The SCFβ-TRCP-ubiquitin ligase complex associates specifically with phosphorylated destruction motifs in IκBα and β-catenin and stimulates IκBα ubiquitination in vitro
The SCFβ-TRCP-ubiquitin ligase complex associates specifically with phosphorylated destruction motifs in IκBα and β-catenin and stimulates IκBα ubiquitination in vitro
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DOI：
10.1101/gad.13.3.270
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发表时间：
1999-02-01
影响因子：
10.5
通讯作者：
Harper, JW
中科院分区：
文献类型：
--
作者：
Winston, JT;Strack, P;Harper, JW
泛素介导的蛋白降解在控制细胞内几个重要调控分子的水平方面起着核心作用，如细胞周期蛋白、细胞周期蛋白I、p53和I kappa Bα。许多不同的致炎信号导致核因子-kappa B抑制蛋白I kappa Bα的特异性磷酸化和随后泛素介导的破坏。泛素化反应中的底物特异性在很大程度上是由E3-泛素连接酶与其底物的特异性结合所介导的。一类E3连接酶是由最近描述的SCF复合体定义的，其原型首先在芽殖酵母中描述，并在bar KP1下包含(S)，在bar dc53下包含(C)，在bar-box蛋白CDC4下包含(F)。这些复合体通过模块化的F-box蛋白以磷酸化依赖的方式识别它们的底物。在这里，我们描述了一种新的哺乳动物SCF复合体SCFβ-TRCP的生化解剖，它以磷酸化依赖的方式特异性识别I kappa Bα(残基21-41)中的19个氨基酸破坏基序。这个SCF复合体还识别β-连环蛋白中保守的破坏基序，β-连环蛋白是一种水平也受磷酸化依赖的泛素化调节的蛋白质。内源性I-kappa Bα-泛素连接酶活性与SCF-β-TRCP协同作用。此外，在哺乳动物细胞中组装的重组SCFbeta-TRCP具有磷酸I kappa Bα特异性泛素连接酶活性。我们的结果表明，SCFβ-TRCP复合体通过激活NF-kappa B途径和抑制β-catenin途径在多种转录程序中发挥作用。
Ubiquitin-mediated proteolysis has a central role in controlling the intracellular levels of several important regulatory molecules such as cyclins, CKIs, p53, and I kappa B alpha. Many diverse proinflammatory signals lead to the specific phosphorylation and subsequent ubiquitin-mediated destruction of the NF-kappa B inhibitor protein I kappa B alpha. Substrate specificity in ubiquitination reactions is, in large part, mediated by the specific association of the E3-ubiquitin ligases with their substrates. One class of E3 ligases is defined by the recently described SCF complexes, the archetype of which was first described in budding yeast and contains (S) under bar kp1, (C) under bar dc53, and the (F) under bar-box protein Cdc4. These complexes recognize their substrates through modular F-box proteins in a phosphorylation-dependent manner. Here we describe a biochemical dissection of a novel mammalian SCF complex, SCFbeta-TRCP, that specifically recognizes a 19-amino-acid destruction motif in I kappa B alpha (residues 21-41) in a phosphorylation-dependent manner. This SCF complex also recognizes a conserved destruction motif in beta-catenin, a protein with levels also regulated by phosphorylation-dependent ubiquitination. Endogenous I kappa B alpha-ubiquitin ligase activity cofractionates with SCFbeta-TRCP. furthermore, recombinant SCFbeta-TRCP assembled in mammalian cells contains phospho-I kappa B alpha-specific ubiquitin ligase activity. Our results suggest that an SCFbeta-TRCP complex functions in multiple transcriptional programs by activating the NF-kappa B pathway and inhibiting the beta-catenin pathway.