Transcriptome-wide profiling and quantification of N⁶-methyladenosine by enzyme-assisted adenosine deamination.

ABSTRACT: N⁶-methyladenosine (m⁶A), the most abundant internal messenger RNA modification in higher eukaryotes, serves myriad roles in regulating cellular processes. Functional dissection of m⁶A is, however, hampered in part by the lack of high-resolution and quantitative detection methods. Here we present evolved TadA-assisted N⁶-methyladenosine sequencing (eTAM-seq), an enzyme-assisted sequencing technology that detects and quantifies m⁶A by global adenosine deamination. With eTAM-seq, we analyze the transcriptome-wide distribution of m⁶A in HeLa and mouse embryonic stem cells. The enzymatic deamination route employed by eTAM-seq preserves RNA integrity, facilitating m⁶A detection from limited input samples. In addition to transcriptome-wide m⁶A profiling, we demonstrate site-specific, deep-sequencing-free m⁶A quantification with as few as ten cells, an input demand orders of magnitude lower than existing quantitative profiling methods. We envision that eTAM-seq will enable researchers to not only survey the m⁶A landscape at unprecedented resolution, but also detect m⁶A at user-specified loci with a simple workflow.

SUBMITTER: Xiao YL 

PROVIDER: S-EPMC10625715 | biostudies-literature | 2023 Jul

REPOSITORIES: biostudies-literature