Loss of p53 enhances catalytic activity of IKKβ through O-linked β-N-acetyl glucosamine modification

The IκB kinase (IKK)-NF-κB pathway plays a critical role in oncogenesis. Recently, we have shown that p53 regulates glucose metabolism through the IKK-NF-κB pathway and that, in the absence of p53, the positive feedback loop between IKK-NF-κB and glycolysis has an integral role in oncogene-induced cell transformation. Here, we demonstrate that IKKβ, a component of the IKK complex, was constitutively modified with O-linked β-N-acetyl glucosamine (O-GlcNAc) in both p53-deficient mouse embryonic fibroblasts (MEFs) and transformed human fibroblasts. In p53-deficient cells, the O-GlcNAcylated IKKβ and the activating phosphorylation of IKK were decreased by p65/NF-κB knockdown or glucose depletion. We also found that high glucose induced the O-GlcNAcylation of IKKβ and sustained the TNFα-dependent IKKβ activity. Moreover, the O-GlcNAcase inhibitor streptozotocin intensified O-GlcNAcylation and concomitant activating phosphorylation of IKKβ. Mutational analysis revealed that O-GlcNAcylation of IKKβ occurred at Ser 733 in the C-terminal domain, which was identified as an inactivating phosphorylation site, suggesting that IKKβ O-GlcNAcylation regulates its catalytic activity. Taken together, we propose a novel mechanism for the enhancement of NF-κB activity by loss of p53, which evokes positive feedback regulation from enhanced glucose metabolism to IKK in oncogenesis.