Modulation of TET2 expression and 5-methylcytosine oxidation by the CXXC domain protein IDAX (original) (raw)

• Letter
• Published: 07 April 2013
• Jungeun An1 na1,
• Hozefa S. Bandukwala1,
• Lukas Chavez1,
• Tarmo Äijö1,2,
• William A. Pastor1 nAff7,
• Matthew F. Segal1,
• Huiming Li3 nAff7,
• Kian Peng Koh3 nAff7,
• Harri Lähdesmäki2,
• Patrick G. Hogan1,
• L. Aravind4 &
• …
• Anjana Rao1,3,5

Nature volume 497, pages 122–126 (2013)Cite this article

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Abstract

TET (ten-eleven-translocation) proteins are Fe(ii)- and α-ketoglutarate-dependent dioxygenases1,2,3 that modify the methylation status of DNA by successively oxidizing 5-methylcytosine to 5-hydroxymethylcytosine, 5-formylcytosine and 5-carboxycytosine1,3,4,5, potential intermediates in the active erasure of DNA-methylation marks5,6. Here we show that IDAX (also known as CXXC4), a reported inhibitor of Wnt signalling7 that has been implicated in malignant renal cell carcinoma8 and colonic villous adenoma9, 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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Accession codes

Accessions

Gene Expression Omnibus

Data deposits

The ChIP-seq data have been deposited in the Gene Expression Omnibus (GEO) under accession number GSE42958.

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Acknowledgements

We thank G. Seumois, M. Ku and J. Day for help with library preparation, B. Ren for use of his Illumina Hi-Seq 2000, J. A. Zepeda-Martínez for the recombinant Flag–TET2CD, and members of the Rao laboratory for discussions. This work was supported by National Institutes of Health (NIH) R01 grants HD065812 and CA151535, grant RM-01729 from the California Institute of Regenerative Medicine and Translational Research, grant TRP 6187-12 from the Leukemia and Lymphoma Society (to A.R.) and NIH R01 grant AI40127 (to P.G.H. and A.R). We also gratefully acknowledge a Special Fellow Award from the Leukemia and Lymphoma Society (to M.K.), postdoctoral fellowships from the Lady Tata Memorial Trust and from the GlaxoSmithKline-Immune Disease Institute Alliance (to H.S.B.) and a predoctoral graduate research fellowship from the National Science Foundation (to W.A.P.).

Author information

Author notes

1. William A. Pastor, Huiming Li & Kian Peng Koh
   Present address: Present addresses: Department of Molecular, Cell and Developmental Biology, University of California at Los Angeles, Terasaki Life Sciences Building, 610 Charles Young Drive East, Los Angeles, California 90095-723905, USA (W.A.P.); Bristol-Myers Squibb, 700 Bay Road, Redwood City, California 94063, USA (H.Li); KU Leuven Department of Development and Regeneration & Stem Cell Institute Leuven, Herestraat 49, 3000 Leuven, Belgium (K.P.K.).,
2. Myunggon Ko and Jungeun An: These authors contributed equally to this work.

Authors and Affiliations

1. Division of Signaling and Gene Expression, La Jolla Institute for Allergy & Immunology, La Jolla, California 92037, USA,
   Myunggon Ko, Jungeun An, Hozefa S. Bandukwala, Lukas Chavez, Tarmo Äijö, William A. Pastor, Matthew F. Segal, Patrick G. Hogan & Anjana Rao
2. Department of Information and Computer Science, Aalto University School of Science, FI-00076 Aalto, Finland
   Tarmo Äijö & Harri Lähdesmäki
3. Harvard Medical School and Program in Cellular and Molecular Medicine, Children’s Hospital, Boston, Massachusetts 02115, USA,
   Huiming Li, Kian Peng Koh & Anjana Rao
4. National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894, USA,
   L. Aravind
5. Sanford Consortium for Regenerative Medicine, La Jolla, California 92037, USA,
   Anjana Rao

Authors

1. Myunggon Ko
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2. Jungeun An
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3. Hozefa S. Bandukwala
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4. Lukas Chavez
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5. Tarmo Äijö
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6. William A. Pastor
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8. Huiming Li
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11. Patrick G. Hogan
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Contributions

L.A., P.G.H. and A.R. conceived the project and supervised project planning and execution. M.K. and J.A. performed cellular and molecular experiments including ChIP-seq, gene knockdown, establishment of stable cell lines, site-directed mutagenesis, dot blot, immunocytochemistry, in vitro caspase and TET assays, and in vitro differentiation studies. J.A. performed the in-cell western blots. H.S.B. obtained the initial data showing downregulation of TET2 protein by IDAX. M.K. conducted the electrophoretic mobility shift assays with help from W.A.P. and M.F.S. H.Li and P.G.H. generated the homology model of the IDAX CXXC domain. K.P.K. provided mRNAs from ESC samples. L.C., T.A. and H.Lähdesmäki performed the bioinformatic analyses of ChIP-seq data. M.K. and A.R. wrote the manuscript with input from other authors.

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Correspondence toAnjana Rao.

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Ko, M., An, J., Bandukwala, H. et al. Modulation of TET2 expression and 5-methylcytosine oxidation by the CXXC domain protein IDAX.Nature 497, 122–126 (2013). https://doi.org/10.1038/nature12052

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• Received: 15 February 2012
• Accepted: 28 February 2013
• Published: 07 April 2013
• Issue Date: 02 May 2013
• DOI: https://doi.org/10.1038/nature12052

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Editorial Summary

IDAX regulates TET2 protein expression

TET family proteins modify the methylation status of DNA by oxidizing 5-methylcytosine to 5-hydroxymethylcytosine (5hmC, sometimes called the 'fifth base' of DNA) and other intermediates. TET1 and TET3 contain a CXXC domain but the ancestral CXXC domain of TET2 is encoded by a distinct gene, IDAX (or CXXC4). This paper demonstrates that IDAX binds unmethylated CpG-rich DNA via its CXXC domain and recruits TET2. The separate and linked CXXC domains of TET2 and TET3 are shown to act as regulators of caspase activation and TET enzymatic activity. The authors suggest that future studies should focus on the genomic targets of TET2, IDAX and the IDAX-related protein CXXC5 in normal development and in cancer.