Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

A riboswitch selective for the queuosine precursor preQ1 contains an unusually small aptamer domain

Nature Structural & Molecular Biology volume 14, pages 308–317 (2007)Cite this article

Abstract

A previous bioinformatics-based search for riboswitches yielded several candidate motifs in eubacteria. One of these motifs commonly resides in the 5′ untranslated regions of genes involved in the biosynthesis of queuosine (Q), a hypermodified nucleoside occupying the anticodon wobble position of certain transfer RNAs. Here we show that this structured RNA is part of a riboswitch selective for 7-aminomethyl-7-deazaguanine (preQ1), an intermediate in queuosine biosynthesis. Compared with other natural metabolite-binding RNAs, the preQ1 aptamer appears to have a simple structure, consisting of a single stem-loop and a short tail sequence that together are formed from as few as 34 nucleotides. Despite its small size, this aptamer is highly selective for its cognate ligand in vitro and has an affinity for preQ1 in the low nanomolar range. Relatively compact RNA structures can therefore serve effectively as metabolite receptors to regulate gene expression.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Queuosine biosynthesis in eubacteria.
Figure 2: The 5′ UTR of the B. subtilis queCDEF mRNA undergoes structural modulation in response to preQ1.
Figure 3: Molecular discrimination by the B. subtilis queC 5′ UTR preQ1-binding region.
Figure 4: Determination of the minimal aptamer sequence required for binding of preQ1.
Figure 5: Evidence for a Watson-Crick pairing interaction between preQ1 and a conserved cytidyl residue of the aptamer.
Figure 6: Effects of variant preQ1 riboswitches on genetic control in vivo.

Similar content being viewed by others

Accession codes

Accessions

GenBank/EMBL/DDBJ

References

  1. Ptashne, M. & Gann, A. Genes and Signals (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, USA, 2002).

    Google Scholar 

  2. Mandal, M. & Breaker, R.R. Gene regulation by riboswitches. Nat. Rev. Mol. Cell Biol. 5, 451–463 (2004).

    Article  CAS  Google Scholar 

  3. Soukup, J.K. & Soukup, G.A. Riboswitches exert genetic control through metabolite-induced conformational change. Curr. Opin. Struct. Biol. 14, 344–349 (2004).

    Article  CAS  Google Scholar 

  4. Winkler, W.C. Riboswitches and the role of noncoding RNAs in bacterial metabolic control. Curr. Opin. Chem. Biol. 9, 594–602 (2005).

    Article  CAS  Google Scholar 

  5. Winkler, W.C. & Breaker, R.R. Regulation of bacterial gene expression by riboswitches. Annu. Rev. Microbiol. 59, 487–517 (2005).

    Article  CAS  Google Scholar 

  6. Breaker, R.R. in The RNA World 3rd edn. (eds. Gesteland, R.F., Cech, T.R. & Atkins, J.F.) 89–107 (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, USA, 2006).

    Google Scholar 

  7. Barrick, J.E. et al. New RNA motifs suggest an expanded scope for riboswitches in bacterial genetic control. Proc. Natl. Acad. Sci. USA 101, 6421–6426 (2004).

    Article  CAS  Google Scholar 

  8. Corbino, K.A. et al. Evidence for a second class of S-adenosylmethionine riboswitches and other regulatory RNA motifs in alpha-proteobacteria. Genome Biol. 6, R70 (2005).

    Article  Google Scholar 

  9. Reader, J.S., Metzgar, D., Schimmel, P. & de Crécy-Lagard, V. Identification of four genes necessary for biosynthesis of the modified nucleoside queuosine. J. Biol. Chem. 279, 6280–6285 (2004).

    Article  CAS  Google Scholar 

  10. Harada, F. & Nishimura, S. Possible anticodon sequences of tRNAHis, tRNAAsn, and tRNAAsp from Escherichia coli B. Universal presence of nucleoside Q in the first position of the anticodons of these transfer ribonucleic acids. Biochemistry 11, 301–308 (1972).

    Article  CAS  Google Scholar 

  11. Van Lanen, S.G. et al. From cyclohydrolase to oxidoreductase: discovery of nitrile reductase activity in a common fold. Proc. Natl. Acad. Sci. USA 102, 4264–4269 (2005).

    Article  CAS  Google Scholar 

  12. Gaur, R. & Varshney, U. Genetic analysis identifies a function for the queC (ybaX) gene product at an initial step in the queuosine biosynthetic pathway in Escherichia coli. J. Bacteriol. 187, 6893–6901 (2005).

    Article  CAS  Google Scholar 

  13. Okada, N. et al. Novel mechanism of post-transcriptional modification of tRNA. Insertion of bases of Q precursors into tRNA by a specific tRNA transglycosylase reaction. J. Biol. Chem. 254, 3067–3073 (1979).

    CAS  PubMed  Google Scholar 

  14. Reuter, K., Slany, R., Ullrich, F. & Kersten, H. Structure and organization of Escherichia coli genes involved in biosynthesis of the deazaguanine derivative queuine, a nutrient factor for eukaryotes. J. Bacteriol. 173, 2256–2264 (1991).

    Article  CAS  Google Scholar 

  15. Salazar, J.C., Ambrogelly, A., Crain, P.F., McCloskey, J.A. & Soll, D. A truncated aminoacyl-tRNA synthetase modifies RNA. Proc. Natl. Acad. Sci. USA 101, 7536–7541 (2004).

    Article  CAS  Google Scholar 

  16. Blaise, M. et al. A minimalist glutamyl-tRNA synthetase dedicated to aminoacylation of the tRNAAsp QUC anticodon. Nucleic Acids Res. 32, 2768–2775 (2004).

    Article  CAS  Google Scholar 

  17. Iwata-Reuyl, D. Biosynthesis of the 7-deazaguanosine hypermodified nucleosides of transfer RNA. Bioorg. Chem. 31, 24–43 (2003).

    Article  CAS  Google Scholar 

  18. Durand, J.M. et al. vacC, a virulence-associated chromosomal locus of Shigella flexneri, is homologous to tgt, a gene encoding tRNA-guanine transglycosylase (Tgt) of Escherichia coli K-12. J. Bacteriol. 176, 4627–4634 (1994).

    Article  CAS  Google Scholar 

  19. Noguchi, S., Nishimura, Y., Hirota, Y. & Nishimura, S. Isolation and characterization of an Escherichia coli mutant lacking tRNA-guanine transglycosylase. Function and biosynthesis of queuosine in tRNA. J. Biol. Chem. 257, 6544–6550 (1982).

    CAS  PubMed  Google Scholar 

  20. Frey, B., Janel, G., Michelsen, U. & Kersten, H. Mutations in the Escherichia coli fnr and tgt genes: control of molybdate reductase activity and the cytochrome d complex by fnr. J. Bacteriol. 171, 1524–1530 (1989).

    Article  CAS  Google Scholar 

  21. Bienz, M. & Kubli, E. Wild type tRNATyr reads the TMV RNA stop codon, but Q base-modified tRNATyr does not. Nature 294, 188–190 (1981).

    Article  CAS  Google Scholar 

  22. Meier, F., Suter, B., Grosjean, H., Keith, G. & Kubli, E. Queuosine modification of the wobble base in tRNAHis influences in vivo decoding properties. EMBO J. 4, 823–827 (1985).

    Article  CAS  Google Scholar 

  23. Urbonavičius, J., Qian, Q., Durand, J.M., Hagervall, T.G. & Björk, G.R. Improvement of reading frame maintenance is a common function for several tRNA modifications. EMBO J. 20, 4863–4873 (2001).

    Article  Google Scholar 

  24. Kuchino, Y., Kasai, H., Nihei, K. & Nishimura, S. Biosynthesis of the modified nucleoside Q in transfer RNA. Nucleic Acids Res. 3, 393–398 (1976).

    Article  CAS  Google Scholar 

  25. Okada, N. et al. Structure determination of a nucleoside Q precursor isolated from E. coli tRNA: 7-(aminomethyl)-7-deazaguanosine. Nucleic Acids Res. 5, 2289–2296 (1978).

    Article  CAS  Google Scholar 

  26. Slany, R.K., Bosl, M., Crain, P.F. & Kersten, H. A new function of S-adenosylmethionine: the ribosyl moiety of AdoMet is the precursor of the cyclopentenediol moiety of the tRNA wobble base queuine. Biochemistry 32, 7811–7817 (1993).

    Article  CAS  Google Scholar 

  27. Slany, R.K., Bosl, M. & Kersten, H. Transfer and isomerization of the ribose moiety of AdoMet during the biosynthesis of queuosine tRNAs, a new unique reaction catalyzed by the QueA protein from Escherichia coli. Biochimie 76, 389–393 (1994).

    Article  CAS  Google Scholar 

  28. Frey, B., McCloskey, J., Kersten, W. & Kersten, H. New function of vitamin B12: cobamide-dependent reduction of epoxyqueuosine to queuosine in tRNAs of Escherichia coli and Salmonella typhimurium. J. Bacteriol. 170, 2078–2082 (1988).

    Article  CAS  Google Scholar 

  29. Gusarov, I. & Nudler, E. The mechanism of intrinsic transcription termination. Mol. Cell 3, 495–504 (1999).

    Article  CAS  Google Scholar 

  30. Yarnell, W.S. & Roberts, J.W. Mechanism of intrinsic transcription termination and antitermination. Science 284, 611–615 (1999).

    Article  CAS  Google Scholar 

  31. Soukup, G.A. & Breaker, R.R. Relationship between internucleotide linkage geometry and the stability of RNA. RNA 5, 1308–1325 (1999).

    Article  CAS  Google Scholar 

  32. Wickiser, J.K., Winkler, W.C., Breaker, R.R. & Crothers, D.M. The speed of RNA transcription and metabolite binding kinetics operate an FMN riboswitch. Mol. Cell 18, 49–60 (2005).

    Article  CAS  Google Scholar 

  33. Wickiser, J.K., Cheah, M.T., Breaker, R.R. & Crothers, D.M. The kinetics of ligand binding by an adenine-sensing riboswitch. Biochemistry 44, 13404–13414 (2005).

    Article  CAS  Google Scholar 

  34. Lemay, J.F., Penedo, J.C., Tremblay, R., Lilley, D.M. & Lafontaine, D.A. Folding of the adenine riboswitch. Chem. Biol. 13, 857–868 (2006).

    Article  CAS  Google Scholar 

  35. Gilbert, S.D., Stoddard, C.D., Wise, S.J. & Batey, R.T. Thermodynamic and kinetic characterization of ligand binding to the purine riboswitch aptamer domain. J. Mol. Biol. 359, 754–768 (2006).

    Article  CAS  Google Scholar 

  36. Mandal, M. & Breaker, R.R. Adenine riboswitches and gene activation by disruption of a transcription terminator. Nat. Struct. Mol. Biol. 11, 29–35 (2004).

    Article  CAS  Google Scholar 

  37. Batey, R.T., Gilbert, S.D. & Montange, R.K. Structure of a natural guanine-responsive riboswitch complexed with the metabolite hypoxanthine. Nature 432, 411–415 (2004).

    Article  CAS  Google Scholar 

  38. Serganov, A. et al. Structural basis for discriminative regulation of gene expression by adenine- and guanine-sensing mRNAs. Chem. Biol. 11, 1729–1741 (2004).

    Article  CAS  Google Scholar 

  39. Noeske, J. et al. An intermolecular base triple as the basis of ligand specificity and affinity in the guanine- and adenine-sensing riboswitch RNAs. Proc. Natl. Acad. Sci. USA 102, 1372–1377 (2005).

    Article  CAS  Google Scholar 

  40. Fuchs, R.T., Grundy, F.J. & Henkin, T.M. The S(MK) box is a new SAM-binding RNA for translational regulation of SAM synthetase. Nat. Struct. Mol. Biol. 13, 226–233 (2006).

    Article  CAS  Google Scholar 

  41. Tatusov, R.L. et al. The COG database: new developments in phylogenetic classification of proteins from complete genomes. Nucleic Acids Res. 29, 22–28 (2001).

    Article  CAS  Google Scholar 

  42. Dewey, V. & Kidder, G.W. Partial purification and properties of a nucleoside hydrolase from Crithidia. Arch. Biochem. Biophys. 157, 380–387 (1973).

    Article  CAS  Google Scholar 

  43. Miller, R.L., Sabourin, C.L., Krenitsky, T.A., Berens, R.L. & Marr, J.J. Nucleoside hydrolases from Trypanosoma cruzi. J. Biol. Chem. 259, 5073–5077 (1984).

    CAS  PubMed  Google Scholar 

  44. Gündüz, U. & Katze, J.R. Salvage of the nucleic acid base queuine from queuine-containing tRNA by animal cells. Biochem. Biophys. Res. Commun. 109, 159–167 (1982).

    Article  Google Scholar 

  45. Gündüz, U. & Katze, J.R. Queuine salvage in mammalian cells. Evidence that queuine is generated from queuosine 5′-phosphate. J. Biol. Chem. 259, 1110–1113 (1984).

    PubMed  Google Scholar 

  46. Nissen, P., Ippolito, J.A., Ban, N., Moore, P.B. & Steitz, T.A. RNA tertiary interactions in the large ribosomal subunit: the A-minor motif. Proc. Natl. Acad. Sci. USA 98, 4899–4903 (2001).

    Article  CAS  Google Scholar 

  47. Akimoto, H., Imamiya, E., Hitaka, T., Nomura, H. & Nishimura, S. Synthesis of queuine, the base of naturally occurring hypermodified nucleoside (queuosine), and its analogues. J. Chem. Soc. [Perkin 1] 1988, 1637–1644 (1988).

    Article  Google Scholar 

  48. Migawa, M.T., Hinkley, J.M., Hoops, G.C. & Townsend, L.B. A two step synthesis of the nucleoside Q precursor 2-amino-5-cyanopyrrolo[2,3-d]pyrimidine-4-one (preQ0). Synth. Commun. 26, 3317–3322 (1996).

    Article  CAS  Google Scholar 

  49. Akimoto, H. et al. Queuine analogues. Their synthesis and inhibition of growth of mouse L5178Y cells in vitro. J. Med. Chem. 29, 1749–1753 (1986).

    Article  CAS  Google Scholar 

  50. West, R.A. 4-Hydroxypyrrolo[2,3-d]pyrimidine: Mannich reaction. J. Org. Chem. 26, 4959–4961 (1961).

    Article  CAS  Google Scholar 

  51. Wellcome Foundation, Ltd. Great Britain patent number 981458 (1965).

  52. Eddy, S.R. & Durbin, R. RNA sequence analysis using covariance models. Nucleic Acids Res. 22, 2079–2088 (1994).

    Article  CAS  Google Scholar 

  53. Weinberg, Z. & Ruzzo, W.L. Sequence-based heuristics for faster annotation of non-coding RNA families. Bioinformatics 22, 35–39 (2006).

    Article  CAS  Google Scholar 

  54. Tatusov, R.L. et al. The COG database: an updated version includes eukaryotes. BMC Bioinformatics 4, 41 (2003).

    Article  Google Scholar 

  55. Gerstein, M., Sonnhammer, E.L. & Chothia, C. Volume changes in protein evolution. J. Mol. Biol. 236, 1067–1078 (1994).

    Article  CAS  Google Scholar 

  56. Guérout-Fleury, A.M., Frandsen, N. & Stragier, P. Plasmids for ectopic integration in Bacillus subtilis. Gene 180, 57–61 (1996).

    Article  Google Scholar 

  57. Roth, A., Nahvi, A., Lee, M., Jona, I. & Breaker, R.R. Characteristics of the glmS ribozyme suggest only structural roles for divalent metal ions. RNA 12, 607–619 (2006).

    Article  CAS  Google Scholar 

  58. Mandal, M., Boese, B., Barrick, J.E., Winkler, W.C. & Breaker, R.R. Riboswitches control fundamental biochemical pathways in Bacillus subtilis and other bacteria. Cell 113, 577–586 (2003).

    Article  CAS  Google Scholar 

  59. Sudarsan, N., Wickiser, J.K., Nakamura, S., Ebert, M.S. & Breaker, R.R. An mRNA structure in bacteria that controls gene expression by binding lysine. Genes Dev. 17, 2688–2697 (2003).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank J.A. Collins for assistance in generating B. subtilis transformants and members of the Breaker laboratory for helpful discussions. This work was supported by grants from the US National Institutes of Health (R33 DK07027 and GM 068819) and the US National Science Foundation (EIA 0323510) to R.R.B. RNA research in the Breaker laboratory is also supported by the Howard Hughes Medical Institute.

Author information

Author notes

  1. Wade C Winkler & Jeffrey E Barrick

    Present address: Present addresses: Department of Biochemistry, The University of Texas Southwestern Medical Center, Dallas, Texas 75390-9038, USA (W.C.W.) and Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan 48824, USA (J.E.B.).,

Authors and Affiliations

  1. Howard Hughes Medical Institute, Yale University, P.O. Box 208103, New Haven, 06520-8103, Connecticut, USA

    Adam Roth, Jeffrey E Barrick & Ronald R Breaker

  2. Department of Molecular, Cellular and Developmental Biology, Yale University, P.O. Box 208103, New Haven, 06520-8103, Connecticut, USA

    Wade C Winkler, Elizabeth E Regulski, Jinsoo Lim, Inbal Jona, Ankita Ritwik, Jane N Kim, Rüdiger Welz & Ronald R Breaker

  3. Department of Molecular Biophysics and Biochemistry, Yale University, P.O. Box 208103, New Haven, 06520-8103, Connecticut, USA

    Jeffrey E Barrick & Ronald R Breaker

  4. Department of Chemistry, Portland State University, P.O. Box 751, Portland, 97207, Oregon, USA

    Bobby W K Lee & Dirk Iwata-Reuyl

Authors
  1. Adam Roth
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Wade C Winkler
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Elizabeth E Regulski
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Bobby W K Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Jinsoo Lim
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Inbal Jona
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Jeffrey E Barrick
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Ankita Ritwik
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Jane N Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Rüdiger Welz
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Dirk Iwata-Reuyl
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Ronald R Breaker
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

A. Roth, W.C.W., E.E.R., I.J., A. Ritwik, J.N.K. and R.W. carried out in vitro biochemical analyses of queC RNA constructs. E.E.R. conducted assays of reporter gene expression in vivo. B.W.K.L., J.L. and D.I.-R. synthesized preQ1 and related compounds. J.E.B. performed bioinformatics searches and prepared sequence alignments. A. Roth and R.R.B. prepared the manuscript.

Corresponding author

Correspondence to Ronald R Breaker.

Ethics declarations

Competing interests

R.R.B. is a cofounder of BioRelix, a company interested in the use of riboswitches as drug targets. Although this publication is not directly related to the development of antimicrobials and thus does not currently constitute a competing financial interest, the discovery of novel riboswitches may ultimately lead to development of new classes of antibiotics.

Supplementary information

Supplementary Fig. 1

PreQ1 aptamer sequences. (PDF 128 kb)

Supplementary Fig. 2

PreQ1 riboswitch locations and associated genes. (PDF 164 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Roth, A., Winkler, W., Regulski, E. et al. A riboswitch selective for the queuosine precursor preQ1 contains an unusually small aptamer domain. Nat Struct Mol Biol 14, 308–317 (2007). https://doi.org/10.1038/nsmb1224

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb1224

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing