Molecular and General Genetics MGG

, Volume 217, Issue 2–3, pp 209–214 | Cite as

Nuclear proteins binding to a cauliflower mosaic virus 35S truncated promoter

  • Salomé Prat
  • L. Willmitzer
  • Jose J. Sánchez-Serrano


Proteins present in tobacco nuclear extracts bind to a truncated cauliflower mosaic virus (CaMV) 35S promoter fragment (from −90 to +2 relative to the transcription start site) in a sequence specific manner. Gel mobility shift assays show the presence of two protein-DNA complexes that are not competed by a −47/+2 promoter fragment. DNAse I protection and DNA methylation interference reveal two protected sites in the slower migrating complex; both include the pentamer TGACG, separated by a stretch of eight nucleotides where G methylation does not prevent the binding of the proteins. The faster complex is the prevalent form at low protein concentrations. As the protein concentration increases a non-linear rise in the amount of the slower migrating complex relative to the faster one is seen suggesting that cooperative effects are involved in the binding to the second site.

Key words

Cauliflower mosaic virus 35S truncated promoter DNA-protein interactions Cooperative binding 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Ausubel F, Brent R, Kingston RE, Moore DD, Smith JA, Seidman JG, Struhl K (1987) Current protocols in Molecular Biology. J. Wiley and Sons, Greene Publishing Associates, Brooklyn, NY, USAGoogle Scholar
  2. Covey SN, Hull R (1985) Advances in Cauliflower Mosaic Virus research. Oxford Surveys of Plant Molecular and Cell Biology, vol. 2, pp 339–346Google Scholar
  3. De Block M, Botterman J, Vandewiele M, Dockx J, Thoen C, Gosselé V, Rao Movva N, Thompson C, Van Montagu M, Leemans J (1987) Engineering herbicide resistance in plants by expression of a detoxifying enzyme. EMBO J 6:2513–2518PubMedGoogle Scholar
  4. Fluhr R, Kuhlmeier C, Nagy F, Chua N-H (1986) Organ-specific and light-induced expression of plant genes. Science 232:1106–1115PubMedGoogle Scholar
  5. Fried M, Crothers DM (1981) Equilibria and kinetics of lac repressor-operator interactions by polyacrylamide gel electrophoresis. Nucleic Acids Res 9:6505–6525PubMedGoogle Scholar
  6. Galas DJ, Schmitz A (1978) DNAase footprinting: a simple method for the detection of protein-DNA binding specificity. Nucleic Acids Res 5:3157–3170PubMedGoogle Scholar
  7. Hendrickson W, Schleif R (1985) A dimer of AraC protein contacts three adjacent major groove regions of the araI DNA site. Proc Natl Acad Sci USA 82:3129–3133PubMedCrossRefGoogle Scholar
  8. Maxam A, Gilbert W (1980) Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol 65:499–560PubMedCrossRefGoogle Scholar
  9. Nagy F, Boutry M, Hsu M-Y, Wong M, Chua N-H (1987) The 5′-proximal region of the wheat Cab-1 gene contains a 268-bp enhancer-like sequence for phytochrome response. EMBO J 6:2537–2542PubMedGoogle Scholar
  10. Odell JT, Nagy F, Chua N-H (1985) Identification of DNA sequences required for activity of the Cauliflower Mosaic Virus 35S promoter. Nature 313:810–812PubMedCrossRefGoogle Scholar
  11. Ow DW, Jacobs JD, Howell SH (1987) Functional regions of the cauliflower mosaic virus 35S RNA promoter determined by use of the firefly luciferase gene as a reporter activity. Proc Natl Acad Sci USA 84:4870–4874PubMedCrossRefGoogle Scholar
  12. Parker CS, Topol J, (1984) ADrosophila RNA polymerase II transcription factor contains a promoter-region-specific DNA-binding activity. Cell 36:357–369PubMedCrossRefGoogle Scholar
  13. Poulsen C, Chua N-H (1988) Dissection of 5′ upstream sequences for selective expression of theNicotiana plumbaginifolia rbcS-8B gene. Mol Gen Genet 214:16–23PubMedCrossRefGoogle Scholar
  14. Powell Abel P, Nelson RS, De B, Hoffmann N, Rogers S, Fraley RT, Beachy RN (1986) Delay of disease development in transgenic plants that express the Tobacco Mosaic Virus coat protein gene. Science 232:738–743Google Scholar
  15. Rocha-Sosa M, Sonnewald U, Frommer W, Stratmann M, Schell J, Willmitzer L (1989) Both developmental and metabolic signals activate the promoter of a class I patatin gene. EMBO J (in press)Google Scholar
  16. Sánchez-Serrano JJ, Keil M, O’Connor A, Schell J, Willmitzer L (1987) Wound-induced expression of a potato proteinase inhibitor II gene in transgenic tobacco plants. EMBO J 6:303–306PubMedGoogle Scholar
  17. Stockhaus J, Eckes P, Rocha-Sosa M, Schell J, Willmitzer L (1987) Analysis of cis-active sequences involved in the leaf-specific expression of a potato gene in transgenic plants. Proc Natl Acad Sci USA 84:7943–7947PubMedCrossRefGoogle Scholar
  18. Vaeck M, Reynaerts R, Hofte H, Janssens S, De Beuckeleer M, Dean C, Zabeau M, Van Montagu M, Leemans J (1987) Transgenic plants protected from insect attack. Nature 328:33–37CrossRefGoogle Scholar
  19. Yanisch-Perron C, Vieira J, Messing J (1985) Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene 33:103–119PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 1989

Authors and Affiliations

  • Salomé Prat
    • 1
  • L. Willmitzer
    • 1
  • Jose J. Sánchez-Serrano
    • 1
  1. 1.Institut für Genbiologische Forschung Berlin GmbHBerlin 33

Personalised recommendations