Crosstalk between DNA Methylation and Histone Modifications

DNA methylation and histone modifications play a central role in the epigenetic regulation of gene expression and cell differentiation. DNA methyltransferase 1 (Dnmt1) is
the most ubiquitously expressed DNA methyltransferase and is essential for maintaining
DNA methylation patterns during semi-conservative DNA replication and DNA repair.
The fidelity and processivity of this process is crucial for genome stability and is based
on the specific recognition of hemimethylated CpG sites emerging at the replication
forks. Remarkably, a variety of proteins have been reported to interact with Dnmt1,
regulating the activation, stabilization and recruitment of Dnmt1 to specific sites and
regions. Recently, Uhrf1 has been found to interact with Dnmt1, indicating together
with genetic studies a central role in the maintenance of DNA methylation. However,
the exact mechanism how Uhrf1 plays a role in DNA methylation remains elusive.
Uhrf1 is a multi-domain protein harbouring distinct functional domains, which specifi-
cally recognize epigenetic marks such as DNA methylation and histone modifications.
To investigate the histone-tail binding specificities of Uhrf1, we developed an in vitro
histone-tail peptide binding assay using GFP-fusion proteins. Moreover, we expanded
this assay to 96-well micro plates, allowing high-throughput screening for histone-tail
peptide binding, DNA binding, protein-protein interactions, detection of endogenous
binding partners and quantification of post-translational modifications. Using this assay, we demonstrated that Uhrf1 preferentially binds to H3K9me3 via a tandem Tudor
domain. The binding is mediated by three highly conserved aromatic amino acids
that form an aromatic cage similar to the chromodomain of HP1 .
Furthermore, we characterized the second member of the Uhrf family, Uhrf2. Binding
assays demonstrate a cooperative interplay of Uhrf2 domains that induces preference
for hemimethylated DNA and enhances binding to H3K9me3 heterochromatin
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