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Intragenic DNA methylation alters chromatin structure and elongation efficiency in mammalian cells

Nature Structural & Molecular Biology volume 11, pages 1068–1075 (2004)Cite this article

Abstract

Transcriptional silencing in mammals is often associated with promoter methylation. However, a considerable number of genomic methylated CpGs exist in transposable elements, which are frequently found in intronic regions. To determine whether intragenic methylation influences transcription efficiency, we used the Cre/loxP-based system, RMCE, to introduce a transgene, methylated exclusively in a region downstream of the promoter, into a specific genomic site. This methylation pattern was maintained in vivo, and yielded a clear decrease in transgene expression relative to an unmethylated control. Notably, RNA polymerase II (Pol II) was depleted exclusively in the methylated region, as was histone H3 di- and trimethylated on Lys4 and acetylated on Lys9 and Lys14. As the methylated region adopts a closed chromatin structure in vivo, we propose that dense intragenic DNA methylation in mammalian cells initiates formation of a chromatin structure that reduces the efficiency of Pol II elongation.

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Figure 1: Generation of the methylated reporter cassette.
Figure 2: Intragenic methylation influences expression level.
Figure 3: Methylation state is preserved in vivo.
Figure 4: Intragenic methylation does not affect Pol II initiation.
Figure 5: Pol II is depleted exclusively in the methylated region.
Figure 6: Intragenic DNA methylation directs a local alteration in histone modifications.
Figure 7: Intragenic DNA methylation generates a compact chromatin structure.

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Acknowledgements

This work was supported by US National Institutes of Health grants DK44746 and HL57620 to M.G. M.L. is a special fellow of the Leukemia and Lymphoma Society. We thank E. Bouhassira for the RL5 MEL cell line, G. Peters for the plasmid p16EXON1α, D. Schübeler, A. Krumm and T. Sawado for technical advice and discussion, and R. Gardner and T. Ragoczy for comments on the manuscript.

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Authors and Affiliations

  1. Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, 98109, Washington, USA

    Matthew C Lorincz, David R Dickerson & Mark Groudine

  2. Department of Radiation Oncology, University of Washington School of Medicine, Seattle, 98195, Washington, USA

    Mike Schmitt & Mark Groudine

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Correspondence to Matthew C Lorincz.

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Supplementary Table 1

Primers used in this study. (PDF 21 kb)

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Lorincz, M., Dickerson, D., Schmitt, M. et al. Intragenic DNA methylation alters chromatin structure and elongation efficiency in mammalian cells. Nat Struct Mol Biol 11, 1068–1075 (2004). https://doi.org/10.1038/nsmb840

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