Signal-dependent and -independent degradation of free and NF-κB-bound IκBα

A family of inhibitory I kappa B molecules regulates the activation of the transcription factor NF-kappa B. One member of the I kappa B family, I kappa B alpha, plays a major role in the rapid signal-induced activation of NF-kappa B. I kappa B alpha itself is transcriptionally regulated by NF-kappa B allowing for a tight autoregulatory loop that is both sensitive to and rapidly influenced by NF-kappa B activating stimuli. For this pathway to remain primed both for rapid activation of NF-kappa B in the presence of signal and then to suppress NF-kappa B activation once that signal is removed, I kappa B alpha must be exquisitely regulated. The regulation of I kappa B alpha is mainly accomplished through phosphorylation, ubiquitination, and subsequent degradation. The mechanism(s) that regulate I kappa B alpha degradation needs to be able to target I kappa B alpha for degradation in both its NF-kappa B bound and free states in the cell. In this study, we utilize a full-length I kappa B alpha mutant that is unable to associate to RelA/p65. We show that the signal-induced I kappa B kinase (IKK) phosphorylation sites on I kappa B alpha can only significantly influence the regulation of signal-dependent but not signal-independent turnover of I kappa B alpha. We also demonstrate that the constitutive carboxyl-terminal casein kinase II phosphorylation sites are necessary for the proper regulation of both signal-dependent and -independent turnover of I kappa B alpha. These findings further elucidate how the phosphorylation of I kappa B alpha influences the complex regulatory mechanisms involved in maintaining a sensitive NF-kappa B pathway.