Topology of the human and mouse m6A RNA methylomes revealed by m6A-seq

Abstract

An extensive repertoire of modifications is known to underlie the versatile coding, structural and catalytic functions of RNA, but it remains largely uncharted territory. Although biochemical studies indicate that N6-methyladenosine (m6A) is the most prevalent internal modification in messenger RNA, an in-depth study of its distribution and functions has been impeded by a lack of robust analytical methods. Here we present the human and mouse m6A modification landscape in a transcriptome-wide manner, using a novel approach, m6A-seq, based on antibody-mediated capture and massively parallel sequencing. We identify over 12,000 m6A sites characterized by a typical consensus in the transcripts of more than 7,000 human genes. Sites preferentially appear in two distinct landmarks—around stop codons and within long internal exons—and are highly conserved between human and mouse. Although most sites are well preserved across normal and cancerous tissues and in response to various stimuli, a subset of stimulus-dependent, dynamically modulated sites is identified. Silencing the m6A methyltransferase significantly affects gene expression and alternative splicing patterns, resulting in modulation of the p53 (also known as TP53) signalling pathway and apoptosis. Our findings therefore suggest that RNA decoration by m6A has a fundamental role in regulation of gene expression.

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Figure 1: m 6 A-seq capture of modified RNA fragments exposes an enriched motif.
Figure 2: The transcriptome landscape of m 6 A reveals a unique topology.
Figure 3: m 6 A methylome conservation between human and mouse.
Figure 4: Transcripts differing in methylation patterns under varying growing conditions.

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Primary accessions

Gene Expression Omnibus

Data deposits

Data have been deposited in NCBI’s Gene Expression Omnibus (GEO) and are accessible through GEO series accession number GSE37005 (http://ncbi.nlm.nih.gov/geo/query/acc.cgi?acc5GSE37005).

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Acknowledgements

We thank H. Cedar (The Hebrew University, Jerusalem) for his comments. We thank the Kahn Family Foundation for their support. This work was supported in part by grants from the Flight Attendant Medical Research Institute (FAMRI), Bio-Med Morasha ISF (grant no. 1942/08), and The Israel Ministry for Science and Technology (Scientific Infrastructure Program). R.S. was supported by the ERC-StG program (grant 260432). G.R. holds the Djerassi Chair in Oncology at the Sackler Faculty of Medicine, Tel Aviv University. This work was performed in partial fulfilment of the requirements for a PhD degree to D.D., Sackler Faculty of Medicine, Tel Aviv University.

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Contributions

D.D., S.M.-M. and G.R. conceived and designed the experiments; D.D., S.M.-M., L.U., K.C., S.O. and J.J.-H. performed the experiments; S.S., M.S.-D. and R.S. performed the bioinformatic analysis; D.D., S.M.-M., M.S.-D., N.A., M.K., S.S., R.S. and G.R. analysed and interpreted results, and wrote the paper.

Corresponding author

Correspondence to Gideon Rechavi.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-18, Supplementary References, Supplementary Tables 1-5, Supplementary Notes and legends for Supplementary Tables 6-8. (PDF 5060 kb)

Supplementary Table 6

This file contains Supplementary Table 6 in USCS format. (TXT 2738 kb)

Supplementary Table 6

This file contains Supplementary Table 6, which shows the dataset of all identified m6A peaks in HepG2 cell line and normal human brain. (XLS 21053 kb)

Supplementary Table 7

This file contains Supplementary Table 7 in UCSC format. (TXT 224 kb)

Supplementary Table 7

This file contains Supplementary Table 7, which shows the dataset of all identified m6A peaks in mouse liver. (XLS 8054 kb)

Supplementary Table 8

This file contains Supplementary Table 8, which shows differential m6A peaks between various experimental conditions. (XLS 242 kb)

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Dominissini, D., Moshitch-Moshkovitz, S., Schwartz, S. et al. Topology of the human and mouse m6A RNA methylomes revealed by m6A-seq. Nature 485, 201–206 (2012). https://doi.org/10.1038/nature11112

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