Investigating the role of NF-κB p50 serine 80 phosphorylation in the regulation of inflammation

QH301 Biology;QH345 Biochemistry;QR180 Immunology

Smith, Emma Louise ; Biotechnology and Biological Sciences Research Council (BBSRC) ; Ruaidhri, Carmody

University:University of Glasgow

Department:Institute of Infection Immunity and Inflammation

关键词: NF-κB, p50, transcription factor, transcriptional regulation, phosphorylation, CRISPR/Cas9, DNA binding.;   

Others  :  http://theses.gla.ac.uk/38985/1/2018smithphd.pdf

来源: University of Glasgow

【 摘 要 】

NF-κB is a key transcription factor involved in the regulation of inflammation. The transcriptional activity of NF-κB is regulated by a number of post-translational modifications, including phosphorylation. Phosphorylation of NF-κB subunits may regulate transcription in a gene selective manner. The NF-κB p50 subunit is an important dual regulator of inflammatory responses, which can either promote or repress gene expression depending on the formation of heterodimer or homodimer complexes, respectively. Although p50 is a critical regulator of the immune system, phosphorylation of this subunit is largely understudied. In this thesis, the role of phosphorylation of NF-κB p50 on serine 80 (S80) in regulating transcriptional responses is investigated, using two human NFKB1S80A knock-in cell lines generated by CRISPR/Cas9 genome editing techniques. Transcriptomic analyses reveal a critical role for S80 in selectively regulating the expression of a subset of NF-κB target genes in response to TNFα and LPS, including pro-inflammatory chemokines and cytokines. DNA binding analyses demonstrate that S80 regulates the binding of p50 to NF-κB binding sites in a sequence-specific manner. Specifically, phosphorylation of S80 reduces the affinity of p50 for κB sites that have an adenine at the -1 position. These data indicate that p50 S80 phosphorylation predominantly regulates transcription through the p50:p65 heterodimer, where S80 phosphorylation acts in trans to limit the NF-κB mediated transcription of pro-inflammatory genes. This advances our understanding of the regulation of transcriptional programmes by the NF-ĸB p50 subunit.