I kappa B alpha ubiquitination is catalyzed by an SCF-like complex containing Skp1, cullin-1, and two F-box/WD40-repeat proteins, beta TrCP1 and betaTrCP2

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 t wo homologous F-box/WD40-repeat proteins, beta TrCP1 and beta TrCP2. Intrig uingly, all these components are cooperatively recruited to bind to a phosp horylated I kappa B alpha (pI kappa B alpha) produced by tumor necrosis fac tor-alpha (TNF-alpha) stimulation. I kappa B alpha-E3 bound to pI kappa B a lpha 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 v itro polyubiquitination activity of I kappa B alpha-E3. These results indic ate that the long-sought I kappa B alpha-E3 is an SCF-like complex consisti ng of multiple proteins which are coordinately assembled during phosphoryla tion of I kappa B alpha in response to external signals. (C) 1999 Academic Press.