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



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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Figure 1: IDAX preferentially binds CpG-rich DNA.
Figure 2: IDAX downregulates TET2 protein through caspase activation.
Figure 3: Reciprocal relationship between TET2 and IDAX in mouse ESCs and human U937 cells.
Figure 4: Negative regulation of TET3 by its CXXC domain.

Accession codes


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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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




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.

Corresponding author

Correspondence to Anjana Rao.

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The authors declare no competing financial interests.

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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).

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