Cytosine modification of the dinucleotide CpG in the
DNA regulatory region is an important epigenetic marker during early embryo development, cellular differentiation, and
cancer progression. In clinical settings, such as anti-
cancer drug treatment, it is desirable to develop research tools to characterize DNA sequences affected by epigenetic perturbations. Here, we describe the construction and characterization of two fusion
endonucleases consisting of the (5)mCpG-binding domain of human MeCP2 (hMeCP2) and the cleavage domains of BmrI and FokI
restriction endonucleases (REases). The chimeric (CH)
endonucleases cleave M.HpaII (C(5)mCGG)-and
M.SssI ((5)mCpG)-modified
DNA. Unmodified
DNA and M.MspI-modified
DNA ((5)mCCGG) are poor substrates for the CH-
endonucleases. Sequencing cleavage products of modified lambda
DNA indicates that cleavage takes place outside the (5)mCpG recognition sequence, predominantly 4-17 bp upstream of the modified base (/N(4-17)(5)mCpG, where / indicates the cleavage site). Such (5)mCpG-specific
endonucleases will be useful to study CpG island modification of the regulatory regions of tumor suppressor genes, and for the construction of cell-specific and
tumor-specific modified CpG island databases.