Identification of the receptor component of the IκBα-ubiquitin ligase

Avraham Yaron, Ada Hatzubai, Matti Davis, Iris Lavon, Sharon Amit, Anthony M. Manning, Jens S. Andersen, Matthias Mann, Frank Mercurio, Yinon Ben-Neriah*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

585 Scopus citations

Abstract

NF-κB, a ubiquitous, inducible transcription factor involved in immune, inflammatory, stress and developmental processes, is retained in a latent form in the cytoplasm of non-stimulated cells by inhibitory molecules, IκBs. Its activation is a paradigm for a signal-transduction cascade that integrates an inducible kinase and the ubiquitin-proteasome system to eliminate inhibitory regulators. Here we isolate the pIκ̄α-ubiquitin ligase (pIκBα-E3) that attaches ubiquitin, a small protein which marks other proteins for degradation by the proteasome system, to the phosphorylated NF-κB inhibitor pIκBα. Taking advantage of its high affinity to pIκBα, we isolate this ligase from HeLa cells by single-step immunoaffinity purification. Using nanoelectrospray mass spectrometry, we identify the specific component of the ligase that recognizes that pIκBα degradation motif as an F-box/WD-domain protein belonging to a recently distinguished family of β-TrCP/Slimb proteins. This component, which we denote E3RS(IκB) (pIκBα-E3 receptor subunit), binds specifically to pIκBα and promotes its in vitro ubiquitination in the presence of two other ubiquitin-system enzymes. E1 and UBC5C, one of many known E2 enzymes. An F- box-deletion mutant of E3RS(IκB), which tightly binds pIκBα but does not support its ubiquitination, acts in vivo as a dominant-negative molecule, inhibiting the degradation of pIκBα and consequently NF-κB activation. E3RS(IκB) represents a family of receptor proteins that are core components of a class of ubiquitin ligases. When these receptor components recognize their specific ligand, which is a conserved, phosphorylation-based sequence motif, they target regulatory proteins containing this motif for proteasomal degradation.

Original languageAmerican English
Pages (from-to)590-594
Number of pages5
JournalNature
Volume396
Issue number6711
DOIs
StatePublished - 10 Dec 1998

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