A mechanistic insight into a proteasome-independent constitutive inhibitor κBα (IκBα) degradation and nuclear factor κB (NF-κB) activation pathway in WEHI-231 B-cells

Abstract

Inducible activation of the transcription factor NF-κB (nuclear factor κB) is classically mediated by proteasomal degradation of its associated inhibitors, IκBα (inhibitory κBα) and IκBβ. However, certain B-lymphocytes maintain constitutively nuclear NF-κB activity (a p50-c-Rel heterodimer) which is resistant to inhibition by proteasome inhibitors. This activity in the WEHI-231 B-cell line is associated with continual and preferential degradation of IκBα, which is also unaffected by proteasome inhibitors. Pharmacological studies indicated that there was a correlation between inhibition of IκBa degradation and constitutive p50-c-Rel activity. Domain analysis of IκBα by deletion mutagenesis demonstrated that an N-terminal 36-amino-acid sequence of IκBα represented an instability determinant for constitutive degradation. Moreover, domain grafting studies indicated that this sequence was sufficient to cause IκBβ, but not chloramphenicol acetyltransferase, to be rapidly degraded in WEHI-231 B-cells. However, this sequence was insufficient to target IκBβ to the non-proteasome degradation pathway, suggesting that there was an additional cis-element(s) in IκBα that was required for complete targeting. Nevertheless, the NF-κB pool associated with IκBβ now became constitutively active by virtue of IκBβ instability in these cells. These findings further support the notion that IκB instability governs the maintenance of constitutive p50-c-Rel activity in certain B-cells via a unique degradation pathway.