Gene expression regulation mediated through reversible m6A RNA methylation

Journal name:
Nature Reviews Genetics
Volume:
15,
Pages:
293–306
Year published:
DOI:
doi:10.1038/nrg3724
Published online

Abstract

Cellular RNAs carry diverse chemical modifications that used to be regarded as static and having minor roles in 'fine-tuning' structural and functional properties of RNAs. In this Review, we focus on reversible methylation through the most prevalent mammalian mRNA internal modification, N6-methyladenosine (m6A). Recent studies have discovered protein 'writers', 'erasers' and 'readers' of this RNA chemical mark, as well as its dynamic deposition on mRNA and other types of nuclear RNA. These findings strongly indicate dynamic regulatory roles that are analogous to the well-known reversible epigenetic modifications of DNA and histone proteins. This reversible RNA methylation adds a new dimension to the developing picture of post-transcriptional regulation of gene expression.

At a glance

Figures

  1. Reversible chemical modifications that regulate the flow of genetic information.
    Figure 1: Reversible chemical modifications that regulate the flow of genetic information.

    In the central dogma, genetic information is passed from DNA to RNA and then to protein. Epigenetic DNA modifications (for example, the formation of 5-methylcytosine (m5C; also known as 5mC) and 5-hydroxymethylcytosine (hm5C; also known as 5hmC)) and histone modifications (for example, methylation (me) and acetylation (ac)) are known to have important roles in regulating cell differentiation and development. Reversible RNA modifications (for example, the formation of N6-methyladenosine (m6A) and N6-hydroxymethyladenosine (hm6A)) add an additional layer of dynamic regulation of biological processes.

  2. Profiling of m6A in RNA by m6A RNA immunoprecipitation.
    Figure 2: Profiling of m6A in RNA by m6A RNA immunoprecipitation.

    Antibody-based N6-methyladenosine (m6A) RNA immunoprecipitation has been developed to profile the transcriptome-wide distribution of m6A. a | Isolated mRNA is fragmented to ~100 nucleotides, immunoprecipitated using m6A-specific antibodies, converted to a cDNA library and subjected to high-throughput sequencing. Comparison between the immunoprecipitated sample and the input sample identifies m6A signal peaks. b | Transcriptome-wide profiling of m6A in mRNA revealed that m6A is enriched around stop codons, at 3′ untranslated regions and within long exons. The 5′ cap contains the N6,2′-O-dimethyladenosine (m6Am) modification, which can also be enriched using the m6A-specific antibody. Me, methyl group.

  3. Reversible m6A methylation of mRNA and other types of nuclear RNA.
    Figure 3: Reversible m6A methylation of mRNA and other types of nuclear RNA.

    The N6-methyladenosine (m6A) modification is installed by a hetero complex of two methyltransferases METTL3–METTL14, assisted by Wilms' tumour 1-associating protein (WTAP), and can be demethylated by the α-ketoglutarate (α-KG)-dependent dioxygenases FTO and ALKBH5. a | Saccharomyces cerevisiae inducer of meiosis 4 (Ime4), and human METTL3 and METTL14 contain the S-adenosyl-l-methionine (SAM)-dependent methyltransferase domain for m6A methylation. The (D/E)P(P/A)(W/L) active site and the SAM-binding motif are conserved. b | Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR–CLIP) reveals that the binding sites of METTL14 and METTL3 on mRNA resemble the consensus sequence of m6A in mammalian mRNA. The sequence bound by WTAP moderately overlaps with those bound by METTL14 and METTL3. c | Mammalian FTO and ALKBH5 contain the active site motif HXDXnH (where X denotes any amino acid) for Fe(II) binding, RXXXXXR for both α-KG binding and substrate recognition, and an extra loop that leads to preferential binding of single-stranded over double-stranded nucleic acids68, 121, 122. Relative to Escherichia coli AlkB, mammalian ALKBH5 has an amino-terminal alanine-rich sequence and a potential coiled-coil structure that could be important for its localization. FTO contains an extra carboxy-terminal domain with a novel fold, possibly to engage in additional protein–protein interactions. d | Methylation and demethylation of m6A on RNA are shown. Whereas ALKBH5 catalyses the direct removal of m6A, FTO can oxidize m6A to N6-hydroxymethyladenosine (hm6A) and N6-formyladenosine (f6A) sequentially; hm6A and f6A are moderately stable (with half-lives of ~3 hours under physiological conditions) and can be hydrolysed to adenine.

  4. Functions of the reader (that is, effector) proteins of m6A.
    Figure 4: Functions of the reader (that is, effector) proteins of m6A.

    a | The characterized YTHDF proteins serve as N6-methyladenosine (m6A) 'readers'. Human YTHDF1–3 proteins contain a carboxy-terminal YTH RNA-binding domain and an amino-terminal P/Q/N-rich region. The YTH domain protein is conserved in the fission yeast Schizosaccharomyces pombe and the budding yeast Saccharomyces cerevisiae. b | The m6A modification is enriched in mRNAs with shorter half-lives in general, which supports the proposed main role of m6A in regulating mRNA stability. c | The m6A-specific RNA-binding proteins are engaged in post-transcriptional regulation of gene expression. YTHDF2 regulates the methylation (me)-dependent RNA degradation. Other reader proteins may exist and affect RNA splicing, storage, trafficking and translation. Data in part b courtesy of X. Wang, laboratory of C.H.

  5. RNA methylation could affect various aspects of RNA metabolism and mRNA translation, and regulate protein expression post-transcrptionally.
    Figure 5: RNA methylation could affect various aspects of RNA metabolism and mRNA translation, and regulate protein expression post-transcrptionally.

    Whereas N6-methyladenosine (m6A) methyltransferases and demethylases shape the methylation (me) landscape, the 'reader' proteins bind to the methylated RNA and mediate specific functions. Various cellular processes could be affected by m6A RNA methylation. In the cell nucleus, m6A may affect RNA export, nuclear retention and splicing, possibly through interactions of reader proteins with RNA export, retention and splicing machineries. After RNAs are exported to the cytoplasm, YTHDF2 can bind to the m6A-containing RNAs and direct them to processing bodies (P-bodies) for accelerated mRNA decay. P-bodies can dynamically form stress granules, in which RNAs could be stored and released back to the translating pool. Besides YTHDF2, other m6A reader proteins may bind to m6A-containing RNAs to control their transport and storage, thereby affecting translation. FTO, α-ketoglutarate-dependent dioxygenase FTO; WTAP, Wilms' tumour 1-associating protein.

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Affiliations

  1. Department of Chemistry and Institute for Biophysical Dynamics, The University of Chicago, 929 East 57th Street, Chicago, Illinois 60637, USA.

    • Ye Fu,
    • Dan Dominissini &
    • Chuan He
  2. Cancer Research Center, Chaim Sheba Medical Center, Tel Hashomer 52621, Israel.

    • Dan Dominissini &
    • Gideon Rechavi
  3. Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.

    • Dan Dominissini &
    • Gideon Rechavi

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Author details

  • Ye Fu

    Ye Fu received his B.Sc. in chemistry from Peking University, Beijing, China, in 2007. He then began to work on the chemical biology of nucleic acid modifications in Chuan He's laboratory at the University of Chicago, Illinois, USA. He has focused on the dynamic regulation of RNA modifications by the proteins alkylated DNA repair protein AlkB homologue 8 (ALKBH8) and α-ketoglutarate-dependent dioxygenase FTO, as well as of other DNA modifications. He received his Ph.D. from the University of Chicago in 2012. He then continued as a postdoctoral researcher in Chuan He's laboratory, where he worked on the genomics of nucleic acid modifications. He will be a postdoctoral fellow in Xiaowei Zhuang's laboratory at Harvard University, Cambridge, Massachusetts, USA, starting in 2014.

  • Dan Dominissini

    Dan Dominissini received his B.Med.Sc. from Tel Aviv University, Israel, in 2007. He went on to study RNA post-transcriptional modifications, focusing on adenosine deamination and methylation, in the laboratory of Gideon Rechavi at Tel-Aviv University and obtained his Ph.D. in 2013. He developed the first global N6-methyladenosine (m6A) sequencing method to reveal the principles that govern m6A distribution and identified the first m6A-binding proteins. He is currently a Human Frontier Science Program postdoctoral fellow in the laboratory of Chuan He at the University of Chicago, Illinois, USA.

  • Gideon Rechavi

    Gideon Rechavi is a professor at the Sackler Faculty of Medicine, Tel Aviv University, Israel, and the Head of the Cancer Research Center at the Sheba Medical Center, Tel Hashomer, Israel. He obtained his M.D. from Tel Aviv University in 1981 and his Ph.D. in molecular biology of cancer from the Weizmann Institute of Science, Rehovot, Israel, with David Givol in 1987. He is certified in Paediatrics, Haematology and Paediatric Haemato-Oncology. In 1992, he established the Paediatric Haematology–Oncology and Bone Marrow Transplantation Department at the Sheba Medical Center. He was appointed associate professor in 1990 and promoted to full professor in 1996 at Tel Aviv University. He is the incumbent of the Djerassi chair in Oncology at Tel Aviv University. His research activities focus on RNA editing and methylation, as well as on the role of transposable genetic elements in cancer. He has recently been selected for the Ernest and Bonnie Beutler Research Program of Excellence in Genomic Medicine.

  • Chuan He

    Chuan He is a professor in the Department of Chemistry and Director of the Institute for Biophysical Dynamics at the University of Chicago, Illinois, USA. He is also a joint professor in the Department of Chemical Biology at Peking University, Beijing, China. He obtained his B.S. from the University of Science and Technology of China, Hefei, in 1994. He received his Ph.D. from the Massachusetts Institute of Technology, Cambridge, USA, in chemistry in 2000 with Stephen J. Lippard. After being trained as a Damon-Runyon postdoctoral fellow at Harvard University, Cambridge, Massachusetts, USA, with Gregory L. Verdine from 2000 to 2002, he joined the University of Chicago as an assistant professor; he was promoted to associate professor in 2008 and full professor in 2010. His current research concerns reversible RNA and DNA methylation in biological regulation. He has recently been selected as an Investigator of the Howard Hughes Medical Institute.

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