An RNA Methylation-Sensitive AIEgen-Aptamer Reporting System for Quantitatively Evaluating m 6 A Methylase and Demethylase Activities.

N 6 -Methyladenosine (m 6 A) chemical modification determines the fate of the mammalian cellular mRNA to modulate crucial physiological and pathological processes. Dysregulations of m 6 A methylase and demethylase have been linked to cancer diseases. Therefore, evaluations of enzyme mutants' activities and related inhibitors for discovery of targeted therapeutic strategies are very necessary. Here, we report an RNA methylation-sensitive fluorescent aptamer reporting assay to measure the catalytic activities of m 6 A enzymes under various conditions. The rationale is that when an RNA aptamer, named A-Pepper, is methylated at a specific adenosine position to generate m 6 A-Pepper, the latter displays stronger fluorescence than the former upon binding the ligand, which is an aggregation-induced emission-active luminogen. The fluorescence signal enhancement is linearly proportional to the RNA methylation extent, which is equivalent to the methylase activity. On the contrary, the m 6 A demethylase activity is measured through calculating the fluorescence signal decrease caused by the switching from m 6 A-Pepper to A-Pepper. The assay has been successfully applied to quantitatively evaluate the mutation and inhibitor effects on the activities of m 6 A methylases METTL3/METTL14 and demethylase FTO, and the obtained results are well-consistent with those quantified by the expensive and time-consuming golden standard LC-MS/MS. Our work provides a simple tool capable of detecting m 6 A enzymes' activities and screening their inhibitors in a rapid, quantitative, cost-effective, and high-throughput manner.