Human and bacterial oxidative demethylases repair alkylation damage in both RNA and DNA

Abstract

Repair of DNA damage is essential for maintaining genome integrity, and repair deficiencies in mammals are associated with cancer, neurological disease and developmental defects. Alkylation damage in DNA is repaired by at least three different mechanisms, including damage reversal by oxidative demethylation of 1-methyladenine and 3-methylcytosine by Escherichia coli AlkB. By contrast, little is known about consequences and cellular handling of alkylation damage to RNA. Here we show that two human AlkB homologues, hABH2 and hABH3, also are oxidative DNA demethylases and that AlkB and hABH3, but not hABH2, also repair RNA. Whereas AlkB and hABH3 prefer single-stranded nucleic acids, hABH2 acts more efficiently on double-stranded DNA. In addition, AlkB and hABH3 expressed in E. coli reactivate methylated RNA bacteriophage MS2 in vivo, illustrating the biological relevance of this repair activity and establishing RNA repair as a potentially important defence mechanism in living cells. The different catalytic properties and the different subnuclear localization patterns shown by the human homologues indicate that hABH2 and hABH3 have distinct roles in the cellular response to alkylation damage.

NSUN 3 and ABH 1 modify the wobble position of mt‐t RNA Met to expand codon recognition in mitochondrial translation

“…ABH1 can demethylate single‐stranded DNA and RNA in vitro with low efficiency, with a preference for oxidation of N3‐methylcytosine (m 3 C) (Westbye et al , 2008), and has been suggested to act as histone demethylase and abasic site lyase (Müller et al , 2010; Ougland et al , 2012). While the homologous E. coli AlkB cannot oxidise m 5 C in vitro (Li et al , 2010) and human ALKBH2 and ALKBH3 preferentially repair alkylation at nucleobase heteroatoms such as m 3 C and 1‐methyladenosine (m 1 A) (Aas et al , 2003; Falnes et al , 2004), the oxidation of m 5 C involves transformation of a pseudobenzylic methyl group. In DNA, this reaction is catalysed by related Fe(II)/α‐ketoglutarate‐dependent oxygenases of the TET enzyme family, and the oxidation products play a significant role in epigenetic regulation in mammals (Tahiliani et al , 2009; Breiling & Lyko, 2015; Li et al , 2015).…”

Adenine methylation in eukaryotes: Apprehending the complex evolutionary history and functional potential of an epigenetic modification

“…It was also predicted then that certain members of the AlkB family were likely to demethylate m 6 A in RNA 96. Subsequently, such demethylation of m 6 A in RNA was indeed observed to be the mechanism for resetting methyl marks generated by N 6 A RNA MTases 68, 99. Oxidation of the methyl group by these enzymes results in formation of N 6 hmA and N 6 fA, which restore the original base, releasing formaldehyde and formate 100.…”

ALKBH1-Mediated tRNA Demethylation Regulates Translation

“…m 1 A has recently been discovered as another mRNA modification, in addition to m 6 A, pseudouridine, and m 5 C. The presence of m 1 A at the 5′ UTR region in mRNA has been suggested to promote translation (Dominissini et al, 2016;Li et al, 2016). ALKBH3, a DNA repair enzyme that demethylates N 1 -methyldeoxyadenosine in single-stranded DNA (Aas et al, 2003;Duncan et al, 2002) has been suggested as a potential demethylase of m 1 A in mRNA , the functional relevance of which still requires further investigations. ALKBH3 is also capable of repairing tRNA alkylation damages (Aas et al, 2003), again the biological relevance of which was not clear.…”

“…ALKBH3, a DNA repair enzyme that demethylates N 1 -methyldeoxyadenosine in single-stranded DNA (Aas et al, 2003;Duncan et al, 2002) has been suggested as a potential demethylase of m 1 A in mRNA , the functional relevance of which still requires further investigations. ALKBH3 is also capable of repairing tRNA alkylation damages (Aas et al, 2003), again the biological relevance of which was not clear. We show that m 1 A in tRNA is the main substrate of ALKBH1.…”