Repair of alkylation damage in DNA is clearly important, as at least three repair mechanisms have been described. On the other hand, little is known about what happens to alkylated RNA. However, Hans Krokan and colleagues now report in Nature that damaged RNA can be repaired in vivo — by some of the same enzymes used in DNA repair.

One of the DNA-repair systems involves the oxidative demethylation of 1-methyladenine and 3-methylcytosine. This is catalysed in Escherichia coli by the enzyme AlkB. Krokan and co-workers identified two human AlkB homologues — ABH2 and ABH3. To test the enzymatic activity of these new enzymes, they did high-performance liquid chromatography (HPLC) analysis of [3H]methylated DNA incubated with recombinant ABH2 and ABH3. And indeed, like AlkB, these enzymes were oxidative DNA demethylases capable of removing 1-methyladenine and 3-methylcytosine from DNA by demethylation.

When determining the substrate specificity of ABH2 and ABH3, Krokan and colleagues found that ABH2 was more active on double-stranded (ds)DNA, whereas ABH3 — like AlkB — had a preference for single-stranded (ss)DNA. Remarkably, AlkB and ABH3, but not ABH2, also repaired RNA oligonucleotides efficiently.

Next, the authors tested whether this newly discovered RNA-repair activity was biologically relevant by measuring the survival of chemically methylated (by methyl methanesulphonate; MMS) bacteriophage in the presence of plasmid-expressed AlkB, ABH2 or ABH3 in an AlkB-deficient E. coli strain. Consistent with the in vitro data, AlkB and ABH3, but not ABH2, were able to reactivate MMS-treated ssRNA phage MS2. Methylated ssDNA phage M13 could be reactivated by ABH2 and AlkB, and only partially by ABH3, whereas AlkB and ABH2, but not ABH3, reactivated dsDNA phage λ.

By studying the subcellular localization of ABH2 and ABH3, Krokan and co-workers noticed that ABH2 colocalizes with replication foci during S phase, which is consistent with its higher activity on dsDNA. By contrast, the dispersed nuclear localization of ABH3 throughout the cell cycle, and its preference for ssDNA and RNA, indicates that it might have a maintenance role in actively transcribing genes.

So, given that about 20 of the 62 known naturally occurring RNA modifications are sequence-specific methylation events, Krokan and colleagues conclude “...it is not surprising that cells have evolved RNA repair mechanisms that operate to prevent dysfunction of RNA as a result of alkylation damage”.