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The SRA protein Np95 mediates epigenetic inheritance by recruiting Dnmt1 to methylated DNA

Abstract

DNA methyltransferase (cytosine-5) 1 (Dnmt1) is the principal enzyme responsible for maintenance of CpG methylation and is essential for the regulation of gene expression, silencing of parasitic DNA elements, genomic imprinting and embryogenesis1,2,3,4. Dnmt1 is needed in S phase to methylate newly replicated CpGs occurring opposite methylated ones on the mother strand of the DNA, which is essential for the epigenetic inheritance of methylation patterns in the genome. Despite an intrinsic affinity of Dnmt1 for such hemi-methylated DNA5, the molecular mechanisms that ensure the correct loading of Dnmt1 onto newly replicated DNA in vivo are not understood. The Np95 (also known as Uhrf1 and ICBP90) protein binds methylated CpG through its SET and RING finger-associated (SRA) domain6. Here we show that localization of mouse Np95 to replicating heterochromatin is dependent on the presence of hemi-methylated DNA. Np95 forms complexes with Dnmt1 and mediates the loading of Dnmt1 to replicating heterochromatic regions. By using Np95-deficient embryonic stem cells and embryos, we show that Np95 is essential in vivo to maintain global and local DNA methylation and to repress transcription of retrotransposons and imprinted genes. The link between hemi-methylated DNA, Np95 and Dnmt1 thus establishes key steps of the mechanism for epigenetic inheritance of DNA methylation.

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Figure 1: Local accumulation of Np95 is dependent on hemi-methylated DNA.
Figure 2: Impairment of DNA methylation status on Np95 gene inactivation.
Figure 3: Misexpression of hypomethylated genes on Np95 gene inactivation.
Figure 4: Np95 is required to target Dnmt1.

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Acknowledgements

This work was supported in part by a grant from the Genome Network Project (to H.K.), by the ‘Ground-based Research Program for Space Utilization’ promoted by the Japan Space Forum (H.K.) and by a grant-in-aid for Scientific Research on Priority Areas (germ-cell development, reprogramming and epigenetics, to M.O. and K.M.) from the Ministry of Education, Culture, Sports, Science and Technology, Japan. We thank W. Reik, N. Brockdorff, P. Burrows, H. Niwa, H. Sano, J. Strouboulis and M. Vidal for critical reading and reagents.

Author Contributions J. Sharif., K.O. and K.M. performed DNA methylation and gene expression analyses; M.M., Y.M.-K. and H.K. generated, maintained and performed phenotypic analyses of knockout mice; M.M., J. Shinga. and A.I. purified the protein complexes and performed mass spectrometry analyses; S. Takebayashi and M.O. performed immunofluorescence analysis; M.M., I.S. and S. Tajima performed the DNA methylation assay; and T.A.E. and T.T. performed statistical analyses. K.M., M.O. and H.K. designed the study, wrote the paper and contributed equally as co-senior authors. All authors discussed the results and commented on the manuscript.

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Correspondence to Haruhiko Koseki.

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Supplementary Information

The file contains Supplementary Figures S1-S8 with Legends and Supplementary Tables 1-3. This document shows biochemical properties of NP95 complexes, supplementary evidences for recognition of hemi-methylated DNA by Np95, generation and phenotypes of Np95-deficient mice, normal expression of Dnmt3a and -3b and cell-cycle progression in Np95-deficient ES cells and others. (PDF 776 kb)

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Sharif, J., Muto, M., Takebayashi, Si. et al. The SRA protein Np95 mediates epigenetic inheritance by recruiting Dnmt1 to methylated DNA. Nature 450, 908–912 (2007). https://doi.org/10.1038/nature06397

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