Negative regulation of NF_-kB action by Set9-mediated lysine methylation of the relA subunit

Proper regulation of NF-_-kB activity is critical to maintain and balance the inflammatory response. Inactivation of the NF-_-kB complex relies in part on the proteasome-mediated degradation of promoter-bound NF-_-kB, but the detailed molecular mechanism initiating this process remains elusive. Here, we show that the methylation of the RelA subunit of NF-_-kB has an important function in this process. Lysine methyltransferase Set9 physically associates with RelA in vitro and in vivo in response to TNF-α stimulation. Mutational and mass spectrometric analyses reveal that RelA is monomethylated by Set9 at lysine residues 314 and 315 in vitro and in vivo. Methylation of RelA inhibits NF-_-kB action by inducing the proteasome-mediated degradation of promoter-associated RelA. Depletion of Set9 by siRNA or mutation of the RelA methylation sites prolongs DNA binding of NF-_-kB and enhances TNF-α-induced expression of NF-_-kB target genes. Together, these findings unveil a novel mechanism by which methylation of RelA dictates the turnover of NF- _-kB and controls the NF-_-kB-mediated inflammatory response.