Structural dynamics of double-stranded DNA with epigenome modification.

Ayako Furukawa,Kyohei Arita,Kenji Sugase,Erik Walinda

doi:10.1093/nar/gkaa1210

Ayako Furukawa, Kyohei Arita + Show 2 more

Open Access

https://doi.org/10.1093/nar/gkaa1210

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Abstract

Modification of cytosine plays an important role in epigenetic regulation of gene expression and genome stability. Cytosine is converted to 5-methylcytosine (5mC) by DNA methyltransferase; in turn, 5mC may be oxidized to 5-hydroxymethylcytosine (5hmC) by ten-eleven translocation enzyme. The structural flexibility of DNA is known to affect the binding of proteins to methylated DNA. Here, we have carried out a semi-quantitative analysis of the dynamics of double-stranded DNA (dsDNA) containing various epigenetic modifications by combining data from imino 1H exchange and imino 1H R1ρ relaxation dispersion NMR experiments in a complementary way. Using this approach, we characterized the base-opening (kopen) and base-closing (kclose) rates, facilitating a comparison of the base-opening and -closing process of dsDNA containing cytosine in different states of epigenetic modification. A particularly striking result is the increase in the kopen rate of hemi-methylated dsDNA 5mC/C relative to unmodified or fully methylated dsDNA, indicating that the Watson–Crick base pairs undergo selective destabilization in 5mC/C. Collectively, our findings imply that the epigenetic modulation of cytosine dynamics in dsDNA mediates destabilization of the GC Watson–Crick base pair to allow base-flipping in living cells.

Highlights

Cytosine modifications are known to play an important role in epigenetic regulation of gene expression and genome (5mC) by DNA methyltransferases [3,4]; in turn, 5mC may be further oxidized to 5-hydroxymethylcytosine (5hmC)by ten-eleven translocation (TET) enzyme [5,6]
We designed six double-stranded DNA (dsDNA) sequences using the deoxyribonucleic acids carrying 5mC and 5hmC modifications based on the sequence recognized by UHRF1 [7]
C/C and 5hmC/C were control samples, each differing from 5mC/C at a singlemodified base, to analyse the effects of epigenetic cytosine modification on dsDNA dynamics

Summary

Introduction

Cytosine modifications are known to play an important role in epigenetic regulation of gene expression and genome (5mC) by DNA methyltransferases [3,4]; in turn, 5mC may be further oxidized to 5-hydroxymethylcytosine (5hmC)by ten-eleven translocation (TET) enzyme [5,6]. UHRF1 (ubiquitin-like containing PHD and RING finger domains 1) recognizes hemimethylated double-stranded DNA (dsDNA), in which one strand is methylated but the other is not. UHRF1 binds to the ‘flipped-out’ 5mC structure [7,8,9], but does not interact with either unmodified dsDNA or fullmethylated dsDNA, in which both strands are methylated [7]. One possibility is that DNA methylation increases the flexibility of dsDNA, thereby enabling the dsDNA at the position of 5mC to more frequently adopt a flipped-out structure or at least to adopt a more open structure that can be recognized by UHRF1. It has been shown that DNA methylation can alter DNA flexibility and affect the properties of proteins binding to the methylated

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