Regulation of Constitutive p50/c-Rel Activity via Proteasome Inhibitor-Resistant IκBα Degradation in B Cells

Abstract

Constitutive NF-κB activity has emerged as an important cell survival component of physiological and pathological processes, including B-cell development. In B cells, constitutive NF-κB activity includes p50/c-Rel and p52/RelB heterodimers, both of which are critical for proper B-cell development. We previously reported that WEHI-231 B cells maintain constitutive p50/c-Rel activity via selective degradation of IκBα that is mediated by a proteasome inhibitor-resistant, now termed PIR, pathway. Here, we examined the mechanisms of PIR degradation by comparing it to the canonical pathway that involves IκB kinase-dependent phosphorylation and β-TrCP-dependent ubiquitylation of the N-terminal signal response domain of IκBα. We found a distinct consensus sequence within this domain of IκBα for PIR degradation. Chimeric analyses of IκBα and IκBβ further revealed that the ankyrin repeats of IκBα, but not IκBβ, contained information necessary for PIR degradation, thereby explaining IκBα selectivity for the PIR pathway. Moreover, we found that PIR degradation of IκBα and constitutive p50/c-Rel activity in primary murine B cells were maintained in a manner different from B-cell-activating-factor-dependent p52/RelB regulation. Thus, our findings suggest that nonconventional PIR degradation of IκBα may play a physiological role in the development of B cells in vivo.

SUPPLEMENTAL MATERIAL

We thank Kei Tashiro for providing K3R- and K5R-IκBα clones, Zhijian Chen for providing c-myc-ΔF-β-TrCP, Bradley Seufzer for technical support, Cindy Chan for the construction of some mutant IκBαs, and the Miyamoto lab members for helpful discussions.

This work was supported in part by funding from an American Heart Association Predoctoral fellowship to S.O.; NIH training grant (T32GM07215) for S.O., S.D.S., and I.J.A.; and NIH R01-CA77474, NIH R01-CA81065, and the Shaw Scientist Award from the Milwaukee Foundation to S.M.