IκBα ubiquitination is catalyzed by an SCF-like complex containing Skp1, cullin-1, and two F-box/WD40-repeat proteins, βTrCP1 and βTrCP2

Destruction of the transcriptional inhibitor I kappa B by the ubiquitin (Ub) system is required for signal-dependent activation of the multifunctional transcriptional factor NF-kappa B, but details of this ubiquitination are largely unknown, We report here that the I kappa B alpha-ubiquitin ligase (I kappa B alpha-E3) is an SCF-like complex containing Skp1, cullin-1, and two homologous F-box/WD40-repeat proteins, beta TrCP1 and beta TrCP2. Intriguingly, all these components are cooperatively recruited to bind to a phosphorylated I kappa B alpha (pI kappa B alpha) produced by tumor necrosis factor-alpha (TNF-alpha) stimulation. I kappa B alpha-E3 bound to pI kappa B alpha catalyzed in vitro ubiquitination of pI kappa B alpha in the presence of ATP, Ub, and E1-activating and E2-conjugating enzymes. Forced expression of beta TrCP1 and beta TrCP2 resulted in dramatic augmentation of the in vitro polyubiquitination activity of I kappa B alpha-E3. These results indicate that the long-sought I kappa B alpha-E3 is an SCF-like complex consisting of multiple proteins which are coordinately assembled during phosphorylation of I kappa B alpha in response to external signals. (C) 1999 Academic Press.