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The Shine-Dalgarno sequence of riboswitch-regulated single mRNAs shows ligand-dependent accessibility bursts

Author

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  • Arlie J. Rinaldi

    (Single Molecule Analysis Group, University of Michigan
    Present address: W.M. Keck Science Center, The Claremont Colleges, Claremont, California 91711, USA)

  • Paul E. Lund

    (Single Molecule Analysis Group, University of Michigan
    Program in Chemical Biology, University of Michigan)

  • Mario R. Blanco

    (Single Molecule Analysis Group, University of Michigan
    Present address: Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, California 91125, USA)

  • Nils G. Walter

    (Single Molecule Analysis Group, University of Michigan)

Abstract

In response to intracellular signals in Gram-negative bacteria, translational riboswitches—commonly embedded in messenger RNAs (mRNAs)—regulate gene expression through inhibition of translation initiation. It is generally thought that this regulation originates from occlusion of the Shine-Dalgarno (SD) sequence upon ligand binding; however, little direct evidence exists. Here we develop Single Molecule Kinetic Analysis of RNA Transient Structure (SiM-KARTS) to investigate the ligand-dependent accessibility of the SD sequence of an mRNA hosting the 7-aminomethyl-7-deazaguanine (preQ1)-sensing riboswitch. Spike train analysis reveals that individual mRNA molecules alternate between two conformational states, distinguished by ‘bursts’ of probe binding associated with increased SD sequence accessibility. Addition of preQ1 decreases the lifetime of the SD’s high-accessibility (bursting) state and prolongs the time between bursts. In addition, ligand-jump experiments reveal imperfect riboswitching of single mRNA molecules. Such complex ligand sensing by individual mRNA molecules rationalizes the nuanced ligand response observed during bulk mRNA translation.

Suggested Citation

  • Arlie J. Rinaldi & Paul E. Lund & Mario R. Blanco & Nils G. Walter, 2016. "The Shine-Dalgarno sequence of riboswitch-regulated single mRNAs shows ligand-dependent accessibility bursts," Nature Communications, Nature, vol. 7(1), pages 1-10, April.
    • Handle: RePEc:nat:natcom:v:7:y:2016:i:1:d:10.1038_ncomms9976
    DOI: 10.1038/ncomms9976
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    Cited by:

    1. Yanyan Xue & Jun Li & Dian Chen & Xizhu Zhao & Liang Hong & Yu Liu, 2023. "Observation of structural switch in nascent SAM-VI riboswitch during transcription at single-nucleotide and single-molecule resolution," Nature Communications, Nature, vol. 14(1), pages 1-14, December.
    2. Adrien Chauvier & Shiba S. Dandpat & Rosa Romero & Nils G. Walter, 2024. "A nascent riboswitch helix orchestrates robust transcriptional regulation through signal integration," Nature Communications, Nature, vol. 15(1), pages 1-18, December.

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