Publications by authors named "Myunggon Ko"

26 Publications

  • Page 1 of 1

Decoding ribosomes to keep growing (in AML).

Blood 2020 06;135(23):2015-2016

Ulsan National Institute of Science and Technology.

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http://dx.doi.org/10.1182/blood.2020006021DOI Listing
June 2020

Decreased vitamin C uptake mediated by SLC2A3 promotes leukaemia progression and impedes TET2 restoration.

Br J Cancer 2020 05 16;122(10):1445-1452. Epub 2020 Mar 16.

Cancer Research Institute, Seoul National University College of Medicine, Seoul, Republic of Korea.

Background: Vitamin C suppresses leukaemogenesis by modulating Tet methylcytosine dioxygenase (TET) activity. However, its beneficial effect in the treatment of patients with acute myeloid leukaemia (AML) remains controversial. In this study, we aimed to identify a potential predictive biomarker for vitamin C treatment in AML.

Methods: Gene expression patterns and their relevance to the survival of AML patients were analysed with The Cancer Genome Atlas (TCGA) and Therapeutically Applicable Research to Generate Effective Treatments (TARGET) database cases. In vitro experiments were performed on AML cell lines, a SLC2A3-knockdown cell line and patient-derived primary AML cells.

Results: SLC2A3 expression was significantly decreased in leukaemic blast cells. Below-median SLC2A3 expression was associated with poor overall survival. Low SLC2A3 expression was associated with less effective demethylation, and a diminished vitamin C effect in the AML and lymphoma cell lines. SLC2A3 knockdown in the KG-1 cell line decreased the response of vitamin C. In patient-derived primary AML cells, vitamin C only restored TET2 activity when SLC2A3 was expressed.

Conclusion: SLC2A3 could be used as a potential biomarker to predict the effect of vitamin C treatment in AML.
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http://dx.doi.org/10.1038/s41416-020-0788-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7217885PMC
May 2020

TET family dioxygenases and DNA demethylation in stem cells and cancers.

Exp Mol Med 2017 04 28;49(4):e323. Epub 2017 Apr 28.

Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan, Korea.

The methylation of cytosine and subsequent oxidation constitutes a fundamental epigenetic modification in mammalian genomes, and its abnormalities are intimately coupled to various pathogenic processes including cancer development. Enzymes of the Ten-eleven translocation (TET) family catalyze the stepwise oxidation of 5-methylcytosine in DNA to 5-hydroxymethylcytosine and further oxidation products. These oxidized 5-methylcytosine derivatives represent intermediates in the reversal of cytosine methylation, and also serve as stable epigenetic modifications that exert distinctive regulatory roles. It is becoming increasingly obvious that TET proteins and their catalytic products are key regulators of embryonic development, stem cell functions and lineage specification. Over the past several years, the function of TET proteins as a barrier between normal and malignant states has been extensively investigated. Dysregulation of TET protein expression or function is commonly observed in a wide range of cancers. Notably, TET loss-of-function is causally related to the onset and progression of hematologic malignancy in vivo. In this review, we focus on recent advances in the mechanistic understanding of DNA methylation-demethylation dynamics, and their potential regulatory functions in cellular differentiation and oncogenic transformation.
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http://dx.doi.org/10.1038/emm.2017.5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6130217PMC
April 2017

DNA Demethylation of the Foxp3 Enhancer Is Maintained through Modulation of Ten-Eleven-Translocation and DNA Methyltransferases.

Mol Cells 2016 Dec 13;39(12):888-897. Epub 2016 Dec 13.

Department of Pathology, Hallym University College of Medicine, Chuncheon 24252, Korea.

Stable expression of Foxp3 is ensured by demethylation of CpG motifs in the intronic element, the conserved non-coding sequence 2 (CNS2), which persists throughout the lifespan of regulatory T cells (Tregs). However, little is known about the mechanisms on how CNS2 demethylation is sustained. In this study, we found that Ten-Eleven-Translocation (Tet) DNA dioxygenase protects the CpG motifs of CNS2 from re-methylation by DNA methyltransferases (Dnmts) and prevents Tregs from losing Foxp3 expression under inflammatory conditions. Upon stimulation of Tregs by interleukin-6 (IL6), Dnmt1 was recruited to CNS2 and induced methylation, which was inhibited by Tet2 recruited by IL2. Tet2 prevented CNS2 re-methylation by not only the occupancy of the CNS2 locus but also by its enzymatic activity. These results show that the CNS2 methylation status is dynamically regulated by a balance between Tets and Dnmts which influences the expression of Foxp3 in Tregs.
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http://dx.doi.org/10.14348/molcells.2016.0276DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5223106PMC
December 2016

DNMT3A and TET2 compete and cooperate to repress lineage-specific transcription factors in hematopoietic stem cells.

Nat Genet 2016 09 18;48(9):1014-23. Epub 2016 Jul 18.

Stem Cells and Regenerative Medicine Center, Baylor College of Medicine, Houston, Texas, USA.

Mutations in the epigenetic modifiers DNMT3A and TET2 non-randomly co-occur in lymphoma and leukemia despite their epistasis in the methylation-hydroxymethylation pathway. Using Dnmt3a and Tet2 double-knockout mice in which the development of malignancy is accelerated, we show that the double-knockout methylome reflects regions of independent, competitive and cooperative activity. Expression of lineage-specific transcription factors, including the erythroid regulators Klf1 and Epor, is upregulated in double-knockout hematopoietic stem cells (HSCs). DNMT3A and TET2 both repress Klf1, suggesting a model of cooperative inhibition by epigenetic modifiers. These data demonstrate a dual role for TET2 in promoting and inhibiting HSC differentiation, the loss of which, along with DNMT3A, obstructs differentiation, leading to transformation.
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http://dx.doi.org/10.1038/ng.3610DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4957136PMC
September 2016

TET2 Regulates Mast Cell Differentiation and Proliferation through Catalytic and Non-catalytic Activities.

Cell Rep 2016 05 5;15(7):1566-1579. Epub 2016 May 5.

Institute for Research in Biomedicine, Universita' della Svizzera italiana (USI), 6500 Bellinzona, Switzerland. Electronic address:

Dioxygenases of the TET family impact genome functions by converting 5-methylcytosine (5mC) in DNA to 5-hydroxymethylcytosine (5hmC). Here, we identified TET2 as a crucial regulator of mast cell differentiation and proliferation. In the absence of TET2, mast cells showed disrupted gene expression and altered genome-wide 5hmC deposition, especially at enhancers and in the proximity of downregulated genes. Impaired differentiation of Tet2-ablated cells could be relieved or further exacerbated by modulating the activity of other TET family members, and mechanistically it could be linked to the dysregulated expression of C/EBP family transcription factors. Conversely, the marked increase in proliferation induced by the loss of TET2 could be rescued exclusively by re-expression of wild-type or catalytically inactive TET2. Our data indicate that, in the absence of TET2, mast cell differentiation is under the control of compensatory mechanisms mediated by other TET family members, while proliferation is strictly dependent on TET2 expression.
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http://dx.doi.org/10.1016/j.celrep.2016.04.044DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5584687PMC
May 2016

Acute loss of TET function results in aggressive myeloid cancer in mice.

Nat Commun 2015 Nov 26;6:10071. Epub 2015 Nov 26.

Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, California 92037, USA.

TET-family dioxygenases oxidize 5-methylcytosine (5mC) in DNA, and exert tumour suppressor activity in many types of cancers. Even in the absence of TET coding region mutations, TET loss-of-function is strongly associated with cancer. Here we show that acute elimination of TET function induces the rapid development of an aggressive, fully-penetrant and cell-autonomous myeloid leukaemia in mice, pointing to a causative role for TET loss-of-function in this myeloid malignancy. Phenotypic and transcriptional profiling shows aberrant differentiation of haematopoietic stem/progenitor cells, impaired erythroid and lymphoid differentiation and strong skewing to the myeloid lineage, with only a mild relation to changes in DNA modification. We also observe progressive accumulation of phospho-H2AX and strong impairment of DNA damage repair pathways, suggesting a key role for TET proteins in maintaining genome integrity.
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http://dx.doi.org/10.1038/ncomms10071DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4674670PMC
November 2015

DNA methylation and hydroxymethylation in hematologic differentiation and transformation.

Curr Opin Cell Biol 2015 Dec 18;37:91-101. Epub 2015 Nov 18.

Division of Signaling and Gene Expression, La Jolla Institute for Allergy & Immunology, La Jolla, CA 92037, USA; Department of Pharmacology and Moores Cancer Center, University of California at San Diego, La Jolla, CA, USA; Sanford Consortium for Regenerative Medicine, La Jolla, CA 92037, USA.

Maintenance of the balance of DNA methylation and demethylation is fundamental for normal cellular development and function. Members of the Ten-Eleven-Translocation (TET) family proteins are Fe(II)-dependent and 2-oxoglutarate-dependent dioxygenases that catalyze sequential oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) and subsequent oxidized derivatives in DNA. In addition to their roles as intermediates in DNA demethylation, these oxidized methylcytosines are novel epigenetic modifications of DNA. DNA methylation and hydroxymethylation profiles are markedly disrupted in a wide range of cancers but how these changes are related to the pathogenesis of cancers is still ambiguous. In this review, we discuss the current understanding of TET protein functions in normal and malignant hematopoietic development and the ongoing questions to be resolved.
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http://dx.doi.org/10.1016/j.ceb.2015.10.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4688184PMC
December 2015

Functions of TET Proteins in Hematopoietic Transformation.

Mol Cells 2015 Nov 10;38(11):925-35. Epub 2015 Nov 10.

School of Life Sciences, Ulsan National Institute of Science and Technology, Korea.

DNA methylation is a well-characterized epigenetic modification that plays central roles in mammalian development, genomic imprinting, X-chromosome inactivation and silencing of retrotransposon elements. Aberrant DNA methylation pattern is a characteristic feature of cancers and associated with abnormal expression of oncogenes, tumor suppressor genes or repair genes. Ten-eleven-translocation (TET) proteins are recently characterized dioxygenases that catalyze progressive oxidation of 5-methylcytosine to produce 5-hydroxymethylcytosine and further oxidized derivatives. These oxidized methylcytosines not only potentiate DNA demethylation but also behave as independent epigenetic modifications per se. The expression or activity of TET proteins and DNA hydroxymethylation are highly dysregulated in a wide range of cancers including hematologic and non-hematologic malignancies, and accumulating evidence points TET proteins as a novel tumor suppressor in cancers. Here we review DNA demethylation-dependent and -independent functions of TET proteins. We also describe diverse TET loss-of-function mutations that are recurrently found in myeloid and lymphoid malignancies and their potential roles in hematopoietic transformation. We discuss consequences of the deficiency of individual Tet genes and potential compensation between different Tet members in mice. Possible mechanisms underlying facilitated oncogenic transformation of TET-deficient hematopoietic cells are also described. Lastly, we address non-mutational mechanisms that lead to suppression or inactivation of TET proteins in cancers. Strategies to restore normal 5mC oxidation status in cancers by targeting TET proteins may provide new avenues to expedite the development of promising anti-cancer agents.
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http://dx.doi.org/10.14348/molcells.2015.0294DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4673406PMC
November 2015

A zebrafish model of myelodysplastic syndrome produced through tet2 genomic editing.

Mol Cell Biol 2015 Mar 15;35(5):789-804. Epub 2014 Dec 15.

Department of Pediatric Oncology, Dana-Farber Cancer Institute and Harvard Medical School, Boston, Massachusetts, USA

The ten-eleven translocation 2 gene (TET2) encodes a member of the TET family of DNA methylcytosine oxidases that converts 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) to initiate the demethylation of DNA within genomic CpG islands. Somatic loss-of-function mutations of TET2 are frequently observed in human myelodysplastic syndrome (MDS), which is a clonal malignancy characterized by dysplastic changes of developing blood cell progenitors, leading to ineffective hematopoiesis. We used genome-editing technology to disrupt the zebrafish Tet2 catalytic domain. tet2(m/m) (homozygous for the mutation) zebrafish exhibited normal embryonic and larval hematopoiesis but developed progressive clonal myelodysplasia as they aged, culminating in myelodysplastic syndromes (MDS) at 24 months of age, with dysplasia of myeloid progenitor cells and anemia with abnormal circulating erythrocytes. The resultant tet2(m/m) mutant zebrafish lines show decreased levels of 5hmC in hematopoietic cells of the kidney marrow but not in other cell types, most likely reflecting the ability of other Tet family members to provide this enzymatic activity in nonhematopoietic tissues but not in hematopoietic cells. tet2(m/m) zebrafish are viable and fertile, providing an ideal model to dissect altered pathways in hematopoietic cells and, for small-molecule screens in embryos, to identify compounds with specific activity against tet2 mutant cells.
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http://dx.doi.org/10.1128/MCB.00971-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4323485PMC
March 2015

TET proteins and 5-methylcytosine oxidation in hematological cancers.

Immunol Rev 2015 Jan;263(1):6-21

Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, CA, USA.

DNA methylation has pivotal regulatory roles in mammalian development, retrotransposon silencing, genomic imprinting, and X-chromosome inactivation. Cancer cells display highly dysregulated DNA methylation profiles characterized by global hypomethylation in conjunction with hypermethylation of promoter CpG islands that presumably lead to genome instability and aberrant expression of tumor suppressor genes or oncogenes. The recent discovery of ten-eleven-translocation (TET) family dioxygenases that oxidize 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC) in DNA has led to profound progress in understanding the mechanism underlying DNA demethylation. Among the three TET genes, TET2 recurrently undergoes inactivating mutations in a wide range of myeloid and lymphoid malignancies. TET2 functions as a bona fide tumor suppressor particularly in the pathogenesis of myeloid malignancies resembling chronic myelomonocytic leukemia (CMML) and myelodysplastic syndromes (MDS) in human. Here we review diverse functions of TET proteins and the novel epigenetic marks that they generate in DNA methylation/demethylation dynamics and normal and malignant hematopoietic differentiation. The impact of TET2 inactivation in hematopoiesis and various mechanisms modulating the expression or activity of TET proteins are also discussed. Furthermore, we also present evidence that TET2 and TET3 collaborate to suppress aberrant hematopoiesis and hematopoietic transformation. A detailed understanding of the normal and pathological functions of TET proteins may provide new avenues to develop novel epigenetic therapies for treating hematological malignancies.
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http://dx.doi.org/10.1111/imr.12239DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4617313PMC
January 2015

Large conserved domains of low DNA methylation maintained by Dnmt3a.

Nat Genet 2014 Jan 24;46(1):17-23. Epub 2013 Nov 24.

1] Stem Cells and Regenerative Medicine Center, Department of Pediatrics and Molecular & Human Genetics, Baylor College of Medicine, Houston, Texas, USA. [2].

Gains and losses in DNA methylation are prominent features of mammalian cell types. To gain insight into the mechanisms that promote shifts in DNA methylation and contribute to changes in cell fate, including malignant transformation, we performed genome-wide mapping of 5-methylcytosine and 5-hydroxymethylcytosine in purified mouse hematopoietic stem cells. We discovered extended regions of low methylation (canyons) that span conserved domains frequently containing transcription factors and are distinct from CpG islands and shores. About half of the genes in these methylation canyons are coated with repressive histone marks, whereas the remainder are covered by activating histone marks and are highly expressed in hematopoietic stem cells (HSCs). Canyon borders are demarked by 5-hydroxymethylcytosine and become eroded in the absence of DNA methyltransferase 3a (Dnmt3a). Genes dysregulated in human leukemias are enriched for canyon-associated genes. The new epigenetic landscape we describe may provide a mechanism for the regulation of hematopoiesis and may contribute to leukemia development.
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http://dx.doi.org/10.1038/ng.2836DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3920905PMC
January 2014

Modulation of TET2 expression and 5-methylcytosine oxidation by the CXXC domain protein IDAX.

Nature 2013 May 7;497(7447):122-6. Epub 2013 Apr 7.

Division of Signaling and Gene Expression, La Jolla Institute for Allergy & Immunology, La Jolla, California 92037, USA.

TET (ten-eleven-translocation) proteins are Fe(ii)- and α-ketoglutarate-dependent dioxygenases that modify the methylation status of DNA by successively oxidizing 5-methylcytosine to 5-hydroxymethylcytosine, 5-formylcytosine and 5-carboxycytosine, potential intermediates in the active erasure of DNA-methylation marks. Here we show that IDAX (also known as CXXC4), a reported inhibitor of Wnt signalling that has been implicated in malignant renal cell carcinoma and colonic villous adenoma, regulates TET2 protein expression. IDAX was originally encoded within an ancestral TET2 gene that underwent a chromosomal gene inversion during evolution, thus separating the TET2 CXXC domain from the catalytic domain. The IDAX CXXC domain binds DNA sequences containing unmethylated CpG dinucleotides, localizes to promoters and CpG islands in genomic DNA and interacts directly with the catalytic domain of TET2. Unexpectedly, IDAX expression results in caspase activation and TET2 protein downregulation, in a manner that depends on DNA binding through the IDAX CXXC domain, suggesting that IDAX recruits TET2 to DNA before degradation. IDAX depletion prevents TET2 downregulation in differentiating mouse embryonic stem cells, and short hairpin RNA against IDAX increases TET2 protein expression in the human monocytic cell line U937. Notably, we find that the expression and activity of TET3 is also regulated through its CXXC domain. Taken together, these results establish the separate and linked CXXC domains of TET2 and TET3, respectively, as previously unknown regulators of caspase activation and TET enzymatic activity.
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http://dx.doi.org/10.1038/nature12052DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3643997PMC
May 2013

D-2-hydroxyglutarate produced by mutant IDH1 perturbs collagen maturation and basement membrane function.

Genes Dev 2012 Sep 27;26(18):2038-49. Epub 2012 Aug 27.

The Campbell Family Institute for Breast Cancer Research, Ontario Cancer Institute, University Health Network, Toronto, Ontario, Canada.

Isocitrate dehydrogenase-1 (IDH1) R132 mutations occur in glioma, but their physiological significance is unknown. Here we describe the generation and characterization of brain-specific Idh1 R132H conditional knock-in (KI) mice. Idh1 mutation results in hemorrhage and perinatal lethality. Surprisingly, intracellular reactive oxygen species (ROS) are attenuated in Idh1-KI brain cells despite an apparent increase in the NADP(+)/NADPH ratio. Idh1-KI cells also show high levels of D-2-hydroxyglutarate (D2HG) that are associated with inhibited prolyl-hydroxylation of hypoxia-inducible transcription factor-1α (Hif1α) and up-regulated Hif1α target gene transcription. Intriguingly, D2HG also blocks prolyl-hydroxylation of collagen, causing a defect in collagen protein maturation. An endoplasmic reticulum (ER) stress response induced by the accumulation of immature collagens may account for the embryonic lethality of these mutants. Importantly, D2HG-mediated impairment of collagen maturation also led to basement membrane (BM) aberrations that could play a part in glioma progression. Our study presents strong in vivo evidence that the D2HG produced by the mutant Idh1 enzyme is responsible for the above effects.
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http://dx.doi.org/10.1101/gad.198200.112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3444730PMC
September 2012

The SWI/SNF-like BAF complex is essential for early B cell development.

J Immunol 2012 Apr 16;188(8):3791-803. Epub 2012 Mar 16.

Department of Biological Sciences, Institute of Molecular Biology and Genetics, Research Center for Functional Cellulomics, Seoul National University, Seoul 151-742, Korea.

During the process of B cell development, transcription factors, such as E2A and Ebf1, have been known to play key roles. Although transcription factors and chromatin regulators work in concert to direct the expression of B lineage-specific genes, little is known about the involvement of regulators for chromatin structure during B lymphopoiesis. In this article, we show that deletion of Srg3/mBaf155, a scaffold subunit of the SWI/SNF-like BAF complex, in the hematopoietic lineage caused defects at both the common lymphoid progenitor stage and the transition from pre-pro-B to early pro-B cells due to failures in the expression of B lineage-specific genes, such as Ebf1 and Il7ra, and their downstream target genes. Moreover, mice that were deficient in the expression of Brg1, a subunit of the complex with ATPase activity, also showed defects in early B cell development. We also found that the expression of Ebf1 and Il7ra is directly regulated by the SWI/SNF-like BAF complex. Thus, our results suggest that the SWI/SNF-like BAF complex facilitates early B cell development by regulating the expression of B lineage-specific genes.
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http://dx.doi.org/10.4049/jimmunol.1103390DOI Listing
April 2012

TET2: epigenetic safeguard for HSC.

Blood 2011 Oct;118(17):4501-3

La Jolla Institute for Allergy and Immunology, USA.

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http://dx.doi.org/10.1182/blood-2011-08-373357DOI Listing
October 2011

Ten-Eleven-Translocation 2 (TET2) negatively regulates homeostasis and differentiation of hematopoietic stem cells in mice.

Proc Natl Acad Sci U S A 2011 Aug 22;108(35):14566-71. Epub 2011 Aug 22.

Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.

The Ten-Eleven-Translocation 2 (TET2) gene encodes a member of TET family enzymes that alters the epigenetic status of DNA by oxidizing 5-methylcytosine to 5-hydroxymethylcytosine (5hmC). Somatic loss-of-function mutations of TET2 are frequently observed in patients with diverse myeloid malignancies, including myelodysplastic syndromes, myeloproliferative neoplasms, and chronic myelomonocytic leukemia. By analyzing mice with targeted disruption of the Tet2 catalytic domain, we show here that Tet2 is a critical regulator of self-renewal and differentiation of hematopoietic stem cells (HSCs). Tet2 deficiency led to decreased genomic levels of 5hmC and augmented the size of the hematopoietic stem/progenitor cell pool in a cell-autonomous manner. In competitive transplantation assays, Tet2-deficient HSCs were capable of multilineage reconstitution and possessed a competitive advantage over wild-type HSCs, resulting in enhanced hematopoiesis into both lymphoid and myeloid lineages. In vitro, Tet2 deficiency delayed HSC differentiation and skewed development toward the monocyte/macrophage lineage. Our data indicate that Tet2 has a critical role in regulating the expansion and function of HSCs, presumably by controlling 5hmC levels at genes important for the self-renewal, proliferation, and differentiation of HSCs.
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http://dx.doi.org/10.1073/pnas.1112317108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3167529PMC
August 2011

Genome-wide mapping of 5-hydroxymethylcytosine in embryonic stem cells.

Nature 2011 May 8;473(7347):394-7. Epub 2011 May 8.

Harvard Medical School, Immune Disease Institute and Program in Cellular and Molecular Medicine, Children's Hospital Boston, Boston, Massachusetts 02115, USA.

5-hydroxymethylcytosine (5hmC) is a modified base present at low levels in diverse cell types in mammals. 5hmC is generated by the TET family of Fe(II) and 2-oxoglutarate-dependent enzymes through oxidation of 5-methylcytosine (5mC). 5hmC and TET proteins have been implicated in stem cell biology and cancer, but information on the genome-wide distribution of 5hmC is limited. Here we describe two novel and specific approaches to profile the genomic localization of 5hmC. The first approach, termed GLIB (glucosylation, periodate oxidation, biotinylation) uses a combination of enzymatic and chemical steps to isolate DNA fragments containing as few as a single 5hmC. The second approach involves conversion of 5hmC to cytosine 5-methylenesulphonate (CMS) by treatment of genomic DNA with sodium bisulphite, followed by immunoprecipitation of CMS-containing DNA with a specific antiserum to CMS. High-throughput sequencing of 5hmC-containing DNA from mouse embryonic stem (ES) cells showed strong enrichment within exons and near transcriptional start sites. 5hmC was especially enriched at the start sites of genes whose promoters bear dual histone 3 lysine 27 trimethylation (H3K27me3) and histone 3 lysine 4 trimethylation (H3K4me3) marks. Our results indicate that 5hmC has a probable role in transcriptional regulation, and suggest a model in which 5hmC contributes to the 'poised' chromatin signature found at developmentally-regulated genes in ES cells.
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http://dx.doi.org/10.1038/nature10102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3124347PMC
May 2011

Impaired hydroxylation of 5-methylcytosine in myeloid cancers with mutant TET2.

Nature 2010 Dec;468(7325):839-43

Department of Pathology, Harvard Medical School, Immune Disease Institute and Program in Cellular and Molecular Medicine, Children’s Hospital Boston, Boston, Massachusetts 02115, USA.

TET2 is a close relative of TET1, an enzyme that converts 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) in DNA. The gene encoding TET2 resides at chromosome 4q24, in a region showing recurrent microdeletions and copy-neutral loss of heterozygosity (CN-LOH) in patients with diverse myeloid malignancies. Somatic TET2 mutations are frequently observed in myelodysplastic syndromes (MDS), myeloproliferative neoplasms (MPN), MDS/MPN overlap syndromes including chronic myelomonocytic leukaemia (CMML), acute myeloid leukaemias (AML) and secondary AML (sAML). We show here that TET2 mutations associated with myeloid malignancies compromise catalytic activity. Bone marrow samples from patients with TET2 mutations displayed uniformly low levels of 5hmC in genomic DNA compared to bone marrow samples from healthy controls. Moreover, small hairpin RNA (shRNA)-mediated depletion of Tet2 in mouse haematopoietic precursors skewed their differentiation towards monocyte/macrophage lineages in culture. There was no significant difference in DNA methylation between bone marrow samples from patients with high 5hmC versus healthy controls, but samples from patients with low 5hmC showed hypomethylation relative to controls at the majority of differentially methylated CpG sites. Our results demonstrate that Tet2 is important for normal myelopoiesis, and suggest that disruption of TET2 enzymatic activity favours myeloid tumorigenesis. Measurement of 5hmC levels in myeloid malignancies may prove valuable as a diagnostic and prognostic tool, to tailor therapies and assess responses to anticancer drugs.
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http://dx.doi.org/10.1038/nature09586DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3003755PMC
December 2010

Chromatin remodeling, development and disease.

Mutat Res 2008 Dec 20;647(1-2):59-67. Epub 2008 Aug 20.

Department of Biological Sciences, Institute of Molecular Biology and Genetics, Seoul National University, Seoul 151-742, Republic of Korea.

Development is a stepwise process in which multi-potent progenitor cells undergo lineage commitment, differentiation, proliferation and maturation to produce mature cells with restricted developmental potentials. This process is directed by spatiotemporally distinct gene expression programs that allow cells to stringently orchestrate intricate transcriptional activation or silencing events. In eukaryotes, chromatin structure contributes to developmental progression as a blueprint for coordinated gene expression by actively participating in the regulation of gene expression. Changes in higher order chromatin structure or covalent modification of its components are considered to be critical events in dictating lineage-specific gene expression during development. Mammalian cells utilize multi-subunit nuclear complexes to alter chromatin structure. Histone-modifying complex catalyzes covalent modifications of histone tails including acetylation, methylation, phosphorylation and ubiquitination. ATP-dependent chromatin remodeling complex, which disrupts histone-DNA contacts and induces nucleosome mobilization, requires energy from ATP hydrolysis for its catalytic activity. Here, we discuss the diverse functions of ATP-dependent chromatin remodeling complexes during mammalian development. In particular, the roles of these complexes during embryonic and hematopoietic development are reviewed in depth. In addition, pathological conditions such as tumor development that are induced by mutation of several key subunits of the chromatin remodeling complex are discussed, together with possible mechanisms that underlie tumor suppression by the complex.
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http://dx.doi.org/10.1016/j.mrfmmm.2008.08.004DOI Listing
December 2008

BAF60a interacts with p53 to recruit the SWI/SNF complex.

J Biol Chem 2008 May 26;283(18):11924-34. Epub 2008 Feb 26.

Department of Biological Sciences, Institute of Molecular Biology and Genetics, Research Center for Functional Cellulomics, Seoul National University, Seoul 151-742, Korea.

To understand the tumor-suppressing mechanism of the SWI/SNF chromatin remodeling complex, we investigated its molecular relationship with p53. Using the pREP4-luc episomal reporter, we first demonstrated that p53 utilizes the chromatin remodeling activity of the SWI/SNF complex to initiate transcription from the chromatin-structured promoter. Among the components of the SWI/SNF complex, we identified BAF60a as a mediator of the interaction with p53 by the yeast two-hybrid assay. p53 directly interacted only with BAF60a, but not with other components of the SWI/SNF complex, such as BRG1, SRG3, SNF5, or BAF57. We found out that multiple residues at the amino acid 108-150 region of BAF60a were involved in the interaction with the tetramerization domain of p53. The N-terminal fragment of BAF60a containing the p53-interacting region as well as small interfering RNA for baf60a inhibited the SWI/SNF complex-mediated transcriptional activity of p53. The uncoupling of p53 with the SWI/SNF complex resulted in the repression of both p53-dependent apoptosis and cell cycle arrest by the regulation of target genes. These results suggest that the SWI/SNF chromatin remodeling complex is involved in the suppression of tumors by the interaction with p53.
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http://dx.doi.org/10.1074/jbc.M705401200DOI Listing
May 2008

Expression of SRG3, a core component of mouse SWI/SNF chromatin-remodeling complex, is regulated by cooperative interactions between Sp1/Sp3 and Ets transcription factors.

Biochem Biophys Res Commun 2005 Dec 26;338(3):1435-46. Epub 2005 Oct 26.

Department of Biological Sciences, Institute of Molecular Biology and Genetics, Research Center for Functional Cellomics, Seoul National University, Seoul 151-742, Republic of Korea.

SRG3, a mouse homolog of yeast SWI3 and human BAF155, is known to be a core component of SWI/SNF chromatin-remodeling complex. We have previously shown that SRG3 plays essential roles in early mouse embryogenesis, brain development, and T-cell development. SRG3 gene expression was differentially regulated depending on the developmental stages and exhibited tissue-specific pattern. In this study, we showed that the functional interactions between Sp and Ets family transcription factors are crucial for the SRG3 expression. Sp1 and Sp3 specifically bound to the two canonical Sp-binding sites (GC boxes) at -152 and -114, and a non-canonical Sp-binding site (CCTCCT motif) at -108 in the SRG3 promoter. Using Drosophila SL2 cells, we found that various Sp or Ets family members activate the SRG3 promoter through these Sp- or Ets-binding sites, respectively, in a dose-dependent manner. Intriguingly, different combinatorial expression of Ets and Sp factors in SL2 cells resulted in either strong synergistic activation or repression of the SRG3 promoter activity. Moreover, the Sp-mediated activation of SRG3 promoter required the intact Ets-binding element. Taken together, these results suggest that diverse interactions between Sp1/Sp3 and Ets factors are crucial for the SRG3 gene expression.
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http://dx.doi.org/10.1016/j.bbrc.2005.10.107DOI Listing
December 2005

Modulation of androgen receptor transactivation by the SWI3-related gene product (SRG3) in multiple ways.

Mol Cell Biol 2005 Jun;25(12):4841-52

Hormone Research Center and School of Biological Sciences and Technology, Chonnam National University, Gwangju 500-757, Republic of Korea.

The SWI3-related gene product (SRG3), a component of the mouse SWI/SNF complex, has been suggested to have an alternative function. Here, we demonstrate that in the prostate transactivation of the androgen receptor (AR) is modulated by SRG3 in multiple ways. The expression of SRG3, which is developmentally regulated in the prostate, is induced by androgen through AR. SRG3 in turn enhances the transactivation of AR, providing a positive feedback regulatory loop. The SRG3 coactivation of AR transactivation is achieved through the recruitment of coactivator SRC-1, the protein level of which is upregulated by SRG3, providing another pathway of positive regulation. Interestingly, SRG3 coactivation of AR transactivation is fully functional in BRG1/BRM-deficient C33A cells and the AR/SRG3/SRC-1 complex formed in vivo contains neither BRG1 nor BRM protein, suggesting the possibility of an SRG3 function independent of the SWI/SNF complex. Importantly, the AR/SRG3/SRC-1 complex occupies androgen response elements on the endogenous SRG3 and PSA promoter in an androgen-dependent manner in mouse prostate and LNCaP cells, respectively, inducing gene expression. These results suggest that the multiple positive regulatory mechanisms of AR transactivation by SRG3 may be important for the rapid proliferation of prostate cells during prostate development and regeneration.
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http://dx.doi.org/10.1128/MCB.25.12.4841-4852.2005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1140583PMC
June 2005

E2A/HEB and Id3 proteins control the sensitivity to glucocorticoid-induced apoptosis in thymocytes by regulating the SRG3 expression.

J Biol Chem 2004 May 11;279(21):21916-23. Epub 2004 Mar 11.

School of Biological Sciences and Institute of Molecular Biology and Genetics, Seoul National University, Seoul 151-742.

The E protein family transcription factors encoded by the E2A and HEB genes are known to play critical roles in the coordinate regulation of lymphocyte development. Previous studies have shown that T cell receptor (TCR) signals rapidly induce Id3, a dominant negative antagonist of E2A activity and allow thymocytes to survive selection events in the thymus. Here we show that SRG3 acts as a novel downstream target of E2A/HeLa E box-binding (HEB) complex and modulates glucocorticoid (GC) susceptibility in thymocytes in response to TCR signals. We have identified a putative E box element in the SRG3 promoter that is required for optimal promoter activity. The transcription factors E2A and HEB specifically associate with the E box element. Moreover, E2A-HEB heterodimers cooperated to activate SRG3 transcription, which was inhibited by the expression of Id proteins. TCR-mediated signals rapidly induced Id3 via MEK/ERK activation and thereby kept the E2A/HEB complex from binding to the E box element in the SRG3 promoter. Retroviral transduction of Id3 also repressed the SRG3 expression by inhibiting the E box binding activity of the E2A/HEB complex. Intriguingly, enforced Id3 expression conferred thymocyte resistance to GCs, which could be overcome by the overexpression of SRG3. Taken together, these results suggest that Id3 may enhance the viability of immature thymocytes by at least rendering them resistant to GCs through SRG3 down-regulation.
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http://dx.doi.org/10.1074/jbc.M402145200DOI Listing
May 2004

T cell receptor signaling inhibits glucocorticoid-induced apoptosis by repressing the SRG3 expression via Ras activation.

J Biol Chem 2004 May 11;279(21):21903-15. Epub 2004 Mar 11.

School of Biological Sciences and Institute of Molecular Biology and Genetics, Seoul National University, Seoul 151-742.

Activation of T cell antigen receptor (TCR) signaling inhibits glucocorticoid (GC)-induced apoptosis of T cells. However, the detailed mechanism regarding how activated T cells are protected from GC-induced apoptosis is unclear. Previously, we have shown that the expression level of SRG3, a murine homolog of BAF155 in humans, correlated well with the GC sensitivity of T cells either in vitro or in vivo. Intriguingly, the expression of SRG3 decreased upon positive selection in the thymus. Here we have shown that TCR signaling inhibits the SRG3 expression via Ras activation and thereby renders primary thymocytes and some thymoma cells resistant to GC-mediated apoptosis. By using pharmacological inhibitors, we have shown that Ras-mediated down-regulation of the SRG3 gene expression is mediated by MEK/ERK and phosphatidylinositol 3-kinase pathways. Moreover, TCR signals repressed the SRG3 transcription through the putative binding sites for E proteins and Ets family transcription factors in the proximal region of the SRG3 promoter. Introduction of mutations in these elements rendered the SRG3 promoter immune to the Ras or TCR signals. Taken together, these observations suggest that TCR signals result in GC desensitization in immature T cells by repressing SRG3 gene expression via Ras activation.
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http://dx.doi.org/10.1074/jbc.M402144200DOI Listing
May 2004