The NEDD8 pathway is essential for SCfβ-TrCP-mediated ubiquitination and processing of the NF-κB precursor p105

Ruthie E. Amir, Kazuhiro Iwai, Aaron Ciechanover

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Abstract

The p50 subunit of NF-κB is generated by limited processing of the precursor p105. IκB kinase-mediated phosphorylation of the C-terminal domain of p105 recruits the SCFβ-TrCP ubiquitin ligase, resulting in rapid ubiquitination and subsequent processing/degradation of p105. NEDD8 is known to activate SCF ligases following modification of their cullin component. Here we show that NEDDylation is required for conjugation and processing of p105 by SCFβ-TrCP following phosphorylation of the molecule. In a crude extract, a dominant negative E2 enzyme, UBC12, inhibits both conjugation and processing of p105, and inhibition is alleviated by an excess of WT- UBC12. In a reconstituted cell-free system, ubiquitination of p105 was stimulated only in the presence of all three components of the NEDD8 pathway, E1, E2, and NEDD8. A Cul-1 mutant that cannot be NEDDylated could not stimulate ubiquitination and processing of p105. Similar findings were observed also in cells. It should be noted that NEDDylation is required only for the stimulated but not for basal processing of p105. Although the mechanisms that underlie processing of p105 are largely obscure, it is clear that NEDDylation and the coordinated activity of SCFβ-Trcp on both p105 and IκBα serve as an important regulatory mechanism controlling NF-κB activity.

Original languageEnglish
Pages (from-to)23253-23259
Number of pages7
JournalJournal of Biological Chemistry
Volume277
Issue number26
DOIs
StatePublished - 28 Jun 2002

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Ubiquitination
SKP Cullin F-Box Protein Ligases
Processing
Cullin Proteins
Phosphorylation
Cell-Free System
Ligases
Complex Mixtures
Phosphotransferases
Enzymes
Ubiquitin
Degradation
Molecules

Cite this

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title = "The NEDD8 pathway is essential for SCfβ-TrCP-mediated ubiquitination and processing of the NF-κB precursor p105",
abstract = "The p50 subunit of NF-κB is generated by limited processing of the precursor p105. IκB kinase-mediated phosphorylation of the C-terminal domain of p105 recruits the SCFβ-TrCP ubiquitin ligase, resulting in rapid ubiquitination and subsequent processing/degradation of p105. NEDD8 is known to activate SCF ligases following modification of their cullin component. Here we show that NEDDylation is required for conjugation and processing of p105 by SCFβ-TrCP following phosphorylation of the molecule. In a crude extract, a dominant negative E2 enzyme, UBC12, inhibits both conjugation and processing of p105, and inhibition is alleviated by an excess of WT- UBC12. In a reconstituted cell-free system, ubiquitination of p105 was stimulated only in the presence of all three components of the NEDD8 pathway, E1, E2, and NEDD8. A Cul-1 mutant that cannot be NEDDylated could not stimulate ubiquitination and processing of p105. Similar findings were observed also in cells. It should be noted that NEDDylation is required only for the stimulated but not for basal processing of p105. Although the mechanisms that underlie processing of p105 are largely obscure, it is clear that NEDDylation and the coordinated activity of SCFβ-Trcp on both p105 and IκBα serve as an important regulatory mechanism controlling NF-κB activity.",
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The NEDD8 pathway is essential for SCfβ-TrCP-mediated ubiquitination and processing of the NF-κB precursor p105. / Amir, Ruthie E.; Iwai, Kazuhiro; Ciechanover, Aaron.

In: Journal of Biological Chemistry, Vol. 277, No. 26, 28.06.2002, p. 23253-23259.

Research output: Contribution to journalArticleResearchpeer-review

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T1 - The NEDD8 pathway is essential for SCfβ-TrCP-mediated ubiquitination and processing of the NF-κB precursor p105

AU - Amir, Ruthie E.

AU - Iwai, Kazuhiro

AU - Ciechanover, Aaron

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