A fluorescent methylation-switchable probe for highly sensitive analysis of FTO N 6 -methyladenosine demethylase activity in cells

摘要: N 6-Methyladenosine (m6A) is one of the most abundant epigenetic modifications on mRNA. It dynamically regulated by m6A demethylases FTO and ALKBH5, which are currently attracting intense medical interest because their strong association with several human diseases. Despite clinical significance, molecular mechanisms remain unclear, hence there tremendous in developing analytical tools to facilitate functional studies, a longer term view validating therapeutic potentials. To date, no method exists permits analysis m6A-demethylase activity cells. overcome this challenge, herein, we describe first example fluorescent m6A-switchable oligonucleotide probe, enables direct detection demethylase both vitro living The probe provides simple, yet powerful visual tool for highly sensitive activity. Through use m6A-probe, were able achieve real-time imaging single-cell flow cytometry analyses HepG2 We also successfully applied monitor dynamic changes methylation levels during 3T3-L1 pre-adipocyte differentiation. strategy outlined here versatile may, principle, be adapted study range RNA and, more widely, other modifying enzymes. best our knowledge, present represents not only assay monitoring cells, but new sensing dynamics.