Functional analysis of ubiquitin ligase by knock-out mise

1) 我们报道了 F-box 蛋白 FWD1 作为泛素连接酶降解 lκB 或 β-catenin。 FWD1与Cul-1、Skp1和Rbx1构成SCF复合物。在这项研究中，我们培育了 Cul-1 和 Skp1（SCF 复合物的组成部分）和另一种 F-box 蛋白 Skp2 的敲除小鼠。对这些小鼠的分析显示了SCF复合系统对蛋白质降解的生物学作用，这是泛素-蛋白酶体系统进行的蛋白水解的一类。2)Skp2是细胞周期蛋白E和p27^<Kip1>的泛素连接酶。在Skp2敲除小鼠中，细胞周期蛋白E和p27^<Kip1>异常积累。突变小鼠中的细胞含有明显增大的多倍体细胞核和多个中心体，并且显示出生长速率降低和细胞凋亡增加。3)我们生成并表征了同时缺乏Skp2和p27^<Kip1>的小鼠。 Skp2^<-/->p27^</-/-> 小鼠没有表现出过度复制表型，表明 p27^<Kip1> 积累是其发育所必需的。4) Cul-1 和 Skp1 敲除小鼠在胚胎第 5.5 天和第 6.5 天之间死亡。在这些胚胎中，细胞周期蛋白 E 积累。据推测，Cul-1和Skp1是SCF复合物的常见成分，并与多种蛋白质的降解有关。基因敲除小鼠的早期胚胎死亡支持SCF复合物参与多种蛋白质降解系统。

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.