Role of TET proteins in myeloid malignancies

Summary

Principal Investigator: Anjana Rao
Abstract: DESCRIPTION (provided by applicant): The methylation status of DNA influences many biological processes during mammalian development and is known to be highly aberrant in cancer. In mammalian cells, DNA methylation occurs primarily as symmetrical methylation of cytosine in the context of the dinucleotide CpG, and the presence of high levels of 5-methyl- cytosine (5mC) at promoters is generally correlated with diminished gene expression. We recently discovered that the TET proteins TET1, TET2 and TET3 constitute a new family of dioxygenases that utilize molecular oxygen and the cofactors Fe(II) and 2-oxoglutarate to oxidize 5mC to 5-hydroxymethylcytosine (5hmC) in DNA. As a result, TET proteins alter DNA methylation status in a novel and hitherto unprecedented way. Simultaneously, several labs reported that TET2 mutations are frequently associated with myelodysplastic syndromes (MDS), myeloproliferative neoplasms (MPN) and myeloid malignancies such as chronic myelomonocytic leukemia (CMML) and acute myeloid leukemia (AML). Two other genes frequently mutated in these patients include those encoding the DNA methyltransferase DNMT3A and the polycomb group protein ASXL1, a component of a complex that deubiquitinates H2A. In this proposal we will explore, at a molecular level using appropriate mouse models, the roles of Tet2 and Tet3 in hematopoiesis and myeloid function (Aim 1). In Aims 2 and 3, we will investigate the relation of Tet2/ Tet3 to Dnmt3a and Asxl1 respectively. We have developed many methods and generated many reagents relevant to these proposed studies, including quantitative methods to measure overall genomic levels of 5hmC in bone marrow samples from patients with MDS/ MPN/ CMML/ secondary AML;and innovative strategies for mapping the genomic location of 5hmC and profiling 5hmC at single-base resolution. We have generated mice with conditional disruption of the Tet2 and Tet3 genes;and have uncovered a novel relation between 5hmC and the polycomb complex. WE have shown in ES cells that 5hmC is present predominantly at the promoters of genes that are (i) inactive but "poised" to be expressed upon ES cell differentiation;(ii) bear dual ("bivalent") H3K4me3 and H3K27me3 marks;and (iii) are bound by components of the polycomb complexes PRC1 and PRC2 (the H3K27me3 mark is deposited by PRC2). By defining the genome-wide changes in DNA methylation, DNA hydroxymethylation and histone modifications that occur as a result of loss of function of Tet2 and Tet3 and selected leukemia-associated mutations in Dnmt3a and Asxl1, our proposed studies will provide fundamental insights into how these proteins control chromatin structure and the epigenetic landscape at their target genes. This information will help us understand how changes that occur as a result of somatic mutations in these gene products in stem cells might predispose to myeloid cancers in humans.
Funding Period: 2012-04-10 - 2017-03-31
more information: NIH RePORT

Top Publications

  1. pmc Mutational spectrum analysis of chronic myelomonocytic leukemia includes genes associated with epigenetic regulation: UTX, EZH2, and DNMT3A
    Anna M Jankowska
    Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, Cleveland Clinic, Cleveland, OH, USA
    Blood 118:3932-41. 2011
  2. pmc Large conserved domains of low DNA methylation maintained by Dnmt3a
    MiRa Jeong
    1 Stem Cells and Regenerative Medicine Center, Department of Pediatrics and Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas, USA 2
    Nat Genet 46:17-23. 2014
  3. pmc Global epigenomic reconfiguration during mammalian brain development
    Ryan Lister
    Genomic Analysis Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037, USA
    Science 341:1237905. 2013
  4. pmc Vitamin C induces Tet-dependent DNA demethylation and a blastocyst-like state in ES cells
    Kathryn Blaschke
    Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, Department of Obstetrics and Gynecology and Center for Reproductive Sciences, University of California San Francisco, San Francisco, California 94143, USA
    Nature 500:222-6. 2013
  5. pmc TETonic shift: biological roles of TET proteins in DNA demethylation and transcription
    William A Pastor
    Sanford Consortium for Regenerative Medicine and La Jolla Institute for Allergy and Immunology, 9420 Athena Circle, La Jolla, California 92037, USA
    Nat Rev Mol Cell Biol 14:341-56. 2013
  6. pmc Modulation of TET2 expression and 5-methylcytosine oxidation by the CXXC domain protein IDAX
    Myunggon Ko
    Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, California 92037, USA
    Nature 497:122-6. 2013
  7. pmc DNA methylation and methylcytosine oxidation in cell fate decisions
    Kian Peng Koh
    KU Leuven Department of Development and Regeneration and Stem Cell Institute Leuven, 3000 Leuven, Belgium
    Curr Opin Cell Biol 25:152-61. 2013
  8. pmc O-GlcNAcylation and 5-methylcytosine oxidation: an unexpected association between OGT and TETs
    Anand Balasubramani
    Sanford Consortium for Regenerative Medicine and La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA
    Mol Cell 49:618-9. 2013
  9. pmc Stage-specific roles for tet1 and tet2 in DNA demethylation in primordial germ cells
    John J Vincent
    Molecular Biology Institute, University of California, Los Angeles, Los Angeles, CA 90095, USA
    Cell Stem Cell 12:470-8. 2013
  10. pmc The GLIB technique for genome-wide mapping of 5-hydroxymethylcytosine
    William A Pastor
    La Jolla Institute for Allergy and Immunology, La Jolla, California, USA
    Nat Protoc 7:1909-17. 2012

Research Grants

Detail Information

Publications16

  1. pmc Mutational spectrum analysis of chronic myelomonocytic leukemia includes genes associated with epigenetic regulation: UTX, EZH2, and DNMT3A
    Anna M Jankowska
    Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, Cleveland Clinic, Cleveland, OH, USA
    Blood 118:3932-41. 2011
    ..Various mutant genotype combinations were observed, indicating molecular heterogeneity in CMML. Our results suggest that molecular defects affecting distinct pathways can lead to similar clinical phenotypes...
  2. pmc Large conserved domains of low DNA methylation maintained by Dnmt3a
    MiRa Jeong
    1 Stem Cells and Regenerative Medicine Center, Department of Pediatrics and Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas, USA 2
    Nat Genet 46:17-23. 2014
    ..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. ..
  3. pmc Global epigenomic reconfiguration during mammalian brain development
    Ryan Lister
    Genomic Analysis Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037, USA
    Science 341:1237905. 2013
    ....
  4. pmc Vitamin C induces Tet-dependent DNA demethylation and a blastocyst-like state in ES cells
    Kathryn Blaschke
    Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, Department of Obstetrics and Gynecology and Center for Reproductive Sciences, University of California San Francisco, San Francisco, California 94143, USA
    Nature 500:222-6. 2013
    ..Collectively, the results of this study establish vitamin C as a direct regulator of Tet activity and DNA methylation fidelity in ES cells...
  5. pmc TETonic shift: biological roles of TET proteins in DNA demethylation and transcription
    William A Pastor
    Sanford Consortium for Regenerative Medicine and La Jolla Institute for Allergy and Immunology, 9420 Athena Circle, La Jolla, California 92037, USA
    Nat Rev Mol Cell Biol 14:341-56. 2013
    ....
  6. pmc Modulation of TET2 expression and 5-methylcytosine oxidation by the CXXC domain protein IDAX
    Myunggon Ko
    Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, California 92037, USA
    Nature 497:122-6. 2013
    ..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...
  7. pmc DNA methylation and methylcytosine oxidation in cell fate decisions
    Kian Peng Koh
    KU Leuven Department of Development and Regeneration and Stem Cell Institute Leuven, 3000 Leuven, Belgium
    Curr Opin Cell Biol 25:152-61. 2013
    ....
  8. pmc O-GlcNAcylation and 5-methylcytosine oxidation: an unexpected association between OGT and TETs
    Anand Balasubramani
    Sanford Consortium for Regenerative Medicine and La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA
    Mol Cell 49:618-9. 2013
    ....
  9. pmc Stage-specific roles for tet1 and tet2 in DNA demethylation in primordial germ cells
    John J Vincent
    Molecular Biology Institute, University of California, Los Angeles, Los Angeles, CA 90095, USA
    Cell Stem Cell 12:470-8. 2013
    ..Instead, Tet1 and Tet2 have a locus-specific role in shaping the PGC epigenome during subsequent development...
  10. pmc The GLIB technique for genome-wide mapping of 5-hydroxymethylcytosine
    William A Pastor
    La Jolla Institute for Allergy and Immunology, La Jolla, California, USA
    Nat Protoc 7:1909-17. 2012
    ..GLIB is capable of quantitatively tagging and precipitating fragments containing a single 5hmC molecule. Sample preparation and GLIB can be conducted in 2-3 d...
  11. pmc The anti-CMS technique for genome-wide mapping of 5-hydroxymethylcytosine
    Yun Huang
    La Jolla Institute for Allergy and Immunology, La Jolla, California, USA
    Nat Protoc 7:1897-908. 2012
    ..Moreover, it does not enrich for CA and CT repeats, as noted for 5hmC DNA IP using antibodies to 5hmC. The anti-CMS protocol takes 3 d to complete...
  12. pmc D-2-hydroxyglutarate produced by mutant IDH1 perturbs collagen maturation and basement membrane function
    Masato Sasaki
    The Campbell Family Institute for Breast Cancer Research, Ontario Cancer Institute, University Health Network, Toronto, Ontario, Canada
    Genes Dev 26:2038-49. 2012
    ..Our study presents strong in vivo evidence that the D2HG produced by the mutant Idh1 enzyme is responsible for the above effects...
  13. pmc PGC7, H3K9me2 and Tet3: regulators of DNA methylation in zygotes
    Jinsuk Kang
    La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA
    Cell Res 23:6-9. 2013
    ....
  14. pmc Nontargeted nucleotide analysis based on benzoylhistamine labeling-MALDI-TOF/TOF-MS: discovery of putative 6-oxo-thymine in DNA
    Poguang Wang
    Pharmaceutical Science and Barnett Institute, Northeastern University, Boston, Massachusetts 02115, United States
    Anal Chem 84:3811-9. 2012
    ..The method provides deoxynucleotide-specific detection, accurate measurement of molecular ions, high sensitivity, semiquantitation, and, to the extent studied to date, normalization of response within a factor of <3...
  15. pmc Ten-Eleven-Translocation 2 (TET2) negatively regulates homeostasis and differentiation of hematopoietic stem cells in mice
    Myunggon Ko
    Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA
    Proc Natl Acad Sci U S A 108:14566-71. 2011
    ..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...
  16. pmc Distinct roles of the methylcytosine oxidases Tet1 and Tet2 in mouse embryonic stem cells
    Yun Huang
    Divisions of Signalling and Gene Expression and Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037
    Proc Natl Acad Sci U S A 111:1361-6. 2014
    ..Together, the data point to a complex interplay between Tet1 and Tet2 in mESC, and to distinct roles for these two proteins in regulating promoter, exon, and polyadenylation site usage in cells. ..

Research Grants30

  1. Epigenetic regulation of transcription by mixed lineage leukemia protein MLL1
    Yali Dou; Fiscal Year: 2013
    ..This project is to understand the fundamental mechanism for these changes and thus, the cause of the disease. It will set up the foundation for potential medical interference in the future. ..
  2. DNA Methylation Control Hematopoietic Stem Cell Lineage Differentiation
    GRANT ANTHONY CHALLEN; Fiscal Year: 2013
    ..I believe the basic research outlined in this project can contribute to the fundamental understanding of epigenetic regulation of stem cell function and has direct relevance to progression of human disease states. ..
  3. HORMONAL REGULATION OF BLOOD PRESSURE
    Michal Laniado Schwartzman; Fiscal Year: 2013
    ..ular tone, in the pathophysiology of hypertension and cardiovascular disease. ..
  4. Targeting the aberrant kinome-epigenome in AML
    Guido Marcucci; Fiscal Year: 2013
    ....
  5. Regulated Accessibility of a cytokine gene locus.
    Anjana Rao; Fiscal Year: 2013
    ..In this proposal we plan to investigate the role of TET proteins in T cells, a major class of cells in the immune system that protect the organism from attack by bacteria, viruses and other pathogens including cancer cells. ..