Nedd8 modification of Cul-1 activates SCF(β(TrCP))-dependent ubiquitination of IκBα

Margaret A. Read, James E. Brownell, Tatiana B. Gladysheva, Maria Hottelet, Lana A. Parent, Michael B. Coggins, Jacqueline W. Pierce, Vladimir N. Podust, Rong Shu Luo, Vincent Chau, Vito J. Palombella

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Abstract

Regulation of NF-κB occurs through phosphorylation-dependent ubiquitination of IκBα, which is degraded by the 26S proteasome. Recent studies have shown that ubiquitination of IκBα is carried out by a ubiquitin-ligase enzyme complex called SCF(β(TrCP)). Here we show that Nedd8 modification of the Cul-1 component of SCF(β(TrCP)) is important for function of SCF(β(TrCP)) in ubiquitination of IκBα. In cells, Nedd8- conjugated Cul-1 was complexed with two substrates of SCF(β(TrCP)), phosphorylated IκBα and β-catenin, indicating that Nedd8-Cul-1 conjugates are part of SCF(β(TrCP)) in vivo. Although only a minute fraction of total cellular Cul-1 is modified by Nedd8, the Cul-1 associated with ectopically expressed βTrCP was highly enriched for the Nedd8-conjugated form. Moreover, optimal ubiquitination of IκBα required Nedd8 and the Nedd8-conjugating enzyme, Ubc12. The site of Nedd8 ligation to Cul-1 is essential, as SCF(β(TrCP)) containing a K720R mutant of Cul-1 only weakly supported IκBα ubiquitination compared to SCF(β(TrCP)) containing WT Cul-1, suggesting that the Nedd8 ligation of Cul-1 affects the ubiquitination activity of SCF(β(TrCP)). These observations provide a functional link between the highly related ubiquitin and Nedd8 pathways of protein modification and show how they operate together to selectively target the signal-dependent degradation of IκBα.

Original languageEnglish (US)
Pages (from-to)2326-2333
Number of pages8
JournalMolecular and cellular biology
Volume20
Issue number7
DOIs
StatePublished - Apr 1 2000

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All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cell Biology

Cite this

Read, M. A., Brownell, J. E., Gladysheva, T. B., Hottelet, M., Parent, L. A., Coggins, M. B., Pierce, J. W., Podust, V. N., Luo, R. S., Chau, V., & Palombella, V. J. (2000). Nedd8 modification of Cul-1 activates SCF(β(TrCP))-dependent ubiquitination of IκBα. Molecular and cellular biology, 20(7), 2326-2333. https://doi.org/10.1128/MCB.20.7.2326-2333.2000