NF-κB p65 dimerization and DNA-binding is important for inflammatory gene expression

FASEB J. 2019 Mar;33(3):4188-4202. doi: 10.1096/fj.201801638R. Epub 2018 Dec 7.


Increasing evidence shows that many transcription factors execute important biologic functions independent from their DNA-binding capacity. The NF-κB p65 (RELA) subunit is a central regulator of innate immunity. Here, we investigated the relative functional contribution of p65 DNA-binding and dimerization in p65-deficient human and murine cells reconstituted with single amino acid mutants preventing either DNA-binding (p65 E/I) or dimerization (p65 FL/DD). DNA-binding of p65 was required for RelB-dependent stabilization of the NF-κB p100 protein. The antiapoptotic function of p65 and expression of the majority of TNF-α-induced genes were dependent on p65's ability to bind DNA and to dimerize. Chromatin immunoprecipitation with massively parallel DNA sequencing experiments revealed that impaired DNA-binding and dimerization strongly diminish the chromatin association of p65. However, there were also p65-independent TNF-α-inducible genes and a subgroup of p65 binding sites still allowed some residual chromatin association of the mutants. These sites were enriched in activator protein 1 (AP-1) binding motifs and showed increased chromatin accessibility and basal transcription. This suggests a mechanism of assisted p65 chromatin association that can be in part facilitated by chromatin priming and cooperativity with other transcription factors such as AP-1.-Riedlinger, T., Liefke, R., Meier-Soelch, J., Jurida, L., Nist, A., Stiewe, T., Kracht, M., Schmitz, M. L. NF-κB p65 dimerization and DNA-binding is important for inflammatory gene expression.

Keywords: chromatin; inflammation; transcription.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Binding Sites / genetics
  • Cell Line, Tumor
  • Chromatin / genetics
  • Chromatin Assembly and Disassembly / genetics
  • DNA / genetics*
  • DNA-Binding Proteins / genetics*
  • Dimerization
  • Gene Expression / genetics*
  • HeLa Cells
  • Humans
  • Inflammation / genetics*
  • Mice
  • Protein Binding / genetics
  • Transcription Factor AP-1 / genetics
  • Transcription Factor RelA / genetics*
  • Transcription Factor RelB / genetics


  • Chromatin
  • DNA-Binding Proteins
  • Transcription Factor AP-1
  • Transcription Factor RelA
  • Transcription Factor RelB
  • DNA