Tumor necrosis factor α-induced phosphorylation of RelA/p65 on Ser529 is controlled by casein kinase II

Nuclear factor kappaB (NF-kappaB)/Rel transcription factors are key regulators of a variety of genes involved in immune and inflammatory responses, growth, differentiation, apoptosis, and development. In unstimulated cells, NF-kappaB/Rel proteins are sequestered in the cytoplasm by I kappaB inhibitor proteins. Many extracellular stimuli, such as tumor necrosis factor alpha (TNF alpha), cause rapid phosphorylation of I kappaB at N-terminal serine residues leading to ubiquitination and degradation of the inhibitor. Subsequently, NF-kappaB proteins translocate to the nucleus and activate gene expression through KB response elements. TNF alpha, as well as certain other stimuli, also induces the phosphorylation of the NP-kappaB proteins. Previously, we have shown that TNF alpha induces RelA/p65 phosphorylation at serine 529 and that this inducible phosphorylation increases NF-kappaB transcriptional activity on an exogenously supplied reporter (1). In this report, we demonstrate that casein kinase II (CKII) interacts with p65 in vivo and can phosphorylate p65 at serine 529 in vitro. A CKII inhibitor (PD144795) inhibited TNF alpha -induced p65 phosphorylation in vivo. Furthermore, our results indicate that the association between I kappaB alpha and p65 inhibits p65 phosphorylation by CKII and that degradation of I kappaB alpha allows CKII to phosphorylate p65 to increase NF-kappaB transactivation potential. These data may explain the ability of CKII to modulate cell growth and demonstrate a mechanism whereby CKII can function in an inducible manner.