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. 1997 Feb 1;25(3):668–674. doi: 10.1093/nar/25.3.668

Altered structure of the DNA duplex recognized by yeast transcription factor Reb1p.

D R Davis 1, D J Stillman 1
PMCID: PMC146484  PMID: 9016611

Abstract

The Saccharomyces cerevisiae REB1 gene encodes a sequence-specific DNA binding protein that has been implicated in chromatin structure, transcription regulation and transcription termination. Previous work has shown that the DNA sequence recognized by Reb1p contains an adenosine residue that is unusually reactive toward chemical modification by dimethylsulfate and that methylation of this nucleoside increases the binding affinity of the Reb1p protein for its target. Prompted by these results, we determined the solution structure of the 13mer Reb1p DNA duplex recognition site d(GTCCGGGTAATGC).d(GCATTACCCGGAC) using 2D NMR, distance geometry and iterative 2D NOESY back-calculation structure refinement. The distance geometry-refined molecule demonstrated an unusual structure in the TAAT region of the sequence that was manifested in cross-strand base stacking, as indicated by unusually strong NOE interactions between H2 protons on three adjacent adenosine bases. This structure was compared to two published NMR studies of DNA duplexes containing the related sequence TAAC. The Reb1p DNA structure does not show the conformational mobility or the 'transient kink' at TpA steps characteristic of the related TAAT-containing sequences.

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Selected References

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  1. Banks K. M., Hare D. R., Reid B. R. Three-dimensional solution structure of a DNA duplex containing the BclI restriction sequence: two-dimensional NMR studies, distance geometry calculations, and refinement by back-calculation of the NOESY spectrum. Biochemistry. 1989 Aug 22;28(17):6996–7010. doi: 10.1021/bi00443a033. [DOI] [PubMed] [Google Scholar]
  2. Brandl C. J., Struhl K. A nucleosome-positioning sequence is required for GCN4 to activate transcription in the absence of a TATA element. Mol Cell Biol. 1990 Aug;10(8):4256–4265. doi: 10.1128/mcb.10.8.4256. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Carmen A. A., Holland M. J. The upstream repression sequence from the yeast enolase gene ENO1 is a complex regulatory element that binds multiple trans-acting factors including REB1. J Biol Chem. 1994 Apr 1;269(13):9790–9797. [PubMed] [Google Scholar]
  4. Chasman D. I., Lue N. F., Buchman A. R., LaPointe J. W., Lorch Y., Kornberg R. D. A yeast protein that influences the chromatin structure of UASG and functions as a powerful auxiliary gene activator. Genes Dev. 1990 Apr;4(4):503–514. doi: 10.1101/gad.4.4.503. [DOI] [PubMed] [Google Scholar]
  5. Erkine A. M., Adams C. C., Gao M., Gross D. S. Multiple protein-DNA interactions over the yeast HSC82 heat shock gene promoter. Nucleic Acids Res. 1995 May 25;23(10):1822–1829. doi: 10.1093/nar/23.10.1822. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Fedor M. J., Lue N. F., Kornberg R. D. Statistical positioning of nucleosomes by specific protein-binding to an upstream activating sequence in yeast. J Mol Biol. 1988 Nov 5;204(1):109–127. doi: 10.1016/0022-2836(88)90603-1. [DOI] [PubMed] [Google Scholar]
  7. Hare D. R., Reid B. R. Three-dimensional structure of a DNA hairpin in solution: two-dimensional NMR studies and distance geometry calculations on d(CGCGTTTTCGCG). Biochemistry. 1986 Sep 9;25(18):5341–5350. doi: 10.1021/bi00366a053. [DOI] [PubMed] [Google Scholar]
  8. Hare D. R., Wemmer D. E., Chou S. H., Drobny G., Reid B. R. Assignment of the non-exchangeable proton resonances of d(C-G-C-G-A-A-T-T-C-G-C-G) using two-dimensional nuclear magnetic resonance methods. J Mol Biol. 1983 Dec 15;171(3):319–336. doi: 10.1016/0022-2836(83)90096-7. [DOI] [PubMed] [Google Scholar]
  9. Hare D., Shapiro L., Patel D. J. Wobble dG X dT pairing in right-handed DNA: solution conformation of the d(C-G-T-G-A-A-T-T-C-G-C-G) duplex deduced from distance geometry analysis of nuclear Overhauser effect spectra. Biochemistry. 1986 Nov 18;25(23):7445–7456. doi: 10.1021/bi00371a029. [DOI] [PubMed] [Google Scholar]
  10. Ju Q. D., Morrow B. E., Warner J. R. REB1, a yeast DNA-binding protein with many targets, is essential for growth and bears some resemblance to the oncogene myb. Mol Cell Biol. 1990 Oct;10(10):5226–5234. doi: 10.1128/mcb.10.10.5226. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kennedy M. A., Nuutero S. T., Davis J. T., Drobny G. P., Reid B. R. Mobility at the TpA cleavage site in the T3A3-containing AhaIII and PmeI restriction sequences. Biochemistry. 1993 Aug 10;32(31):8022–8035. doi: 10.1021/bi00082a025. [DOI] [PubMed] [Google Scholar]
  12. Kim S. G., Lin L. J., Reid B. R. Determination of nucleic acid backbone conformation by 1H NMR. Biochemistry. 1992 Apr 14;31(14):3564–3574. doi: 10.1021/bi00129a003. [DOI] [PubMed] [Google Scholar]
  13. Kintanar A., Klevit R. E., Reid B. R. Two-dimensional NMR investigation of a bent DNA fragment: assignment of the proton resonances and preliminary structure analysis. Nucleic Acids Res. 1987 Jul 24;15(14):5845–5862. doi: 10.1093/nar/15.14.5845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lang W. H., Morrow B. E., Ju Q., Warner J. R., Reeder R. H. A model for transcription termination by RNA polymerase I. Cell. 1994 Nov 4;79(3):527–534. doi: 10.1016/0092-8674(94)90261-5. [DOI] [PubMed] [Google Scholar]
  15. Lang W. H., Reeder R. H. The REB1 site is an essential component of a terminator for RNA polymerase I in Saccharomyces cerevisiae. Mol Cell Biol. 1993 Jan;13(1):649–658. doi: 10.1128/mcb.13.1.649. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Liaw P. C., Brandl C. J. Defining the sequence specificity of the Saccharomyces cerevisiae DNA binding protein REB1p by selecting binding sites from random-sequence oligonucleotides. Yeast. 1994 Jun;10(6):771–787. doi: 10.1002/yea.320100608. [DOI] [PubMed] [Google Scholar]
  17. Lin C. H., Hill G. C., Hurley L. H. Characterization of a 12-mer duplex d(GGCGGAGTTAGG).d(CCTAACTCCGCC) containing a highly reactive (+)-CC-1065 sequence by 1H and 31P NMR, hydroxyl-radical footprinting, and NOESY restrained molecular dynamics calculations. Chem Res Toxicol. 1992 Mar-Apr;5(2):167–182. doi: 10.1021/tx00026a005. [DOI] [PubMed] [Google Scholar]
  18. Morrow B. E., Ju Q., Warner J. R. A bipartite DNA-binding domain in yeast Reb1p. Mol Cell Biol. 1993 Feb;13(2):1173–1182. doi: 10.1128/mcb.13.2.1173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Morrow B. E., Ju Q., Warner J. R. Purification and characterization of the yeast rDNA binding protein REB1. J Biol Chem. 1990 Dec 5;265(34):20778–20783. [PubMed] [Google Scholar]
  20. Radha P. K., Madan A., Nibedita R., Hosur R. V. Solution structure of the conserved segment of the Myb cognate DNA sequence by 2D NMR, spectral simulation, restrained energy minimization, and distance geometry calculations. Biochemistry. 1995 May 2;34(17):5913–5922. doi: 10.1021/bi00017a021. [DOI] [PubMed] [Google Scholar]
  21. Reid B. R., Banks K., Flynn P., Nerdal W. NMR distance measurements in DNA duplexes: sugars and bases have the same correlation times. Biochemistry. 1989 Dec 26;28(26):10001–10007. doi: 10.1021/bi00452a019. [DOI] [PubMed] [Google Scholar]
  22. Reid B. R. Sequence-specific assignments and their use in NMR studies of DNA structure. Q Rev Biophys. 1987 Aug;20(1-2):1–34. doi: 10.1017/s0033583500004212. [DOI] [PubMed] [Google Scholar]
  23. Remacle J. E., Holmberg S. A REB1-binding site is required for GCN4-independent ILV1 basal level transcription and can be functionally replaced by an ABF1-binding site. Mol Cell Biol. 1992 Dec;12(12):5516–5526. doi: 10.1128/mcb.12.12.5516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Schlaepfer I. R., Mattoon J. R., Bajszár G. The sequence and potential regulatory elements of the HEM2 promoter of Saccharomyces cerevisiae. Yeast. 1994 Feb;10(2):227–229. doi: 10.1002/yea.320100209. [DOI] [PubMed] [Google Scholar]
  25. Scott E. W., Baker H. V. Concerted action of the transcriptional activators REB1, RAP1, and GCR1 in the high-level expression of the glycolytic gene TPI. Mol Cell Biol. 1993 Jan;13(1):543–550. doi: 10.1128/mcb.13.1.543. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Wang H., Nicholson P. R., Stillman D. J. Identification of a Saccharomyces cerevisiae DNA-binding protein involved in transcriptional regulation. Mol Cell Biol. 1990 Apr;10(4):1743–1753. doi: 10.1128/mcb.10.4.1743. [DOI] [PMC free article] [PubMed] [Google Scholar]

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