Antibody-free enzyme-assisted chemical approach for detection of N6-methyladenosine

Nat Chem Biol. 2020 Aug;16(8):896-903. doi: 10.1038/s41589-020-0525-x. Epub 2020 Apr 27.

Abstract

The inert chemical property of RNA modification N6-methyladenosine (m6A) makes it very challenging to detect. Most m6A sequencing methods rely on m6A-antibody immunoprecipitation and cannot distinguish m6A and N6,2'-O-dimethyladenosine modification at the cap +1 position (cap m6Am). Although the two antibody-free methods (m6A-REF-seq/MAZTER-seq and DART-seq) have been developed recently, they are dependent on m6A sequence or cellular transfection. Here, we present an antibody-free, FTO-assisted chemical labeling method termed m6A-SEAL for specific m6A detection. We applied m6A-SEAL to profile m6A landscapes in humans and plants, which displayed the known m6A distribution features in transcriptome. By doing a comparison with all available m6A sequencing methods and specific m6A sites validation by SELECT, we demonstrated that m6A-SEAL has good sensitivity, specificity and reliability for transcriptome-wide detection of m6A. Given its tagging ability and FTO's oxidation property, m6A-SEAL enables many applications such as enrichment, imaging and sequencing to drive future functional studies of m6A and other modifications.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine / analogs & derivatives*
  • Adenosine / analysis
  • Adenosine / chemistry
  • Affinity Labels / chemistry*
  • Alpha-Ketoglutarate-Dependent Dioxygenase FTO / chemistry
  • Alpha-Ketoglutarate-Dependent Dioxygenase FTO / metabolism
  • Humans
  • Immunoprecipitation / methods
  • Methylation
  • RNA / chemistry
  • RNA, Messenger / chemistry
  • RNA, Messenger / metabolism
  • Reproducibility of Results
  • Sensitivity and Specificity
  • Transcriptome

Substances

  • Affinity Labels
  • RNA, Messenger
  • RNA
  • N-methyladenosine
  • Alpha-Ketoglutarate-Dependent Dioxygenase FTO
  • FTO protein, human
  • Adenosine