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

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

Author information

1
Institute of Biochemistry, Member of the German Center for Lung Research, Justus-Liebig-University, Giessen, Germany.
2
Institute of Molecular Biology and Tumor Research (IMT), Philipps University Marburg, Marburg, Germany.
3
Rudolf-Buchheim-Institute of Pharmacology, Member of the German Center for Lung Research, Justus-Liebig-University, Giessen, Germany; and.
4
Genomics Core Facility-Institute of Molecular Oncology, Philipps University Marburg, Marburg, Germany.

Abstract

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

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