Abstract
The multi-domain protein UHRF1 is essential for DNA methylation maintenance and binds DNA via a base-flipping mechanism with a preference for hemi-methylated CpG sites. We investigated its binding to hemi- and symmetrically modified DNA containing either 5-methylcytosine (mC), 5-hydroxymethylcytosine (hmC), 5-formylcytosine (fC), or 5-carboxylcytosine (caC). Our experimental results indicate that UHRF1 binds symmetrically carboxylated and hybrid methylated/carboxylated CpG dyads in addition to its previously reported substrates. Complementary molecular dynamics simulations provide a possible mechanistic explanation of how the protein could differentiate between modification patterns. First, we observe different local binding modes in the nucleotide binding pocket as well as the protein's NKR finger. Second, both DNA modification sites are coupled through key residues within the NKR finger, suggesting a communication pathway affecting protein-DNA binding for carboxylcytosine modifications. Our results suggest a possible additional function of the hemi-methylation reader UHRF1 through binding of carboxylated CpG sites. This opens the possibility of new biological roles of UHRF1 beyond DNA methylation maintenance and of oxidised methylcytosine derivates in epigenetic regulation.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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5-Methylcytosine / metabolism*
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Animals
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Base Sequence
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CCAAT-Enhancer-Binding Proteins / chemistry
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CCAAT-Enhancer-Binding Proteins / metabolism*
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CpG Islands / genetics*
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Cytosine / analogs & derivatives*
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Cytosine / metabolism
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Epigenesis, Genetic
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Mice
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Molecular Dynamics Simulation
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Protein Binding
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Protein Domains
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Substrate Specificity
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Ubiquitin-Protein Ligases / chemistry
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Ubiquitin-Protein Ligases / metabolism*
Substances
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5-carboxylcytosine
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CCAAT-Enhancer-Binding Proteins
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5-Methylcytosine
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Cytosine
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Ubiquitin-Protein Ligases
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Uhrf1 protein, mouse
Grants and funding
This work was supported by the Deutsche Forschungsgemeinschaft (SFB 1035/A10 and SFB749/C08 to IA, SFB 1064/A17 and SFB 1243/A01 to HL, CIPSM to IA and HL, CA198/10-1 to MCC). MS was supported by the TUM International Graduate School of Science and Engineering (IGSSE). CB and CT were supported by the International Max Planck Research School for Molecular Life Sciences (IMPRS-LS). AS was trained and supported by the graduate school GRK1721 of the Deutsche Forschungsgemeinschaft. PZ received support from the “Vereinigung von Freunden der Technischen Universität Darmstadt”. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.