Splice site m6A methylation prevents binding of U2AF35 to inhibit RNA splicing

Cell. 2021 Jun 10;184(12):3125-3142.e25. doi: 10.1016/j.cell.2021.03.062. Epub 2021 Apr 29.


The N6-methyladenosine (m6A) RNA modification is used widely to alter the fate of mRNAs. Here we demonstrate that the C. elegans writer METT-10 (the ortholog of mouse METTL16) deposits an m6A mark on the 3' splice site (AG) of the S-adenosylmethionine (SAM) synthetase pre-mRNA, which inhibits its proper splicing and protein production. The mechanism is triggered by a rich diet and acts as an m6A-mediated switch to stop SAM production and regulate its homeostasis. Although the mammalian SAM synthetase pre-mRNA is not regulated via this mechanism, we show that splicing inhibition by 3' splice site m6A is conserved in mammals. The modification functions by physically preventing the essential splicing factor U2AF35 from recognizing the 3' splice site. We propose that use of splice-site m6A is an ancient mechanism for splicing regulation.

Keywords: 3' splice site; METT-10; METTL16; SAM homeostasis; SAM synthetase; U2AF35/65; U6 snRNA; m(6)A; spermatogenesis; splicing.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine / analogs & derivatives*
  • Adenosine / metabolism
  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Caenorhabditis elegans / genetics
  • Conserved Sequence / genetics
  • Diet
  • HeLa Cells
  • Humans
  • Introns / genetics
  • Methionine Adenosyltransferase
  • Methylation
  • Methyltransferases / chemistry
  • Mice
  • Mutation / genetics
  • Nucleic Acid Conformation
  • Protein Binding
  • RNA Precursors / chemistry
  • RNA Precursors / genetics
  • RNA Precursors / metabolism
  • RNA Splice Sites / genetics*
  • RNA Splicing / genetics*
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • RNA, Small Nuclear
  • S-Adenosylmethionine
  • Splicing Factor U2AF / metabolism*
  • Transcriptome / genetics


  • RNA Precursors
  • RNA Splice Sites
  • RNA, Messenger
  • RNA, Small Nuclear
  • Splicing Factor U2AF
  • U2AF1 protein, human
  • U6 small nuclear RNA
  • S-Adenosylmethionine
  • N-methyladenosine
  • METTL16 protein, human
  • Methyltransferases
  • Methionine Adenosyltransferase
  • Adenosine