Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1998 Jul 1;333(Pt 1):51–56. doi: 10.1042/bj3330051

The first high-mobility-group box of upstream binding factor assembles across-over DNA junction by basic residues.

C H Hu 1, J M Wang 1, H B Tseng 1
PMCID: PMC1219554  PMID: 9639561

Abstract

Upstream binding factor (UBF) is a eukaryotic RNA polymerase I-specific transcription factor. Its predominant DNA-binding motif, ubfHMG box 1, preserves DNA assembling activity that can bind two or more DNA duplexes simultaneously to form a crossover DNA junction. Here we investigate the basis of crossover DNA-assembling activity of ubfHMG box 1 by extensive mutagenesis analyses and mobility shift assay. Although the ubfHMG box 1 preserves a high mobility group (HMG) core structure, changing a number of the consensus hydrophobic and aromatic residues to alanine did not inhibit its crossover-assembling activity. This indicates that these residues do not directly participate in protein-DNA interaction. However, altering a series of basic residues in the helices 1 and 2 regions or the N-terminal extended strand of the ubfHMG box 1 motif had severe effects on DNA-assembling activity; however, certain non-specific DNA binding activity still remained. This suggests that the ubfHMG box 1 motif might extensively contact the backbone of a crossover junction through its multiple basic residues. Mutating a hydrophobic residue in the terminal dimerization domain inhibited the association of truncated Xenopus UBF, but had little effect on its crossover-assembling activity. This indicates that the UBF-crossover DNA complex is not established by the association of individual DNA-bound peptides.

Full Text

The Full Text of this article is available as a PDF (433.2 KB).

Selected References

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

  1. Bachvarov D., Moss T. The RNA polymerase I transcription factor xUBF contains 5 tandemly repeated HMG homology boxes. Nucleic Acids Res. 1991 May 11;19(9):2331–2335. doi: 10.1093/nar/19.9.2331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Baxevanis A. D., Bryant S. H., Landsman D. Homology model building of the HMG-1 box structural domain. Nucleic Acids Res. 1995 Mar 25;23(6):1019–1029. doi: 10.1093/nar/23.6.1019. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Baxevanis A. D., Landsman D. The HMG-1 box protein family: classification and functional relationships. Nucleic Acids Res. 1995 May 11;23(9):1604–1613. doi: 10.1093/nar/23.9.1604. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bazett-Jones D. P., Leblanc B., Herfort M., Moss T. Short-range DNA looping by the Xenopus HMG-box transcription factor, xUBF. Science. 1994 May 20;264(5162):1134–1137. doi: 10.1126/science.8178172. [DOI] [PubMed] [Google Scholar]
  5. Bell S. P., Jantzen H. M., Tjian R. Assembly of alternative multiprotein complexes directs rRNA promoter selectivity. Genes Dev. 1990 Jun;4(6):943–954. doi: 10.1101/gad.4.6.943. [DOI] [PubMed] [Google Scholar]
  6. Bianchi M. E., Beltrame M., Paonessa G. Specific recognition of cruciform DNA by nuclear protein HMG1. Science. 1989 Feb 24;243(4894 Pt 1):1056–1059. doi: 10.1126/science.2922595. [DOI] [PubMed] [Google Scholar]
  7. Copenhaver G. P., Putnam C. D., Denton M. L., Pikaard C. S. The RNA polymerase I transcription factor UBF is a sequence-tolerant HMG-box protein that can recognize structured nucleic acids. Nucleic Acids Res. 1994 Jul 11;22(13):2651–2657. doi: 10.1093/nar/22.13.2651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Falciola L., Murchie A. I., Lilley D. M., Bianchi M. Mutational analysis of the DNA binding domain A of chromosomal protein HMG1. Nucleic Acids Res. 1994 Feb 11;22(3):285–292. doi: 10.1093/nar/22.3.285. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Grosschedl R., Giese K., Pagel J. HMG domain proteins: architectural elements in the assembly of nucleoprotein structures. Trends Genet. 1994 Mar;10(3):94–100. doi: 10.1016/0168-9525(94)90232-1. [DOI] [PubMed] [Google Scholar]
  10. Hardman C. H., Broadhurst R. W., Raine A. R., Grasser K. D., Thomas J. O., Laue E. D. Structure of the A-domain of HMG1 and its interaction with DNA as studied by heteronuclear three- and four-dimensional NMR spectroscopy. Biochemistry. 1995 Dec 26;34(51):16596–16607. doi: 10.1021/bi00051a007. [DOI] [PubMed] [Google Scholar]
  11. Hisatake K., Nishimura T., Maeda Y., Hanada K., Song C. Z., Muramatsu M. Cloning and structural analysis of cDNA and the gene for mouse transcription factor UBF. Nucleic Acids Res. 1991 Sep 11;19(17):4631–4637. doi: 10.1093/nar/19.17.4631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hu C. H., McStay B., Jeong S. W., Reeder R. H. xUBF, an RNA polymerase I transcription factor, binds crossover DNA with low sequence specificity. Mol Cell Biol. 1994 May;14(5):2871–2882. doi: 10.1128/mcb.14.5.2871. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jantzen H. M., Admon A., Bell S. P., Tjian R. Nucleolar transcription factor hUBF contains a DNA-binding motif with homology to HMG proteins. Nature. 1990 Apr 26;344(6269):830–836. doi: 10.1038/344830a0. [DOI] [PubMed] [Google Scholar]
  14. Kermekchiev M., Workman J. L., Pikaard C. S. Nucleosome binding by the polymerase I transactivator upstream binding factor displaces linker histone H1. Mol Cell Biol. 1997 Oct;17(10):5833–5842. doi: 10.1128/mcb.17.10.5833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Khadake J. R., Rao M. R. Condensation of DNA and chromatin by an SPKK-containing octapeptide repeat motif present in the C-terminus of histone H1. Biochemistry. 1997 Feb 4;36(5):1041–1051. doi: 10.1021/bi961617p. [DOI] [PubMed] [Google Scholar]
  16. Kuhn A., Voit R., Stefanovsky V., Evers R., Bianchi M., Grummt I. Functional differences between the two splice variants of the nucleolar transcription factor UBF: the second HMG box determines specificity of DNA binding and transcriptional activity. EMBO J. 1994 Jan 15;13(2):416–424. doi: 10.1002/j.1460-2075.1994.tb06276.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Lai Y. S., Tseng H. B., Hu C. H. The dimerization domain upstream binding factor contains multiple helical structures. Biochem Biophys Res Commun. 1996 Mar 27;220(3):816–823. doi: 10.1006/bbrc.1996.0487. [DOI] [PubMed] [Google Scholar]
  18. Laudet V., Stehelin D., Clevers H. Ancestry and diversity of the HMG box superfamily. Nucleic Acids Res. 1993 May 25;21(10):2493–2501. doi: 10.1093/nar/21.10.2493. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Love J. J., Li X., Case D. A., Giese K., Grosschedl R., Wright P. E. Structural basis for DNA bending by the architectural transcription factor LEF-1. Nature. 1995 Aug 31;376(6543):791–795. doi: 10.1038/376791a0. [DOI] [PubMed] [Google Scholar]
  20. Maeda Y., Hisatake K., Kondo T., Hanada K., Song C. Z., Nishimura T., Muramatsu M. Mouse rRNA gene transcription factor mUBF requires both HMG-box1 and an acidic tail for nucleolar accumulation: molecular analysis of the nucleolar targeting mechanism. EMBO J. 1992 Oct;11(10):3695–3704. doi: 10.1002/j.1460-2075.1992.tb05454.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. McStay B., Frazier M. W., Reeder R. H. xUBF contains a novel dimerization domain essential for RNA polymerase I transcription. Genes Dev. 1991 Nov;5(11):1957–1968. doi: 10.1101/gad.5.11.1957. [DOI] [PubMed] [Google Scholar]
  22. McStay B., Hu C. H., Pikaard C. S., Reeder R. H. xUBF and Rib 1 are both required for formation of a stable polymerase I promoter complex in X. laevis. EMBO J. 1991 Aug;10(8):2297–2303. doi: 10.1002/j.1460-2075.1991.tb07766.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Neil K. J., Ridsdale R. A., Rutherford B., Taylor L., Larson D. E., Glibetic M., Rothblum L. I., Harauz G. Structure of recombinant rat UBF by electron image analysis and homology modelling. Nucleic Acids Res. 1996 Apr 15;24(8):1472–1480. doi: 10.1093/nar/24.8.1472. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. O'Mahony D. J., Rothblum L. I. Identification of two forms of the RNA polymerase I transcription factor UBF. Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3180–3184. doi: 10.1073/pnas.88.8.3180. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. O'Mahony D. J., Smith S. D., Xie W., Rothblum L. I. Analysis of the phosphorylation, DNA-binding and dimerization properties of the RNA polymerase I transcription factors UBF1 and UBF2. Nucleic Acids Res. 1992 Mar 25;20(6):1301–1308. doi: 10.1093/nar/20.6.1301. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Pil P. M., Lippard S. J. Specific binding of chromosomal protein HMG1 to DNA damaged by the anticancer drug cisplatin. Science. 1992 Apr 10;256(5054):234–237. doi: 10.1126/science.1566071. [DOI] [PubMed] [Google Scholar]
  27. Putnam C. D., Copenhaver G. P., Denton M. L., Pikaard C. S. The RNA polymerase I transactivator upstream binding factor requires its dimerization domain and high-mobility-group (HMG) box 1 to bend, wrap, and positively supercoil enhancer DNA. Mol Cell Biol. 1994 Oct;14(10):6476–6488. doi: 10.1128/mcb.14.10.6476. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Putnam C. D., Pikaard C. S. Cooperative binding of the Xenopus RNA polymerase I transcription factor xUBF to repetitive ribosomal gene enhancers. Mol Cell Biol. 1992 Nov;12(11):4970–4980. doi: 10.1128/mcb.12.11.4970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Rost B., Sander C. Conservation and prediction of solvent accessibility in protein families. Proteins. 1994 Nov;20(3):216–226. doi: 10.1002/prot.340200303. [DOI] [PubMed] [Google Scholar]
  30. Rost B., Sander C. Improved prediction of protein secondary structure by use of sequence profiles and neural networks. Proc Natl Acad Sci U S A. 1993 Aug 15;90(16):7558–7562. doi: 10.1073/pnas.90.16.7558. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rost B., Sander C. Prediction of protein secondary structure at better than 70% accuracy. J Mol Biol. 1993 Jul 20;232(2):584–599. doi: 10.1006/jmbi.1993.1413. [DOI] [PubMed] [Google Scholar]
  32. Rost B., Sander C., Schneider R. PHD--an automatic mail server for protein secondary structure prediction. Comput Appl Biosci. 1994 Feb;10(1):53–60. doi: 10.1093/bioinformatics/10.1.53. [DOI] [PubMed] [Google Scholar]
  33. Stros M., Stokrová J., Thomas J. O. DNA looping by the HMG-box domains of HMG1 and modulation of DNA binding by the acidic C-terminal domain. Nucleic Acids Res. 1994 Mar 25;22(6):1044–1051. doi: 10.1093/nar/22.6.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Treiber D. K., Zhai X., Jantzen H. M., Essigmann J. M. Cisplatin-DNA adducts are molecular decoys for the ribosomal RNA transcription factor hUBF (human upstream binding factor). Proc Natl Acad Sci U S A. 1994 Jun 7;91(12):5672–5676. doi: 10.1073/pnas.91.12.5672. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Varga-Weisz P., van Holde K., Zlatanova J. Preferential binding of histone H1 to four-way helical junction DNA. J Biol Chem. 1993 Oct 5;268(28):20699–20700. [PubMed] [Google Scholar]
  36. Weir H. M., Kraulis P. J., Hill C. S., Raine A. R., Laue E. D., Thomas J. O. Structure of the HMG box motif in the B-domain of HMG1. EMBO J. 1993 Apr;12(4):1311–1319. doi: 10.1002/j.1460-2075.1993.tb05776.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Wellman S. E. Carboxyl-terminal peptides from histone H1 variants: DNA binding characteristics and solution conformation. Biopolymers. 1996 Oct;39(4):491–501. doi: 10.1002/(SICI)1097-0282(199610)39:4%3C491::AID-BIP2%3E3.0.CO;2-S. [DOI] [PubMed] [Google Scholar]
  38. Wen L., Huang J. K., Johnson B. H., Reeck G. R. A human placental cDNA clone that encodes nonhistone chromosomal protein HMG-1. Nucleic Acids Res. 1989 Feb 11;17(3):1197–1214. doi: 10.1093/nar/17.3.1197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Werner M. H., Huth J. R., Gronenborn A. M., Clore G. M. Molecular basis of human 46X,Y sex reversal revealed from the three-dimensional solution structure of the human SRY-DNA complex. Cell. 1995 Jun 2;81(5):705–714. doi: 10.1016/0092-8674(95)90532-4. [DOI] [PubMed] [Google Scholar]
  40. van Houte L. P., Chuprina V. P., van der Wetering M., Boelens R., Kaptein R., Clevers H. Solution structure of the sequence-specific HMG box of the lymphocyte transcriptional activator Sox-4. J Biol Chem. 1995 Dec 22;270(51):30516–30524. doi: 10.1074/jbc.270.51.30516. [DOI] [PubMed] [Google Scholar]
  41. van Houte L., van Oers A., van de Wetering M., Dooijes D., Kaptein R., Clevers H. The sequence-specific high mobility group 1 box of TCF-1 adopts a predominantly alpha-helical conformation in solution. J Biol Chem. 1993 Aug 25;268(24):18083–18087. [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES