Serine 32 and serine 36 of IκBα are directly phosphorylated by protein kinase CKII in Vitro

IκBα is an inherently unstable protein which binds to and retains the ubiquitous transcription factor NFκB in the cytoplasm of resting cells. A continuous low level translocation of NFκB to the nucleus, secondary to the basal turnover of IκBα, is hypothesized to be necessary for cellular maturation, survival and, potentially, transformation. In response to cellular stimulation by inflammatory cytokines or mitogens, IκBα is rapidly degraded allowing larger pools of NFκB to translocate to the nucleus. Phosphorylation of IκBα at serine 32 (S32) and serine 36 (S36) is necessary for this stimuli-induced degradation. IKKα/β kinases and p90 rsk1 are involved in stimuli-induced targeting of one or both of these IκBα sites. Whether other kinases phosphorylate S32 and S36 directly, and if so, what function they serve in NFκB activation remains unknown. Here we present evidence of a direct phosphorylation of IκBα at both S32 and S36 by purified or immunoprecipitated protein kinase CKII (PK-CKII) and a specific in vivo association between IκBα and PK-CKII. This PK-CKII-specific kinase activity is not found within the IKKα/β-containing signalsome complex and is biochemically distinct from that of the IKKα/β kinases. The identification of an additional N-terminal IκBα kinase which is constitutively active and not significantly inducible raises numerous possibilities as to its role in cellular function.