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

Nature volume 468, pages 839–843 (2010)Cite this article

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Abstract

TET2 is a close relative of TET1, an enzyme that converts 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) in DNA1,2. 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 malignancies3. 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)4,5,6,7,8,9,10,11,12. 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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Figure 1: The catalytic activity of Tet2 is compromised by mutations in predicted catalytic residues.
Figure 2: TET2 mutational status correlates with 5hmC levels in patients with myeloid malignancies.
Figure 4: Relation of 5hmC levels to DNA methylation status.
Figure 3: Tet2 regulates myeloid differentiation.

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Primary accessions

Gene Expression Omnibus

Data deposits

Data have been deposited at GEO under accession number GSE25706 (methylation status of each CpG site (beta value) can be found in Supplementary Table 3).

Change history

  • 09 December 2010

    A panel was replaced in Fig. 1c and data accession information was added.

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Acknowledgements

This work was supported by NIH grants R01 AI44432 and RC1 DA028422 (to A.R.), NIH grants K24 HL077522 and R01 HL098522, an Established Investigator award from the Aplastic Anemia & MDS Foundation, and an award from the Bob Duggan Memorial Research Fund (to J.P.M), NIH grant R01 HG4069 (to X.S.L.) and a pilot grant from Harvard Catalyst, The Harvard Clinical and Translational Science Center (NIH Grant #1 UL1 RR 025758-02, to S.A.). Y.H. was supported by postdoctoral fellowships from the GlaxoSmithKline-Immune Disease Institute (GSK-IDI) Alliance and the Leukemia and Lymphoma Society of America. H.S.B. is supported by a postdoctoral fellowship from the GSK-IDI Alliance.

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  1. Myunggon Ko, Yun Huang, Jungeun An & Anjana Rao

    Present address: Present address: La Jolla Institute for Allergy and Immunology, La Jolla, California 92037, USA (M.K., Y.H., J.A., A.R.).,

  2. Myunggon Ko and Yun Huang: These authors contributed equally to this work.

Authors and Affiliations

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

    Myunggon Ko, Yun Huang, Utz J. Pape, Mamta Tahiliani, Hozefa S. Bandukwala, Jungeun An, Edward D. Lamperti, Kian Peng Koh & Anjana Rao

  2. Department of Translational Hematology and Oncology Research, and Department of Hematologic Oncology and Blood Disorders, Taussig Cancer Institute, Cleveland Clinic, Cleveland, 44195, Ohio, USA

    Anna M. Jankowska, Rebecca Ganetzky & Jaroslaw P. Maciejewski

  3. Department of Biostatistics and Computational Biology, Dana-Farber Cancer Institute and Harvard School of Public Health, Boston, 02115, Massachusetts, USA

    Utz J. Pape & X. Shirley Liu

  4. National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, 20894, Maryland, USA

    L. Aravind

  5. Division of Pediatric Hematology/Oncology, Children's Hospital Boston and Dana-Farber Cancer Institute, Harvard Stem Cell Institute, Boston, 02115, Massachusetts, USA

    Suneet Agarwal

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Contributions

M.K. analysed the biochemical effects of patient-associated TET2 mutations and performed the in vitro differentiation studies; Y.H. generated and characterized the anti-CMS antiserum, developed the quantitative dot-blot assay and quantified 5hmC in DNA samples from patients and healthy controls. A.M.J., R.G. and J.P.M. provided patient and control DNA for 5hmC quantification, performed DNA methylation arrays and analysed TET2 mutational status in patients. U.J.P. and X.S.L. carried out the statistical analysis of 5hmC levels and methylation data; M.T., H.S.B. and K.P.K. provided critical reagents; J.A. and E.D.L. contributed to molecular cloning and mouse maintenance respectively; and L.A. and S.A. provided essential intellectual input. A.R. set overall goals, coordinated collaborations and wrote the manuscript.

Corresponding authors

Correspondence to Jaroslaw P. Maciejewski or Anjana Rao.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Methods, Supplementary Text, legends for Supplementary Tables 1-2 and Supplementary Figures 1-11 with legends. The Supplementary Methods were added on 8 December 2010. (PDF 3833 kb)

Supplementary Table 1

This file contains Supplementary Table 1. (XLS 85 kb)

Supplementary Table 2

This file contains Supplementary Table 2. (XLS 1382 kb)

Supplementary Table 3

The file contains Supplementary Table 3. Supplementary Table 3 was added on 8 December 2010. (ZIP 25684 kb)

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Ko, M., Huang, Y., Jankowska, A. et al. Impaired hydroxylation of 5-methylcytosine in myeloid cancers with mutant TET2. Nature 468, 839–843 (2010). https://doi.org/10.1038/nature09586

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