Binding of nuclear factor kappa B to non-canonical consensus sites reveals its multimodal role during the early inflammatory response

Abstract

Mammalian cells have developed intricate mechanisms to interpret, integrate, and respond to extra-cellular stimuli. For example, tumor necrosis factor (TNF) rapidly activates proinflammatory genes, but our understanding of how this occurs against the ongoing transcriptional program of the cell is far from complete. We monitor the early phase of this cascade at high spatio-temporal resolution in TNFα-stimulated human endothelial cells. The main driver of the response, NF-κB, hijacks the regulatory machinery by binding active enhancers already-interacting with gene promoters. Notably, a large fraction of these enhancers do not encode the canonical NF-κB recognition motif. Using a combination of genomics tools we find that site selection is a key determinant of NF-κΒ participation in both transcriptional activation and repression, and we exemplify the repressive NF-κΒ function via its synergy with the JDP2 co-repressor. Finally, detailed analysis of a 3-Mbp locus using both targeted chromatin conformation capture and genome editing uncovers how newly-introduced NF-κΒ exploits pre-existing chromatin looping to exert its multimodal role. This work dissects the involvement of topology in the function of cis-regulatory elements during acute transcriptional responses, where primary sequence and higher-order structure constitute the regulatory context leading to either gene activation or repression.