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Structural basis of a small molecule targeting RNA for a specific splicing correction

Nature Chemical Biology volume 15, pages 1191–1198 (2019)Cite this article

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Abstract

Splicing modifiers promoting SMN2 exon 7 inclusion have the potential to treat spinal muscular atrophy, the leading genetic cause of infantile death. These small molecules are SMN2 exon 7 selective and act during the early stages of spliceosome assembly. Here, we show at atomic resolution how the drug selectively promotes the recognition of the weak 5ʹ splice site of SMN2 exon 7 by U1 snRNP. The solution structure of the RNA duplex formed following 5ʹ splice site recognition in the presence of the splicing modifier revealed that the drug specifically stabilizes a bulged adenine at this exon–intron junction and converts the weak 5ʹ splice site of SMN2 exon 7 into a stronger one. The small molecule acts as a specific splicing enhancer cooperatively with the splicing regulatory network. Our investigations uncovered a novel concept for gene-specific alternative splicing correction that we coined 5ʹ splice site bulge repair.

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Fig. 1: SMN-C5 recognizes the interface between U1 snRNP and the SMN2 E7 5ʹ SS.
Fig. 2: Structural basis for SMN-C5 selectivity.
Fig. 3: Validation of the SMN-C5 mode of action in cellular models.
Fig. 4: Positive cooperativity between SMN-C5 and the splicing regulatory network.
Fig. 5: The concept of 5ʹ-SS bulge repair.

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Data availability

The chemical shifts and the atomic coordinates of the RNA duplex with or without SMN-C5 have been deposited in the Biological Magnetic Resonance Bank (BMRB ID: 34311 and 34312) and in the Protein Data Bank (PDB ID: 6HMI and 6HMO). Other data and materials are available from the authors upon reasonable request.

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Acknowledgements

We thank F. Tessaro, A. Gossert and M. Krepl for their assistance with the molecular dynamics setup, K. McCarthy and F. Metzger for useful discussions and the RNA synthesis platform of the NCCR RNA and Diseases. This work was supported by the Swiss National Science Foundation, the NCCR RNA and Diseases, cure-SMA and SMA-Europe.

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Authors and Affiliations

  1. Department of Biology, Institute of Molecular Biology and Biophysics, ETH Zurich, Zurich, Switzerland

    Sébastien Campagne, Sarah Boigner, Simon Rüdisser, Ahmed Moursy, Laurent Gillioz, Antoine Cléry & Frédéric H.-T. Allain

  2. Biomolecular NMR Spectroscopy Platform, ETH Zurich, Zurich, Switzerland

    Simon Rüdisser

  3. Department of Chemistry and Applied Biosciences, Institute of Pharmaceutical Sciences, ETH Zurich, Zurich, Switzerland

    Anna Knörlein & Jonathan Hall

  4. F. Hoffmann-La Roche Ltd, Pharma Research & Early Development, Roche Innovation Center, Basel, Switzerland

    Hasane Ratni

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  1. Sébastien Campagne
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Contributions

S.C., A.C. and F.H.-T.A. designed the research. S.C. and L.G. performed gel shift assays. S.C., S.B. and A.M. performed the splicing assays in cellular models. S.C. and S.R. performed NMR data collection. A.K. and J.H. synthesized the isotopically labeled RNA. H.R. furnished the splicing modifiers. S.C. analyzed the data and solved the structures. S.C. and F.H.-T.A. wrote the manuscript.

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Correspondence to Sébastien Campagne or Frédéric H.-T. Allain.

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H.R. is an employee of F. Hoffmann-La Roche Ltd.

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Supplementary information

Supplementary Information

Supplementary Table 1 and Supplementary Figs 1–10.

Reporting Summary

Supplementary Video 1

Solution structure of the apo-RNA duplex.

Supplementary Video 2

Solution structure of the RNA duplex bound to SMN-C5.

Supplementary Video 3

RNA conformational switch induced by SMN-C5—major groove view.

Supplementary Video 4

RNA conformational switch induced by SMN-C5—minor groove view.

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Campagne, S., Boigner, S., Rüdisser, S. et al. Structural basis of a small molecule targeting RNA for a specific splicing correction. Nat Chem Biol 15, 1191–1198 (2019). https://doi.org/10.1038/s41589-019-0384-5

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