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The formation of m6A RNA appears to be a co-transcriptional event taking place early on in the RNA lifecycle and is mediated by a multi-protein methyltransferase complex composed, in part, of METTL enzymes. Chemical addition of a methyl group at the nitrogen-6 position of adenosine (A) residues to form m6A occurs via the METTL3/14 heterodimer. Through FTO-mediated oxidative demethylation, m6A is converted in a step-wise manner to hm6A and subsequently f6A before finally reverting back to A. The recent discoveries of these m6A methylase “writers” and their associated demethylase “erasers” in mammals uncovered the reversibility of the m6A modification, exposing potential therapeutic targets for m6A dysregulation-related diseases.
EpigenTek’s ELISA-based Epigenase assays provide rapid measures of m6A methylase and demethylase activity levels from cell/tissue nuclear extracts or purified enzymes in a high-throughput format. A unique m6A substrate is stably coated on the assay wells. Bioactive enzymes from input samples will transfer methyl groups to (methylases), or remove them from (demethylases), the bound substrate. Substrate m6A methylation is subsequently detected by a high-affinity antibody specific for this modification. Quantitative measurements of methylase/demethylase activity and inhibition (for enzyme inhibitor screening) can be quickly obtained within only a few hours.
