Regulation of NF-κB activity by competition between RelA acetylation and ubiquitination

The NF-κB transcription factor plays essential roles in inflammation and oncogenesis. Its ubiquitous RelA subunit is regulated by several post-translational modifications (PTMs) including phosphorylation, ubiquitination, and acetylation. Ubiquitination promotes the termination of RelA-dependent transcription, but its regulation is incompletely understood. Through mass spectrometry analysis of ubiquitinated RelA, we identified 7 lysines that were attached to degradative and non-degradative forms of polyubiquitin. Interestingly, lysines targeted for acetylation were among the residues identified as ubiquitin acceptor sites. Mutation of these particular sites resulted in decreased polyubiquitination. Acetylation and ubiquitination were found to inhibit each other consistent with their use of overlapping sites. Reconstitution of rela(−/−) fibroblasts with wild-type and mutant forms of RelA revealed that modifications at these residues can play activating and inhibitory functions depending on the target gene context. Altogether, this study elucidates that ubiquitination and acetylation can modulate each other and regulate nuclear NF-κB function in a gene-specific manner.