Anti-ALKBH2 (aa 108-157) polyclonal antibody

Environmental and endogenous genotoxic agents can result in a variety of alkylated and carboxymethylated DNA lesions, including N3-ethylthymidine (N3-EtdT), O-2-EtdT, and O-4-EtdT as well as N3-carboxymethylthymidine (N3-CMdT) and O-4-CMdT. By using nonreplicative double-stranded vectors harboring a site-specifically incorporated DNA lesion, we assessed the potential roles of alkyladenine DNA glycosylase (Aag); alkylation repair protein B homologue 2 (Alkbh2); or Alkbh3 in modulating the effects of N3-EtdT, O-2-EtdT, O-4-EtdT, N3-CMdT, or O-4-CMdT on DNA transcription in mammalian cells. We found that the depletion of Aag did not significantly change the transcriptional inhibitory or mutagenic properties of all five examined lesions, suggesting a negligible role of Aag in the repair of these DNA adducts in mammalian cells. In addition, our results revealed that N3-EtdT, but not other lesions, could be repaired by Alkbh2 and Alkbh3 in mammalian cells. Furthermore, we demonstrated the direct reversal of N3-EtdT by purified human Alkbh2 protein in vitro. These findings provided important new insights into the repair of the carboxymethylated and alkylated thymidine lesions in mammalian cells.

ALKB-8 is a 2-oxoglutarate-dependent dioxygenase homologous to bacterial AlkB, which oxidatively demethylates DNA substrates. The mammalian AlkB family contains AlkB homologues denominated ALKBH1 to 8 and FTO. The C. elegans genome includes five AlkB-related genes, homologues of ALKBH1, 4, 6, 7, and 8, but lacks homologues of ALKBH2, 3, and 5 and FTO. ALKBH8 orthologues differ from other AlkB family members by possessing an additional methyltransferase module and an RNA binding N-terminal module. The ALKBH8 methyltransferase domain generates the wobble nucleoside 5-methoxycarbonylmethyluridine from its precursor 5-carboxymethyluridine and its (R)- and (S)-5-methoxycarbonylhydroxymethyluridine hydroxylated forms in tRNA(UCG)(Arg) and tRNA(UCC)(Gly). The ALKBH8/ALKB-8 methyltransferase domain is highly similar to yeast TRM9, which selectively modulates translation of mRNAs enriched with AGA and GAA codons under both normal and stress conditions. In this report, we studied the role of alkb-8 in C. elegans. We show that downregulation of alkb-8 increases detection of lysosome-related organelles visualized by Nile red in vivo. Reversely, forced expression of alkb-8 strongly decreases the detection of this compartment. In addition, overexpression of alkb-8 applied in a pulse during the L1 larval stage increases the C. elegans lifespan.