Abstract:
:RNA:5-methylcytosine (m5C) methyltransferases are currently the focus of intense research following a series of high-profile reports documenting their physiological links to several diseases. However, no methods exist which permit the specific analysis of RNA:m5C methyltransferases in cells. Herein, we described how a combination of biophysical studies led us to identify distinct duplex-remodelling effects of m5C on RNA and DNA duplexes. Specifically, m5C induces a C3'-endo to C2'-endo sugar-pucker switch in CpG RNA duplex but triggers a B-to-Z transformation in CpG DNA duplex. Inspired by these different 'structural signatures', we developed a m5C-sensitive probe which fluoresces spontaneously in response to m5C-induced sugar-pucker switch, hence useful for sensing RNA:m5C methyltransferase activity. Through the use of this probe, we achieved real-time imaging and flow cytometry analysis of NOP2/Sun RNA methyltransferase 2 (NSUN2) activity in HeLa cells. We further applied the probe to the cell-based screening of NSUN2 inhibitors. The developed strategy could also be adapted for the detection of DNA:m5C methyltransferases. This was demonstrated by the development of DNA m5C-probe which permits the screening of DNA methyltransferase 3A inhibitors. To our knowledge, this study represents not only the first examples of m5C-responsive probes, but also a new strategy for discriminating RNA and DNA m5C methyltransferase activity in cells.
journal_name
Nucleic Acids Resjournal_title
Nucleic acids researchauthors
Yang T,Low JJA,Woon ECYdoi
10.1093/nar/gkz1047subject
Has Abstractpub_date
2020-01-10 00:00:00pages
e5issue
1eissn
0305-1048issn
1362-4962pii
5613678journal_volume
48pub_type
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