Functional analysis of ubiquitin ligase by knock-out mise

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1) We have reported that F-box protein, FWD1 works as a ubiquitin ligase on the degradation of lκB or β-catenin. FWD1 constitutes SCF complex with Cul-1, Skp1, and Rbx1. In this study, we generated knock-out mice of Cul-1 and Skp1, which are components of SCF complex, and another F-box protein, Skp2. Analysis of these mice shows the biological role of protein degradation by SCF complex system, one category of proteolysis by ubiquitin-proteasome system.2) Skp2 is a ubiquitin ligase of cyclin E and p27^<Kip1>. In Skp2 knock-out mice, cyclin E and p27^<Kip1> was accumulated abnormally. Cells in the mutant mice contain markedly enlarged nuclei with polyploidy and multiple centrosomes, and show a reduced growth rate and increased apoptosis.3) We generated and characterized mice lacking both Skp2 and p27^<Kip1>. The Skp2^<-/->p27^<-/-> mice did not exhibit the overreplication phenotype, suggesting hat p27^<Kip1> accumulation is required for its development.4) Cul-1 and Skp1 knock-out mice are died between on embryonic day 5.5 and 6.5. In these embryos, cyclin E is accumulated. It is postulated that Cul-1 and Skp1 are common components of SCF complex and relate to degradation of many kinds of protein. Early embryonic death of knock-out mice supports that SCF complex participate in many kinds of protein degradation system.