Letter | Published:

m6A potentiates Sxl alternative pre-mRNA splicing for robust Drosophila sex determination

Nature volume 540, pages 301304 (08 December 2016) | Download Citation

Abstract

N6-methyladenosine (m6A) is the most common internal modification of eukaryotic messenger RNA (mRNA) and is decoded by YTH domain proteins1,2,3,4,5,6,7. The mammalian mRNA m6A methylosome is a complex of nuclear proteins that includes METTL3 (methyltransferase-like 3), METTL14, WTAP (Wilms tumour 1-associated protein) and KIAA1429. Drosophila has corresponding homologues named Ime4 and KAR4 (Inducer of meiosis 4 and Karyogamy protein 4), and Female-lethal (2)d (Fl(2)d) and Virilizer (Vir)8,9,10,11,12. In Drosophila, fl(2)d and vir are required for sex-dependent regulation of alternative splicing of the sex determination factor Sex lethal (Sxl)13. However, the functions of m6A in introns in the regulation of alternative splicing remain uncertain3. Here we show that m6A is absent in the mRNA of Drosophila lacking Ime4. In contrast to mouse and plant knockout models5,7,14, Drosophila Ime4-null mutants remain viable, though flightless, and show a sex bias towards maleness. This is because m6A is required for female-specific alternative splicing of Sxl, which determines female physiognomy, but also translationally represses male-specific lethal 2 (msl-2) to prevent dosage compensation in females. We further show that the m6A reader protein YT521-B decodes m6A in the sex-specifically spliced intron of Sxl, as its absence phenocopies Ime4 mutants. Loss of m6A also affects alternative splicing of additional genes, predominantly in the 5′ untranslated region, and has global effects on the expression of metabolic genes. The requirement of m6A and its reader YT521-B for female-specific Sxl alternative splicing reveals that this hitherto enigmatic mRNA modification constitutes an ancient and specific mechanism to adjust levels of gene expression.

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Acknowledgements

We thank J. Horabin, N. Perrimon and the Bloomington, Harvard and Kyoto stock centres for fly lines, BacPAc for DNA clones, E. Zaharieva and M. L. Li for help with imaging, W. Arlt and R. Michell for comments on the manuscript, and J.-Y. Roignant for communication of results before publication. We acknowledge funding from the BBSRC (BB/M008606/1) to R.F.

Author information

Author notes

    • Zsuzsanna Bodi
    • , Eugenio Sanchez-Moran
    •  & Nigel P. Mongan

    These authors contributed equally to this work.

Affiliations

  1. School of Biosciences, College of Life and Environmental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK

    • Irmgard U. Haussmann
    • , Eugenio Sanchez-Moran
    •  & Matthias Soller
  2. School of Life Science, Faculty of Health and Life Sciences, Coventry University, Coventry CV1 5FB, UK

    • Irmgard U. Haussmann
  3. School of Biosciences, Plant Science Division, University of Nottingham, Sutton Bonington, Loughborough LE12 5RD, UK

    • Zsuzsanna Bodi
    • , Nathan Archer
    •  & Rupert G. Fray
  4. School of Veterinary Medicine and Sciences, University of Nottingham, Sutton Bonington, Loughborough LE12 5RD, UK

    • Nigel P. Mongan

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Contributions

I.U.H. and M.S. performed biochemistry, cell biology and genetic experiments, E.S.M. stained chromosomes, and Z.B., N.A. and R.F. performed biochemistry experiments. N.M. analysed sequencing data. I.U.H., R.F. and M.S. conceived the project and wrote the manuscript with help from N.M. and Z.B.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Matthias Soller.

Extended data

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    This file contains graphical source data, uncropped gels, Western blots and 1D TLCs.

Excel files

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    Supplementary Table 1

    This file contains the alternative splicing analysis.

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    Supplementary Table 2

    This file contains the differential gene expression analysis.

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DOI

https://doi.org/10.1038/nature20577

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