL1 activation is mediated by an interchromosomal interaction in human t cell acute lymphoblastic leukemia
Leukemia, 2014Co-Authors: Bhavita Patel, Yi Qiu, Changwang Deng, Michael D Litt, Kairong Cui, Yuanyuan Kang, M S J Riberio, Tal Salz, S Casada, Keji ZhaoAbstract:Long-range chromatin interactions control metazoan gene transcription. However, the involvement of intra- and interchromosomal interactions in development and oncogenesis remains unclear. TAL1/SCL is a critical transcription factor required for the development of all hematopoietic lineages; yet, aberrant TAL1 transcription often occurs in T-cell acute lymphoblastic leukemia (T-ALL). Here, we report that oncogenic TAL1 expression is regulated by different intra- and interchromosomal loops in normal hematopoietic and leukemic cells, respectively. These intra- and interchromosomal loops alter the cell-type-specific enhancers that interact with the TAL1 promoter. We show that human SET1 (hSET1)-mediated H3K4 methylations promote a long-range chromatin loop, which brings the +51 enhancer in close proximity to TAL1 promoter 1 in erythroid cells. The CCCTC-binding factor (CTCF) facilitates this long-range enhancer/promoter interaction of the TAL1 locus in erythroid cells while blocking the same enhancer/promoter interaction of the TAL1 locus in human T-cell leukemia. In human T-ALL, a T-cell-specific transcription factor c-Maf-mediated interchromosomal interaction brings the TAL1 promoter into close proximity with a T-cell-specific regulatory element located on chromosome 16, activating aberrant TAL1 oncogene expression. Thus, our study reveals a novel molecular mechanism involving changes in three-dimensional chromatin interactions that activate the TAL1 oncogene in human T-cell leukemia.
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dynamic interaction between TAL1 oncoprotein and lsd1 regulates TAL1 function in hematopoiesis and leukemogenesis
Oncogene, 2012Co-Authors: Changwang Deng, Yi Qiu, Keji Zhao, Bhavita Patel, Marjorie Brand, Suming HuangAbstract:TAL1/SCL is a hematopoietic-specific oncogene and its activity is regulated by associated transcriptional co-activators and corepressors. Dysregulation of TAL1 activity has been associated with T-cell leukemogenesis. However, it remains unclear how the interactions between TAL1 and corepressors versus co-activators are properly regulated. Here, we reported that protein kinase A (PKA)-mediated phosphorylation regulates TAL1 interaction with the lysine-specific demethylase (LSD1) that removes methyl group from methylated Lys 4 on histone H3 tails. Phosphorylation of serine 172 in TAL1 specifically destabilizes the TAL1-LSD1 interaction leading to promoter H3K4 hypermethylation and activation of target genes that have been suppressed in normal and malignant hematopoiesis. Knockdown of TAL1 or LSD1 led to a derepression of the TAL1 target genes in T-cell acute lymphoblast leukemia (T-ALL) Jurkat cells, which is accompanied by elevating promoter H3K4 methylation. Similarly, treatment of PKA activator forskolin resulted in derepression of target genes by reducing its interaction with LSD1 while PKA inhibitor H89 represses them by suppressing H3K4 methylation levels. Consistent with the dual roles of TAL1 in transcription, TAL1-associated LSD1 is decreased while recruitment of hSET1 is increased at the TAL1 targets during erythroid differentiation. This process is accompanied by a dramatic increase in H3K4 methylation. Thus, our data revealed a novel interplay between PKA phosphorylation and TAL1-mediated epigenetic regulation that regulates hematopoietic transcription and differentiation programs during hematopoiesis and leukemogenesis.
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ctcf mediated enhancer and promoter interaction regulates differential expression of TAL1 oncogene in normal and malignant hematopoiesis
Blood, 2012Co-Authors: Bhavita Patel, Yi Qiu, Keji Zhao, Changwang Deng, Michael D Litt, Mariana St Just Riberio, Kairong Cui, Yuanyuan Kang, Suming HuangAbstract:Abstract Abstract 281 The key hematopoietic transcription factor TAL1/SCL is essential for the specification of hematopoietic stem cells (HSCs) and differentiation along myeloid and erythroid lineages. However, ectopic activation of TAL1 gene is the most frequent gain-of-function mutation associated with childhood T-cell acute lymphoblastic leukemia (T-ALL) in approximately 60% of patients. To understand underlying epigenetic mechanisms of TAL1 activation in normal and in malignant hematopoiesis, we performed ChIP, ChIP-seq, and chromatin conformation capture assays to investigate chromatin structure profile which correlates with transcriptional activation, at the TAL1 locus comparing erythroid progenitors, K562 and CD36+ cells, with T-ALL cells, Jurkat and Rex cells. We found distinct epigenetic landscape and chromatin organization across the TAL1 locus in these two different types of hematopoietic cells. Although H3K4me3 is enriched at the TAL1 promoter in both erythroid progenitors and in T-ALL cells, the erythroid specific +51 enhancer is marked by active histone modifications only in erythroid progenitors, but not in T-ALL cells. Furthermore, we demonstrated that +51 enhancer interacts with the TAL1 promoter 1a via a long range chromatin loop in vivo in erythroid CD36+ and K562 cells. The recruitment of hSET1 HMT complex facilitates this enhancer/promoter chromatin interaction and depletion of hSET1 leads to loss of H3K4 methylation, enhancer/promoter interaction, RNA PolII loading, and TAL1 transcription at both the TAL1 promoter 1a and the +51 enhancer. In addition, loss of SET1 in CD34+ HSCs exhibited a significant reduction in the formation of CFU-E and BFU-E colonies. In contrast, neither H3K4 methylation nor enhancer/promoter interaction was detected at the +51 enhancer in T-ALL cells. Finally, we investigated the role of insulator protein CTCF in the regulation of TAL1 expression in normal and in malignant cells. We found that regardless of similar CTCF binding patterns at the TAL1 locus in erythroid progenitors and T-ALL cells, CTCF elements flanking the TAL1 locus mediate a long-range loop that encloses TAL1 promoter and enhancer in the same chromatin domain in erythroid progenitors, whereas in T-ALL Jurkat cells the CTCF site located between the promoter 1a and the +51 enhancer forms a repressive loop that expels the +51 enhancer from the same domain with the promoter 1a. Overall, our study demonstrates that a novel long range intrachromosomal interaction between the TAL1 promoter 1a and the +51 enhancer regulated and organized by hSET1 histone methyltransferase and insulator protein CTCF controls TAL1 promoter activity in normal versus malignant hematopoiesis. Disclosures: No relevant conflicts of interest to declare.
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a t cell specific element activates the TAL1 oncogene via an interchromosomal interaction during leukemogenesis
Blood, 2012Co-Authors: Yuanyuan Kang, Yi Qiu, Keji Zhao, Bhavita Patel, Kairong Cui, Suming HuangAbstract:Abstract 3507 T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignant disease of thymocytes that mainly affects children and has very poor prognosis with high rates of relapse. A prominent feature observed in 60% of T-ALL childhood patients is the ectopic expression of a key hematopoietic transcription factor TAL1/SCL. Although several enhancers has been identified to play an important role in normal hematopoietic differentiation, the histone modification patterns and chromatin organization over the whole TAL1 locus reveled that none of them is active in T-ALL cell lines such as Jurkat and Rex cells. It remains currently unknown how TAL1 is activated in the majority of T-ALL patients lacking the TAL1 locus rearrangements. To understand the molecular mechanism underlying regulation of the TAL1 oncogene in leukemic T-cells, we employed circularized chromosome conformation capture (4C) methodology to identify new regulatory elements that activate TAL1 specifically in T-ALL leukemia. Using the TAL1 promoter 1a as the bait, we discovered that the TAL1 promoter 1a interacts with the TIL16 element (TAL1 interacting locus in chromosome 16) that is located at ∼15 Kb downstream of T-cell specific CD2BP2 gene in T-ALL cell line Jurkat, but not in erythroid progenitor K562 cells. The CD2BP2 protein is a cellular adapter protein that was originally identified as a binding partner of the T cell adhesion protein CD2 in the context of T cell signaling. The TIL16 element contains the bind sites for several transcription factors that are important for hematopoiesis such as C-Maf, Pax5, HoxA7 and USF2. The inter-chromosomal interaction between the TIL16 and the TAL1 promoter 1a was further confirmed by chromosome conformation capture (3C) assay in three TAL1 over-expressing T-ALL cell lines, Jurkat, REX and Molt4, but not in K562 cells. Recent genome wide study has correlates H3K4 mono- or dimethyl marks with distal enhancers while trimethyl H3K4 is enriched in promoters of active genes. To further test if the TIL16 acts as T-cell specific enhancer for TAL1 activation in T-ALL cells, we carried out ChIP-seq and ChIP analysis in CD4 T cells, Jurkat, and K562 cells. We found that the TIL16 element is specifically marked by H3K4me1 in Jurkat and CD4+ T-cells but not in K562 cells. The enrichment of H3K4me1 is correlated with the binding of c-Maf, a T-cell specific transcription factor. To further test whether the TIL16 element contributes to transcription activity, a DNA fragments containing the TIL16 element were cloneed into SV40 minimal promoter driven luciferase reporter and introduced into K562 and several T-ALL cell lines. Compared to the pGL3-SV40 vector that showed only minimal luciferase activity, the 1 Kb TIL element specifically activated transcription of the luciferase reporter in T-ALL cells, but not in erythroid progenitor K562 cells suggesting that the TIL16 element functions as a T-cell specific TAL1 enhancer. Thus, our data revealed a novel epigenetic mechanism by which the TAL1 oncogene is ectopically activated in T-cell leukemia. Disclosures: No relevant conflicts of interest to declare.
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Serine Phosphorylation On TAL1 Regulates Its Interaction with Histone Demethylase LSD1.
Blood, 2009Co-Authors: River Ybarra, Yi Qiu, Keji Zhao, Suming HuangAbstract:Abstract 1460 Poster Board I-483 TAL1, originally identified by virtue of its involvement in a T-cell acute lymphoblastic leukemia (T-ALL)-specific chromosomal translocation, is a member of the basic helix-loop-helix (bHLH) family of transcription factors and is required for the development of all hematopoietic cell lineages. TAL1 is a phosphorylated protein and its activities are mediated by the corepressors and coactivators that associate with TAL1. However, the functional link between phosphorylation and the recruitment of co-regulators by TAL1 is currently unknown. We undertook the biochemical purification of the TAL1 containing complexes and showed that TAL1 is associated with histone demethylase complexes containing LSD1, CoREST, HDAC1 and HDAC2. This complex mediates TAL1 directed transcriptional repression during hematopoiesis. The interaction between TAL1 and LSD1 are dynamically regulated and is required for TAL1's function in erythroid differentiation (Proc. Natl. Acad. Sci. USA 106: 10141-10146). To further understand the molecular mechanism that regulates the TAL1 and LSD1 interaction during hematopoiesis, we determined whether TAL1 directly interacts with LSD1 and characterized the domains required for this interaction. TAL1 and its various deletion mutants were tested for their ability to interact with LSD1 in vitro . TAL1 directly interacts with LSD1, and the interacting domain encompasses amino acids 142-185 proximal to the bHLH domain, which contains a serine 172 residue that becomes phosphorylated during transcriptional activation of TAL1. We further mutated serine 172 of TAL1 to Alanine (Ala) or to Aspartic acid (Asp) to mimic unphosphorylated or phosphorylated TAL1, respectively. While the TAL1Ser172Ala mutant remains the interaction with LSD1, TAL1Ser172Asp specifically loses its interaction with LSD1 indicating that serine 172 phosphorylation of TAL1 destabilizes the TAL1 and LSD1 interaction. Given that our previous results indicated that LSD1 inhibits TAL1 mediated erythroid differentiation, to further test whether the activity of TAL1 is mediated through interaction with LSD1, we expressed the TAL1 mutant that deleted the LSD1 interacting domain in erythroid progenitor cells and showed that the deletion of the LSD1 interacting domain of TAL1 lead to a promotion of erythroid differentiation and inhibition of proliferation. Furthermore, consistent with the rapid decline of TAL1-associated LSD1 complex during differentiation, the ChIP and ChIP-seq data showed that H3K4 di- and tri-methylation are enriched at the promoters of TAL1 target genes upon erythroid differentiation. Thus, our data revealed that histone lysine demethylase LSD1 may negatively regulate TAL1-mediated transcription and erythroid differentiation. The results suggest that the dynamic regulation of TAL1-associated LSD1/HDAC1 complex may determine the onset of erythroid differentiation programs. * These authors contribute equally to this work. Disclosures No relevant conflicts of interest to declare.
