Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics. Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics

NFkB RelA is the potent transcriptional activator of inflammatory response genes. We stringently defined a list of direct RelA target genes by integrating physical (ChIPseq) and functional (RNAseq in knockouts) datasets. We then dissected each gene's regulatory strategy by testing RelA variants in a novel primary-cell genetic complementation assay. All endogenous target genes required that RelA makes DNA-base-specific contacts, and none could be activated by the DNA binding domain alone. However, endogenous target genes differed widely in how they employ the two transactivation domains. Through model-aided analysis of the dynamic timecourse data we reveal gene-specific synergy and redundancy of TA1 and TA2. Given that post-translational modifications control TA1 activity and intrinsic affinity for coactivators determines TA2 activity, the differential TA logics suggests context-dependent vs. context-independent control of endogenous RelA-target genes. While some inflammatory initiators appear to require co-stimulatory TA1 activation, inflammatory resolvers are a part of the NFkB RelA core response. Overall design: We performed ChIP-seq with antibody specific for NF-kappaB RelA subunit from retroviral complementation of RelA-deficient primary mouse embryonic fibroblasts with RelA (WT/mutants) stimulated with TNF