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 novel DNA recognition mode by the NF-κB p65 homodimer

Nature Structural Biology volume 5, pages 67–73 (1998)Cite this article

Abstract

The crystal structure of the NF-κB p65 (RelA) homodimer in complex with a DNA target has been determined to 2.4 Å resolution. The two p65 subunits are not symmetrically disposed on the DNA target. The homodimer should optimally bind to a pseudo-palindromic nine base pair target with each subunit recognizing a 5′GGAA-3′ half site separated by a central A–T base pair. However, one of the subunits (subunit B) encounters a half site of 5′-GAAA-3′. The single base-pair change from G–C to A–T results in highly unfavorable interactions between this half site and the base contacting protein residues in subunit B, which leads to an 18° rotation of the N-terminal terminal domain from its normal conformation. Remarkably, subunit B retains all the interactions with the sugar phosphate backbone of the DNA target. This mode of interaction allows the NF-κB p65 homodimer to recognize DNA targets containing only one cognate half site. Differences in the sequence of the other half site provide variations in conformation and affinity of the complex.

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

Similar content being viewed by others

References

  1. Baeurle, P.A. & Henkel, T. Function and activation of NF-kappa B in the immune system. Annu. Rev. Immunol. 12, 141–179 (1994).

    Article  Google Scholar 

  2. Baldwin, A.S., NF-kappa B and I kappa B proteins: new discoveries and insights. Annu. Rev. Immunol. 14, 649–683 (1996).

    Article  CAS  PubMed  Google Scholar 

  3. Siebenlist, U., Franzoso, G. & Brown, K., Structure, regulation and function of NF-kappa B. Annu. Rev. Cell. Biol. 10, 405–455 (1994).

    Article  CAS  PubMed  Google Scholar 

  4. Grilli, M., Chiu, J.J. & Lenardo, M.J. NF-kappa B and Rel: participants in a multiform transcriptional regulatory system. Int Rev Cytol 143, 1–62 (1993).

    Article  CAS  PubMed  Google Scholar 

  5. Kunsch, C., Ruben, S.M. & Rosen, C.A. Selection of optimal kappa B/Rel DNA-binding motifs: interaction of both subunits of NF-kappa B with DNA is required for transcriptional activation. Mol Cell Biol 12, 4412–4421 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Casoiaro, V. et al. Inhibition of NF-AT-dependent transcription by NF-kappa B: implications for differential gene expression in T helper cell subsets. Proc. Natl. Acad. Sci. USA 92, 11623–11627 (1995).

    Article  Google Scholar 

  7. Ghosh, G., van Duyne, G., Ghosh, S. & Sigler, P.B. Structure of NF-kappa B p50 homodimer bound to a kappa B site. Nature 373, 303–310 (1995).

    Article  CAS  PubMed  Google Scholar 

  8. Muller, C.W., Rey, F.A., Sodeoka, M., Verdine, G.L. & Harrison, S.C. Structure of the NF-kappa B p50 homodimer bound to DNA. Nature 373, 311–317 (1995).

    Article  CAS  PubMed  Google Scholar 

  9. Thanos, D. & Maniatis, T. Identification of the rel family members required for virus induction of the human beta interferon gene. Mol. Cell. Biol. 15, 152–164 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Huang, D.-B., Huxford, T., Chen, Y.-Q. & Ghosh, G. The role of DNA in the mechanism of NFkB dimer formation: crystal structures of the dimerization domains of the p50 and p65 subunits. Structure 5 1427–1436 (1997).

    Article  CAS  PubMed  Google Scholar 

  11. Falvo, J.V., Thanos, D. & Maniatis, T. Reversal of intrinsic DNA bends in the IFN beta gene enhancer by transcription factors and the architectural protein HMG I(Y). Cell 83, 1101–1111(1995).

    Article  CAS  PubMed  Google Scholar 

  12. Bell, S., Matthews, J.R., Jaffray, E. & Hay, R.T. l(kappa)B(gamma) inhibits DNA binding of NF-kappaB p50 homodimers by interacting with residues that contact DNA. Mol. Cell. Biol. 16, 6477–6485 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Bressler, P. et al. Mutational analysis of the p50 subunit of NF-kappa B and inhibition of NF-kappa B activity by trans-dominant p50 mutants. J. Virol. 67, 288–293 (1993).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Coleman, T.A., Kunsch, C., Maher, M., Ruben, S.M. & Rosen, C.A. Acquisition of NFKB1-selective DNA binding by substitution of four amino acid residues from NFKB1 into RelA. Mol. Cell. Biol. 13, 3850–3859 (1993).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Toledano, M.B., Ghosh, D., Trinh, F. & Leonard, W.J. N-terminal DNA-binding domains contribute to differential DNA-binding specificities of NF-kappa B p50 and p65. Mol. Cell. Biol. 13, 852–860 (1993).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Kumar, S., Rabson, A.B. & Gelinas, C., RxxRxRxxC motif conserved in all Rel/kappa B proteins is essential for the DNA-binding activity and redox regulation of the v-Rel oncoprotein. Mol Cell Biol 12, 3094–3106 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Ledebur, H.C. & Parks, T.P. Transcriptional regulation of the intercellular adhesion molecule-1 gene by inflammatory cytokines in human endothelial cells. Essential roles of a variant NF-kappa B site and p65 homodimers. J Biol Chem 270, 933–943 (1995).

    Article  CAS  PubMed  Google Scholar 

  18. Sica, A. et al. The c-rel protooncogene product c-Rel but not NF-kappa B binds to the intronic region of the human interferon-gamma gene at a site related to an interferon-stimulable response element. Proc Natl Acad Sci U S A 89, 1740–1744 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Collart, M.A., Baeuerle, P. & Vassalli, P. Regulation of tumor necrosis factor alpha transcription in macrophages: involvement of four kappa B-like motifs and of constitutive and inducible forms of NF-kappa B. Mol. Cell. Biol. 10, 1498–1506 (1990).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Hansen, S.K., Guerrini, L. & Blasi, F. Differential DNA sequence specificity and regulation of HIV-1 enhancer activity by cRel-RelA transcription factor. J. Biol. Chem. 269, 22230–22237 (1994).

    CAS  PubMed  Google Scholar 

  21. Whitley, M.Z., Thanos, D., Read, M.A., Maniatis, T. & Collins, T. A striking similarity in the organization of the E-selectin and beta interferon gene promoters. Mol. Cell. Biol. 14, 6464–6475 (1994).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Xu, X. & Gelinas, C. A mutant Rel-homology domain promotes transcription by p50/NFkappaB1. Oncogene 14, 1521–1530 (1997).

    Article  CAS  PubMed  Google Scholar 

  23. Fujita, T., Nolan, G.P., Ghosh, S. & Baltimore, D. Independent modes of transcriptional activation by the p50 and p65 subunits of NF-kappa B. Genes Dev 6, 775–787 (1992).

    Article  CAS  PubMed  Google Scholar 

  24. LeClair, K.P., Blanar, M.A. & Sharp, P.A. The p50 subunit of NF-kappa B associates with the NF-IL6 transcription factor. Proc Natl Acad Sci U S A 89, 8145–8149 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Stein, B. et al. Cross-coupling of the NF-kappa B p65 and Fos/Jun transcription factors produces potentiated biological function. EmboJ 12, 3879–3891 (1993).

    Article  CAS  Google Scholar 

  26. Sif, S. & Gilmore, T.D. Interaction of the v-Rel oncoprotein with cellular transcription factor Sp1. J. Virol. 68, 7131–7138 (1994).

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Lefstin, J.A., Thomas, J.R. & Yamamoto, K.R. Influence of a steroid receptor DNA-binding domain on transcriptional regulatory functions. Genes Dev 8, 2842–2856 (1994).

    Article  CAS  PubMed  Google Scholar 

  28. Walker, S., Hayes, S. & O′Hare, P.Site-specific conformational alteration of the Oct-1 POU domain-DNA complex as the basis for differential recognition by Vmw65 (VP16). Cell 79, 841–852 (1994).

    Article  CAS  PubMed  Google Scholar 

  29. Yin, M.J. & Gaynor, R.B. HTLV-1 21 bp repeat sequences facilitate stable association between Tax and CREB to increase CREB binding affinity. J Mol Biol 264, 20–31 (1996).

    Article  CAS  PubMed  Google Scholar 

  30. Otwinoswki, Z., Oscillian Data Reduction Programme, in Proceedings of the CCP4 Study Weekend: “Data Collection and Processing,” 29–30 January 1993, (Compiled by: L. Sawyer, N. Issacs S. Bailey) 56–62 (SERC Daresbury Laboratory, England)

  31. Brunger, A.T. X-PLOR Version 3.1: A system for x-ray crystallography and NMR, 382 (Yale University Press, New–Heaven, 1992

    Google Scholar 

  32. Collaborative Computing Project, No. 4 (1994). The CCP4 suite: programs for protein crystallogy. Acta Cryst. D50, 760–763.

  33. Evans, S.V. SETOR: hardware-lighted three-dimensional solid model representations of macromolecules.J. Mol. Graph. 11, 134–138 (1993).

    Article  CAS  PubMed  Google Scholar 

  34. Lavery, R. & Sklenar, H. The definition of generalized helicoidal parameters and of axis curvature for irregular nucleic acids. J. Biomol. Struct. Dyn. 6, 63–91 (1988).

    Article  CAS  PubMed  Google Scholar 

  35. Himes, S.R., Coles, L.S., Katsikeros, R., Lang, R.K. & Shannon, M.F. HTLV-1 tax activation of the GM-CSF and G-CSF promoters requires the interaction of NF-kB with other transcription factor families. Oncogene 8, 3189–3197 (1993).

    CAS  PubMed  Google Scholar 

  36. Himes, S.R., Coles, L.S., Reeves, R. & Shannon, M.F. High mobility group protein I(Y) is required for function and for c-Rel binding to CD28 response elements within the GM-CSF and IL-2 promoters. Immunity 5, 479–489 (1996).

    Article  CAS  PubMed  Google Scholar 

  37. Ghosh, P., Tan, T.H., Rice, N.R., Sica, A. & Young, H.A. The interleukin 2 CD28-responsive complex contains at least three members of the NF kappa B family: c-Rel, p50, and p65. Proc Natl Acad Sci U S A 90, 1696–1700 (1993).

    Article  CAS  Google Scholar 

  38. Parry, G.C. & Mackman, N. A set of inducible genes expressed by activated human monocytic and endothelial cells contain kappa B-like sites that specifically bind c-Rel-p65 heterodimers.J Biol Chem 269, 20823–20825 (1994).

    CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Chemistry and Biochemistry, University of California San Diego, 9500 Gilman Drive, La Jolla, California, 92093-0359, USA

    Yong-Qing Chen & Gourisankar Ghosh

  2. Section of Immunobiology & Department of Molecular Biochemistry & Biophysics, Yale University and the Howard Hughes Medical Institute, 295 Congress Avenue, New Haven, Connecticut, 06510, USA

    Sankar Ghosh

Authors
  1. Yong-Qing Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sankar Ghosh
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Gourisankar Ghosh
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Gourisankar Ghosh.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Chen, YQ., Ghosh, S. & Ghosh, G. A novel DNA recognition mode by the NF-κB p65 homodimer. Nat Struct Mol Biol 5, 67–73 (1998). https://doi.org/10.1038/nsb0198-67

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsb0198-67

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