Control of alternative RNA splicing and gene expression by eukaryotic riboswitches

Abstract

Bacteria make extensive use of riboswitches1,2 to sense metabolites and control gene expression, and typically do so by modulating premature transcription termination or translation initiation. The most widespread riboswitch class known in bacteria responds to the coenzyme thiamine pyrophosphate (TPP)3,4, which is a derivative of vitamin B1. Representatives of this class have also been identified5,6 in fungi and plants, where they are predicted5,7 to control messenger RNA splicing or processing. We examined three TPP riboswitches in the filamentous fungus Neurospora crassa, and found that one activates and two repress gene expression by controlling mRNA splicing. A detailed mechanism involving riboswitch-mediated base-pairing changes and alternative splicing control was elucidated for precursor NMT1 mRNAs, which code for a protein involved in TPP metabolism. These results demonstrate that eukaryotic cells employ metabolite-binding RNAs to regulate RNA splicing events that are important for the control of key biochemical processes.

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Figure 1: Three N. crassa genes carry TPP riboswitches in 5′ introns.
Figure 2: Alternative splicing and gene control by the NMT1 TPP riboswitch.
Figure 3: Short uORFs in unspliced and alternatively spliced mRNAs cause NMT1 repression.
Figure 4: Mechanism of TPP riboswitch-mediated alternative splicing of mRNA in N. crassa.

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Acknowledgements

. We thank J. C. Dunlap for hosting and training M.T.C. in fungal genetics techniques and for supplying us with N. crassa strains, genomic DNA of N. crassa and the luciferase reporter vector. Also, we thank members of the Dunlap laboratory (M. Shi and L. Larrondo) for technical assistance and helpful discussions. A.W. was supported by a postdoctoral fellowship from the German Research Foundation (DFG) and M.T.C. was supported by a Yale College Dean’s Fellowship and an HHMI Future Scientist Fellowship. This work also was supported by an NIH grant (R.R.B).

Author Contributions M.T.C. and A.W. conducted all genetic and molecular biology analyses, and M.T.C. and N.S. conducted biochemical analyses. M.T.C., A.W. and R.R.B. co-wrote the manuscript. All authors contributed to discussions regarding the data and their interpretation.

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Correspondence to Ronald R. Breaker.

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[Competing Interests Statement: ‘R.R.B. is a cofounder of BioRelix, a company interested in using riboswitches as antimicrobial drug targets.’]

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This file contains Supplementary Methods, Supplementary Figures S1-S10 with Legends and additional references. (PDF 1154 kb)

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Cheah, M., Wachter, A., Sudarsan, N. et al. Control of alternative RNA splicing and gene expression by eukaryotic riboswitches. Nature 447, 497–500 (2007). https://doi.org/10.1038/nature05769

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