N(6)-methyladenosine (m(6)A) is the most prevalent internal (non-cap) modification present in the messenger RNA of all higher eukaryotes. Although essential to cell viability and development, the exact role of m(6)A modification remains to be determined. The recent discovery of two m(6)A demethylases in mammalian cells highlighted the importance of m(6)A in basic biological functions and disease. Here we show that m(6)A is selectively recognized by the human YTH domain family 2 (YTHDF2) 'reader' protein to regulate mRNA degradation. We identified over 3,000 cellular RNA targets of YTHDF2, most of which are mRNAs, but which also include non-coding RNAs, with a conserved core motif of G(m(6)A)C. We further establish the role of YTHDF2 in RNA metabolism, showing that binding of YTHDF2 results in the localization of bound mRNA from the translatable pool to mRNA decay sites, such as processing bodies. The carboxy-terminal domain of YTHDF2 selectively binds to m(6)A-containing mRNA, whereas the amino-terminal domain is responsible for the localization of the YTHDF2-mRNA complex to cellular RNA decay sites. Our results indicate that the dynamic m(6)A modification is recognized by selectively binding proteins to affect the translation status and lifetime of mRNA.

/pdf/n6-methyladenosine-dependent-regulation-of-messenger-rna-2payhjmq6g.pdf

N6-methyladenosine-dependent regulation of messenger RNA stability

https://www.cell.com/cell/pdf/S0092-8674(15)00562-0.pdf

N6-methyladenosine Modulates Messenger RNA Translation Efficiency

Certain adenosine residues within mammalian RNAs undergo reversible N6 methylation. Two methyltransferase enzymes, METTL3 and METTL14, as well as the splicing factor WTAP are identified as core components of the multiprotein complex that deposits RNA N6-methyladenosine (m6A) in nuclear RNAs.

/pdf/a-mettl3-mettl14-complex-mediates-mammalian-nuclear-rna-n6-3zjwfen4xs.pdf

A METTL3-METTL14 complex mediates mammalian nuclear RNA N6-adenosine methylation

Dynamic RNA Modifications in Gene Expression Regulation

The methyltransferase like 3 (METTL3)-containing methyltransferase complex catalyzes the N6-methyladenosine (m6A) formation, a novel epitranscriptomic marker; however, the nature of this complex remains largely unknown. Here we report two new components of the human m6A methyltransferase complex, Wilms' tumor 1-associating protein (WTAP) and methyltransferase like 14 (METTL14). WTAP interacts with METTL3 and METTL14, and is required for their localization into nuclear speckles enriched with pre-mRNA processing factors and for catalytic activity of the m6A methyltransferase in vivo. The majority of RNAs bound by WTAP and METTL3 in vivo represent mRNAs containing the consensus m6A motif. In the absence of WTAP, the RNA-binding capability of METTL3 is strongly reduced, suggesting that WTAP may function to regulate recruitment of the m6A methyltransferase complex to mRNA targets. Furthermore, transcriptomic analyses in combination with photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) illustrate that WTAP and METTL3 regulate expression and alternative splicing of genes involved in transcription and RNA processing. Morpholino-mediated knockdown targeting WTAP and/or METTL3 in zebrafish embryos caused tissue differentiation defects and increased apoptosis. These findings provide strong evidence that WTAP may function as a regulatory subunit in the m6A methyltransferase complex and play a critical role in epitranscriptomic regulation of RNA metabolism.

/pdf/mammalian-wtap-is-a-regulatory-subunit-of-the-rna-n6-19zlrd1d5c.pdf

Florian Bogdan

Papers

ALKBH5 Is a Mammalian RNA Demethylase that Impacts RNA Metabolism and Mouse Fertility