Crystal structure of p50/p65 heterodimer of transcription factor NF-κB bound to DNA

The NF-κB p50/p65 heterodimer is the classical member of the Relfamily of transcription factors which regulate diverse cellular functions such as immune response, cell growth, and development. Other mammalian Rel family members, including theproteins p52, proto-oncoprotein c-Rel, and RelB, all have amino-terminal Rel-homology regions (RHRs). The RHR is responsible for the dimerization, DNA binding and cytosolic localization of these proteins by virtue of complex formation with inhibitor κB proteins. Signal-induced removal of κB inhibitors allows translocation of dimers to the cell nucleus and transcriptional regulation of κB DNA-containing genes. NF-κB specifically recognizes κB DNA elements,, with a consensus sequence of 5′-GGGRNYYYCC-3′ (R is an unspecified purine; Y is an unspecified pyrimidine; and N is any nucleotide). Here we report the crystal structure at 2.9 Å resolution of the p50/p65 heterodimer bound to the κB DNA of the intronic enhancer of the immunoglobulin light-chain gene. Our structure reveals a 5-base-pair 5′ subsite for p50, and a 4-base-pair 3′ subsite for p65. This structure indicates why the p50/p65 heterodimer interface is stronger than that of either homodimer. A comparison of this structure with those of other Rel dimers reveals that both subunits adopt variable conformations in a DNA-sequence-dependent manner. Our results explain the different behaviour of the p50/p65 heterodimer with heterologous promoters.

[1]  D. Giedroc,et al.  The role of protein-protein interactions in the assembly of the presynaptic filament for T4 homologous recombination. , 1993, The Journal of biological chemistry.

[2]  D. Baltimore,et al.  I kappa B: a specific inhibitor of the NF-kappa B transcription factor. , 1988, Science.

[3]  S. Harrison,et al.  Structure of the NF-kappa B p50 homodimer bound to DNA. , 1995, Nature.

[4]  P. Sigler,et al.  Structure of NF-κB p50 homodimer bound to a κB site , 1998, Nature.

[5]  W. Leonard,et al.  N-terminal DNA-binding domains contribute to differential DNA-binding specificities of NF-kappa B p50 and p65 , 1993, Molecular and cellular biology.

[6]  L. Guerrini,et al.  Differential DNA sequence specificity and regulation of HIV-1 enhancer activity by cRel-RelA transcription factor. , 1994, The Journal of biological chemistry.

[7]  S V Evans,et al.  SETOR: hardware-lighted three-dimensional solid model representations of macromolecules. , 1993, Journal of molecular graphics.

[8]  David Baltimore,et al.  Inducibility of κ immunoglobulin enhancer-binding protein NF-κB by a posttranslational mechanism , 1986, Cell.

[9]  S. L. Cross,et al.  Functionally distinct NF-kappa B binding sites in the immunoglobulin kappa and IL-2 receptor alpha chain genes. , 1989, Science.

[10]  P. Baeuerle,et al.  The 65-kD subunit of NF-kappa B is a receptor for I kappa B and a modulator of DNA-binding specificity. , 1990, Genes & development.

[11]  Axel T. Brunger,et al.  X-PLOR Version 3.1: A System for X-ray Crystallography and NMR , 1992 .

[12]  D. Baltimore,et al.  Inducibility of kappa immunoglobulin enhancer-binding protein Nf-kappa B by a posttranslational mechanism. , 1986, Cell.

[13]  Dimitris Thanos,et al.  The High Mobility Group protein HMG I(Y) is required for NF-κB-dependent virus induction of the human IFN-β gene , 1992, Cell.

[14]  G. Nolan,et al.  DNA binding and IκB inhibition of the cloned p65 subunit of NF-κB, a rel-related polypeptide , 1991, Cell.

[15]  G. Franzoso,et al.  Structure, regulation and function of NF-kappa B. , 1994, Annual review of cell biology.

[16]  R. Huber,et al.  Accurate Bond and Angle Parameters for X-ray Protein Structure Refinement , 1991 .

[17]  C. Cambillau,et al.  TOM: a FRODO subpackage for protein-ligand fitting with interactive energy minimization , 1988 .

[18]  U Zabel,et al.  DNA binding of purified transcription factor NF-kappa B. Affinity, specificity, Zn2+ dependence, and differential half-site recognition. , 1991, The Journal of biological chemistry.

[19]  T. Maniatis,et al.  Signal-induced site-specific phosphorylation targets I kappa B alpha to the ubiquitin-proteasome pathway. , 1995, Genes & development.

[20]  D. Baltimore,et al.  NF-kappa B: a pleiotropic mediator of inducible and tissue-specific gene control. , 1989, Cell.

[21]  G. Nolan,et al.  Cloning of the p50 DNA binding subunit of NF-κB: Homology to rel and dorsal , 1990, Cell.

[22]  Dimitris Thanos,et al.  Reversal of intrinsic DNA bends in the IFNβ gene enhancer by transcription factors and the architectural protein HMG I(Y) , 1995, Cell.

[23]  Juli D. Klemm,et al.  Crystal structure of the Oct-1 POU domain bound to an octamer site: DNA recognition with tethered DNA-binding modules , 1994, Cell.

[24]  R Lavery,et al.  The definition of generalized helicoidal parameters and of axis curvature for irregular nucleic acids. , 1988, Journal of biomolecular structure & dynamics.

[25]  A. Israël,et al.  The DNA binding subunit of NF-κB is identical to factor KBF1 and homologous to the rel oncogene product , 1990, Cell.

[26]  J. Thornton,et al.  PROCHECK: a program to check the stereochemical quality of protein structures , 1993 .

[27]  J. Zou,et al.  Improved methods for building protein models in electron density maps and the location of errors in these models. , 1991, Acta crystallographica. Section A, Foundations of crystallography.

[28]  T. Maniatis,et al.  A striking similarity in the organization of the E-selectin and beta interferon gene promoters , 1994, Molecular and cellular biology.

[29]  P. Baeuerle,et al.  NF‐kappa B contacts DNA by a heterodimer of the p50 and p65 subunit. , 1991, The EMBO journal.

[30]  S. Ruben,et al.  Isolation of a rel-related human cDNA that potentially encodes the 65-kD subunit of NF-kappa B. , 1991, Science.

[31]  A. Baldwin,et al.  THE NF-κB AND IκB PROTEINS: New Discoveries and Insights , 1996 .