Transient nuclear factor kappa B (NF-kappa B) activation stimulated by interleukin-1 beta may be partly dependent on proteasome activity, but not phosphorylation and ubiquitination of the I kappa B alpha molecule, in C6 glioma cells - Regulation of NF-kappa B linked to chemokine production

We previously reported that several stresses can induce cytokine-induced ne utrophil chemoattractant expression in a nuclear factor kappa B (NF-kappa B )-dependent manner. In this study, we focused further on the regulation of NF-kappa B, The activation of NF-kappa B and the subsequent cytokine-induce d neutrophil chemoattractant induction in response to interleukin-1 beta (I L-1 beta) were inhibited by proteasome inhibitors, MG132 and proteasome inh ibitor I. Translocation of NF-kappa B into nuclei occurs by the phosphoryla tion, multi-ubiquitination, and degradation of I kappa B alpha, a regulator y protein of NF-kappa B, Nascent I kappa B alpha began to degrade 5 min aft er treatment with IL-1 beta and disappeared completely after 15 min. Howeve r, I kappa B alpha returned to basal levels after 45-60 min. Interestingly, resynthesized I kappa B alpha was already phosphorylated at Ser-32, These results suggest that 1) the upstream signals are still activated, although the translocation of NF-kappa B peaks at 15 min; and 2) the regulated prote in(s) acts downstream of I kappa B alpha phosphorylation, Western blotting showed that the resynthesized and phosphorylated I kappa B molecules were a lso upward-shifted by multi-ubiquitination in response to IL-1 beta treatme nt. On the other hand, ATP-dependent Leu-Leu-Val-Tyr cleaving activity tran siently increased, peaked at 15 min, and then decreased to basal levels at 60 min. Furthermore, the cytosolic fraction that was stimulated by IL-1 bet a for 15 min, but not for 0 and 60 min, could degrade phosphorylated and mu lti-ubiquitinated I kappa B alpha, These results indicate that the transien t translocation of NF-kappa B in response to IL-1 beta may be partly depend ent on transient proteasome activation.