Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1995 Jun 11;23(11):1936–1941. doi: 10.1093/nar/23.11.1936

Overcoming potassium-mediated triplex inhibition.

W M Olivas 1, L J Maher 3rd 1
PMCID: PMC306966  PMID: 7596821

Abstract

Sequence-specific duplex DNA recognition by oligonucleotide-directed triple helix formation is a possible approach to in vivo gene inhibition. However, triple helix formation involving guanine-rich oligonucleotides is inhibited by physiological ions, particularly K+, most likely due to oligonucleotide aggregation via guanine quartets. Three oligodeoxynucleotide (ODN) derivatives were tested for their ability to resist guanine quartet-mediated aggregation, yet form stable triplexes. Electrophoretic mobility shift and dimethyl sulfate footprinting assays were used to analyze the formation of triplexes involving these oligonucleotide derivatives. In the absence of K+, all ODNs had similar binding affinities for the duplex target. Triplexes involving a 14mer ODN derivative containing 7-deazaxanthine substituted for three thymine bases or an 18mer ODN containing two additional thymines on both the 5' and 3' termini were abolished by 50 mM K+. Remarkably, triplexes involving an ODN derivative containing four 6-thioguanine bases substituted for guanine resisted K+ inhibition up to 200 mM. We hypothesize that the increased radius and decreased electronegativity of sulfur in the 6-position of guanine destabilize potential guanine quartets. These results improve the prospects for creating ODNS that might serve as specific and efficient gene repressors in vivo.

Full text

PDF
1936

Images in this article

1938Image
on p.1938
1939Image
on p.1939
1939Image
on p.1939
1940Image
on p.1940

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beal P. A., Dervan P. B. Second structural motif for recognition of DNA by oligonucleotide-directed triple-helix formation. Science. 1991 Mar 15;251(4999):1360–1363. doi: 10.1126/science.2003222. [DOI] [PubMed] [Google Scholar]
  2. Cheng A. J., Van Dyke M. W. Monovalent cation effects on intermolecular purine-purine-pyrimidine triple-helix formation. Nucleic Acids Res. 1993 Dec 11;21(24):5630–5635. doi: 10.1093/nar/21.24.5630. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cooney M., Czernuszewicz G., Postel E. H., Flint S. J., Hogan M. E. Site-specific oligonucleotide binding represses transcription of the human c-myc gene in vitro. Science. 1988 Jul 22;241(4864):456–459. doi: 10.1126/science.3293213. [DOI] [PubMed] [Google Scholar]
  4. Elion G. B., Hitchings G. H. Metabolic basis for the actions of analogs of purines and pyrimidines. Adv Chemother. 1965;2:91–177. doi: 10.1016/b978-1-4831-9930-6.50008-3. [DOI] [PubMed] [Google Scholar]
  5. Guschlbauer W., Chantot J. F., Thiele D. Four-stranded nucleic acid structures 25 years later: from guanosine gels to telomer DNA. J Biomol Struct Dyn. 1990 Dec;8(3):491–511. doi: 10.1080/07391102.1990.10507825. [DOI] [PubMed] [Google Scholar]
  6. Ing N. H., Beekman J. M., Kessler D. J., Murphy M., Jayaraman K., Zendegui J. G., Hogan M. E., O'Malley B. W., Tsai M. J. In vivo transcription of a progesterone-responsive gene is specifically inhibited by a triplex-forming oligonucleotide. Nucleic Acids Res. 1993 Jun 25;21(12):2789–2796. doi: 10.1093/nar/21.12.2789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Laughlan G., Murchie A. I., Norman D. G., Moore M. H., Moody P. C., Lilley D. M., Luisi B. The high-resolution crystal structure of a parallel-stranded guanine tetraplex. Science. 1994 Jul 22;265(5171):520–524. doi: 10.1126/science.8036494. [DOI] [PubMed] [Google Scholar]
  8. Maher L. J., 3rd DNA triple-helix formation: an approach to artificial gene repressors? Bioessays. 1992 Dec;14(12):807–815. doi: 10.1002/bies.950141204. [DOI] [PubMed] [Google Scholar]
  9. Maher L. J., 3rd, Dervan P. B., Wold B. Analysis of promoter-specific repression by triple-helical DNA complexes in a eukaryotic cell-free transcription system. Biochemistry. 1992 Jan 14;31(1):70–81. doi: 10.1021/bi00116a012. [DOI] [PubMed] [Google Scholar]
  10. Milligan J. F., Krawczyk S. H., Wadwani S., Matteucci M. D. An anti-parallel triple helix motif with oligodeoxynucleotides containing 2'-deoxyguanosine and 7-deaza-2'-deoxyxanthosine. Nucleic Acids Res. 1993 Jan 25;21(2):327–333. doi: 10.1093/nar/21.2.327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Moser H. E., Dervan P. B. Sequence-specific cleavage of double helical DNA by triple helix formation. Science. 1987 Oct 30;238(4827):645–650. doi: 10.1126/science.3118463. [DOI] [PubMed] [Google Scholar]
  12. Olivas W. M., Maher L. J., 3rd Competitive triplex/quadruplex equilibria involving guanine-rich oligonucleotides. Biochemistry. 1995 Jan 10;34(1):278–284. doi: 10.1021/bi00001a034. [DOI] [PubMed] [Google Scholar]
  13. Orson F. M., Thomas D. W., McShan W. M., Kessler D. J., Hogan M. E. Oligonucleotide inhibition of IL2R alpha mRNA transcription by promoter region collinear triplex formation in lymphocytes. Nucleic Acids Res. 1991 Jun 25;19(12):3435–3441. doi: 10.1093/nar/19.12.3435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Pieles U., Zürcher W., Schär M., Moser H. E. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry: a powerful tool for the mass and sequence analysis of natural and modified oligonucleotides. Nucleic Acids Res. 1993 Jul 11;21(14):3191–3196. doi: 10.1093/nar/21.14.3191. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Postel E. H., Flint S. J., Kessler D. J., Hogan M. E. Evidence that a triplex-forming oligodeoxyribonucleotide binds to the c-myc promoter in HeLa cells, thereby reducing c-myc mRNA levels. Proc Natl Acad Sci U S A. 1991 Sep 15;88(18):8227–8231. doi: 10.1073/pnas.88.18.8227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Rao T. S., Durland R. H., Seth D. M., Myrick M. A., Bodepudi V., Revankar G. R. Incorporation of 2'-deoxy-6-thioguanosine into G-rich oligodeoxyribonucleotides inhibits G-tetrad formation and facilitates triplex formation. Biochemistry. 1995 Jan 24;34(3):765–772. doi: 10.1021/bi00003a009. [DOI] [PubMed] [Google Scholar]
  17. Sen D., Gilbert W. Novel DNA superstructures formed by telomere-like oligomers. Biochemistry. 1992 Jan 14;31(1):65–70. doi: 10.1021/bi00116a011. [DOI] [PubMed] [Google Scholar]
  18. Singleton S. F., Dervan P. B. Influence of pH on the equilibrium association constants for oligodeoxyribonucleotide-directed triple helix formation at single DNA sites. Biochemistry. 1992 Nov 17;31(45):10995–11003. doi: 10.1021/bi00160a008. [DOI] [PubMed] [Google Scholar]
  19. Williamson J. R., Raghuraman M. K., Cech T. R. Monovalent cation-induced structure of telomeric DNA: the G-quartet model. Cell. 1989 Dec 1;59(5):871–880. doi: 10.1016/0092-8674(89)90610-7. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES