Article | Published:

RNA catalyses nuclear pre-mRNA splicing

Nature volume 503, pages 229234 (14 November 2013) | Download Citation

Abstract

In nuclear pre-messenger RNA splicing, introns are excised by the spliceosome, a dynamic machine composed of both proteins and small nuclear RNAs (snRNAs). Over thirty years ago, after the discovery of self-splicing group II intron RNAs, the snRNAs were proposed to catalyse splicing. However, no definitive evidence for a role of either RNA or protein in catalysis by the spliceosome has been reported so far. By using metal rescue strategies in spliceosomes from budding yeast, here we show that the U6 snRNA catalyses both of the two splicing reactions by positioning divalent metals that stabilize the leaving groups during each reaction. Notably, all of the U6 catalytic metal ligands we identified correspond to the ligands observed to position catalytic, divalent metals in crystal structures of a group II intron RNA. These findings indicate that group II introns and the spliceosome share common catalytic mechanisms and probably common evolutionary origins. Our results demonstrate that RNA mediates catalysis within the spliceosome.

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Acknowledgements

We thank C. Guthrie for plasmids; S. -C. Cheng for anti-Cwc25p serum; D. Semlow for strains; J. Olvera for reagents and experimental assistance; R. -J. Lin for sharing unpublished data; members of the Staley and Piccirilli laboratories for discussions; and D. Herschlag, A. Macmillan and T. Nilsen for comments on the manuscript. N.T. was supported by an NSF Graduate Research Fellowship and by a CBI Training Grant (5T32GM008720). This work was funded by a grant from the Chicago Biomedical Consortium, with support from The Searle Funds at the Chicago Community Trust, to J.P.S, A. S. Mankin and E. J. Sontheimer, and by a grant from the National Institutes of Health (R01GM088656) to J.P.S. and J.A.P.

Author information

Author notes

    • Sebastian M. Fica
    •  & Nicole Tuttle

    These authors contributed equally to this work.

Affiliations

  1. Graduate Program in Cell and Molecular Biology, The University of Chicago, Chicago, Illinois 60637, USA

    • Sebastian M. Fica
  2. Department of Molecular Genetics and Cell Biology, Cummings Life Sciences Center, 920 East 58th Street, The University of Chicago, Chicago, Illinois 60637, USA

    • Sebastian M. Fica
    • , Prakash Koodathingal
    •  & Jonathan P. Staley
  3. Department of Chemistry, 929 East 57th Street, The University of Chicago, Chicago, Illinois 60637, USA

    • Nicole Tuttle
    • , Jun Lu
    • , Qing Dai
    •  & Joseph A. Piccirilli
  4. Department of Biochemistry and Molecular Biology, Gordon Center for Integrative Science, 929 East 57th Street, The University of Chicago, Chicago, Illinois 60637, USA

    • Thaddeus Novak
    • , Nan-Sheng Li
    •  & Joseph A. Piccirilli

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Contributions

S.M.F., N.T., T.N., J.P.S. and J.A.P. designed the study; T.N. and P.K. performed initial screening of U6 sulphur substitutions; S.M.F. performed all experiments related to branching; N.T. performed all experiments related to exon ligation; S.M.F. and N.T. together performed Prp8p experiments; J.L., N.-S.L. and Q.D. synthesized RNA oligonucleotides; S.M.F., N.T., J.P.S. and J.A.P. analysed the data and wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Jonathan P. Staley or Joseph A. Piccirilli.

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https://doi.org/10.1038/nature12734

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