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A methylation and phosphorylation switch between an adjacent lysine and serine determines human DNMT1 stability

Nature Structural & Molecular Biology volume 18, pages 42–48 (2011)Cite this article

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Abstract

The protein lysine methyltransferase SET7 regulates DNA methyltransferase-1 (DNMT1) activity in mammalian cells by promoting degradation of DNMT1 and thus allows epigenetic changes via DNA demethylation. Here we reveal an interplay between monomethylation of DNMT1 Lys142 by SET7 and phosphorylation of DNMT1 Ser143 by AKT1 kinase. These two modifications are mutually exclusive, and structural analysis suggests that Ser143 phosphorylation interferes with Lys142 monomethylation. AKT1 kinase colocalizes and directly interacts with DNMT1 and phosphorylates Ser143. Phosphorylated DNMT1 peaks during DNA synthesis, before DNMT1 methylation. Depletion of AKT1 or overexpression of dominant-negative AKT1 increases methylated DNMT1, resulting in a decrease in DNMT1 abundance. In mammalian cells, phosphorylated DNMT1 is more stable than methylated DNMT1. These results reveal cross-talk on DNMT1, through modifications mediated by AKT1 and SET7, that affects cellular DNMT1 levels.

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Figure 1: Lysine methylation and serine phosphorylation on DNMT1.
Figure 2: Structure of the SET7–DNMT1 complex.
Figure 3: DNMT1 association with AKT1 kinase.
Figure 4: AKT1 kinase–mediated phosphorylation stabilizes DNMT1.

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Acknowledgements

We thank J. Benner for nucleotide analysis of the genomic DNA, S.M. Kinney for critical reading of this manuscript, H.G. Chin (New England Biolabs) for recombinant enzymes, X. Zhang for insightful discussion, and D.G. Comb, R.J. Roberts, J.V. Ellard at New England Biolabs for supporting the basic research. The work in the Cheng laboratory was supported by US National Institutes of Health (NIH) grants GM049245 and GM068680. X.C. is a Georgia Research Alliance Eminent Scholar.

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Authors and Affiliations

  1. New England Biolabs, Ipswich, Massachusetts, USA

    Pierre-Olivier Estève, Mala Samaranayake, George R Feehery & Sriharsa Pradhan

  2. Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia, USA

    Yanqi Chang, Anup K Upadhyay, John R Horton & Xiaodong Cheng

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  1. Pierre-Olivier Estève
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Contributions

P.-O.E. performed cell biology and biochemistry experiments. M.S. made constructs, tested kinetics and performed the pull-down assay. G.R.F. performed quantitative PCR. Y.C. performed SET7 purifications, MS-based methylation assays on DNMT1 peptide, mutagenesis of K142R and crystallization of SET7–DNMT1 peptide. A.K.U. purified the DNMT1 N-terminal domain and performed MS-based methylation assays on this fragment. J.R.H. performed crystallographic experiments. X.C. and S.P. organized and analyzed data and wrote the manuscript.

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Correspondence to Sriharsa Pradhan.

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Estève, PO., Chang, Y., Samaranayake, M. et al. A methylation and phosphorylation switch between an adjacent lysine and serine determines human DNMT1 stability. Nat Struct Mol Biol 18, 42–48 (2011). https://doi.org/10.1038/nsmb.1939

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