Khoddami, V. & Cairns, B.R. Nat. Biotechnol. 31, 458–464 (2013).
RNA is methylated at specific cytosine residues. The modification occurs mainly in noncoding species such as tRNA and rRNA and is carried out by methyltransferases that form a transient covalent bond with their target cytosines. Khoddami and Cairns use 5-azacytidine, a cytidine analog that can be incorporated into RNA by living cells, to render the bond permanent, which allows the enzyme and its bound targets to be enriched by immunoprecipitation. 5-Azacytidine also causes C-to-G changes at methylation target sites; as a consequence, targets can be precisely located through sequencing of the enriched RNA. Starting with overexpressed tagged methyltransferases DNMT2 and NSUN2, the researchers enriched known target sites over 200-fold and discovered new targets in human cancer and fibroblast cell lines.
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Targets of RNA cytosine methyltransferases. Nat Methods 10, 463 (2013). https://doi.org/10.1038/nmeth.2492