The carboxy-terminus of IκBα determines susceptibility to degradation by the catalytic core of the proteasome

Abstract

The Rel/NF-κB family of transcription factors controls the expression of a wide variety of genes that are implicated in immune and inflammatory responses and cellular proliferation. Disregulation of NF-κB is associated with cellular transformation and the maintenance of a high anti-apoptotic threshold in transformed cells. NF-κB activity is in turn regulated by its sequestration in the cytoplasm by the inhibitor IκB. IκBα, the most abundant and well-characterized member of the IκB multiprotein family, is rapidly degraded in response to multiple physiologic stimuli. In the present study we show that not only the amino-terminus, but also the carboxy-terminus of IκBα contain transferable signals that must be simultaneously present in an unrelated protein to render it susceptible to activation-induced, proteasome-mediated degradation. We show here that IκBα amino-terminal modifications occur independently of the carboxy-terminus. Moreover, we present evidence indicating a critical role for the carboxy-terminal region in facilitating proteolysis by the catalytic core of the proteasome. When incubated with 20S proteasome extracted from rat liver, IκBα was quickly degraded while a deletion mutant lacking the carboxy-terminus was resistant to proteolysis. Likewise, chimeric proteins of beta-galactosidase with the IκBα carboxy-terminus were degraded in vitro independently of the presence of the IκBα amino-terminus, whereas chimeric proteins lacking the IκBα carboxy-terminus were stable. Our results identify the carboxy-terminus of IκBα as a domain critical for degradation through interaction with an as yet unidentified component of the proteasome.