A Decade of Research on DNA Methyltransferases Concluded with a Publication in Nucleic Acids Research
The recent publication from the Laboratory of Structural Biology at the IIMCB and its partners summarizes almost 10 years of research efforts. This work follows the final fate of cytosine analogues in double-stranded DNA, from their initial interaction with DNA methylating enzymes up to their eventual removal.
Methylation of DNA provides an additional, “sequence-independent” layer of information and (epi)genetic memory. It can tell cells if a gene is to be active or not. So, it is not surprising that “misinformation” arising from aberrant methylation is a cause of multiple diseases, such as myelodysplastic syndrome (MDS), chronic myelomonocytic leukemia (CMML), acute myelogenous leukemia (AML), and β hemoglobinopathies. The damage resulting from aberrant epigenetic states could be limited with the help of small molecules interacting with DNA methyltransferases.
“It took us a long time to put together all elements of the puzzle, but it is extremely rewarding to see the final version of the paper in advanced online form in Nucleic Acids Research,” says Honorata Czapińska from the Laboratory of Structural Biology, IIMCB. “We used a simple prokaryotic M.MpeI methyltransferase as a proxy for the much more complex human DNMT1 enzyme. Some of the cytosine analogues (5-azacytidine and its derivative, 2ʹ-deoxy-5-azacytidine) are already used therapeutically as DNMT1 inhibitors. Our aim was to biochemically and structurally characterize complexes of cytosine derivatives with DNA methyltransferases to understand the fine details of their interaction with the enzymes and their final fate.”
The results proved much more interesting than the research team originally thought. They discovered that in the presence of small thiol-reducing agents typically used in work with methyltransferases, the compounds containing a halogen atom in the place that should be methylated, inhibit the enzyme but, with time, get modified in the same way as the cytosine substrate.
“Since it was widely believed that the inhibitory complexes with halogenated compounds are covalent and irreversible, it took time to understand and document our findings in a way that convinced the experts reviewing our work. Nonetheless, it was a fascinating journey towards the complete comprehension of the phenomenon,” says Marek Wojciechowski from the Plant Breeding and Acclimatization Institute. The understanding of this phenomenon can lead to finding new approaches to modify the (epi)genetic information.
The researchers emphasize that their work was more exciting thanks to making use of the recent theoretical analysis published by the head of the team, Prof. Matthias Bochtler, in Structure early this year. A deeper understanding of the differences between cryo-EM electrostatic potential and crystallographic electron density maps helped explain the appearance of halogenated compounds in the maps obtained by the IIMCB team and others with the two techniques.
The publication, entitled “Cytosine Analogues as DNA Methyltransferase Substrates,” in Nucleic Acids Research is available here: https://tinyurl.com/yyjp8m73
The work was done by scientists from the International Institute of Molecular and Cell Biology in Warsaw, Plant Breeding and Acclimatization Institute - National Research Institute in Radzików, Institute of Biochemistry and Biophysics PAS in Warsaw, and Department of Biochemistry of the University of Oxford.