The NEDD8 pathway is essential for SCF(beta -TrCP)-mediated ubiquitination and processing of the NF-kappa B precursor p105.
The NEDD8 pathway is essential for SCF(beta -TrCP)-mediated ubiquitination and processing of the NF-kappa B precursor p105.
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发表时间：
2002
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通讯作者：
R. Amir;K. Iwai;A. Ciechanover
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作者：
R. Amir;K. Iwai;A. Ciechanover
NF-κ B的p50亚基通过前体p105的有限加工产生。IkappaB激酶介导的p105的C-末端结构域的磷酸化募集SCF（β-TrCP）泛素连接酶，导致p105的快速泛素化和随后的加工/降解。已知NEDD 8在修饰SCF连接酶的cullin组分后激活SCF连接酶。在这里，我们表明，NEDD化是必需的共轭和SCF（β-TrCP）的p105的分子磷酸化后的加工。在粗提物中，显性负性E2酶UBC 12抑制p105的缀合和加工，并且抑制通过过量的WT-UBC 12而减轻。在重建的无细胞系统中，p105的泛素化仅在NEDD 8通路的所有三个组分E1、E2和NEDD 8存在下才被刺激。不能被NEDDylated的Cul-1突变体不能刺激p105的泛素化和加工。在细胞中也观察到类似的发现。应该注意的是，NEDD化仅对p105的刺激加工是必需的，而对p105的基础加工不是必需的。尽管p105的加工机制在很大程度上还不清楚，但很明显，NEDD化和SCF（β-TrCP）对p105和IkappaB α的协调活性是控制NF-κ B活性的重要调节机制。
The p50 subunit of NF-kappaB is generated by limited processing of the precursor p105. IkappaB kinase-mediated phosphorylation of the C-terminal domain of p105 recruits the SCF(beta-TrCP) ubiquitin ligase, resulting in rapid ubiquitination and subsequent processing/degradation of p105. NEDD8 is known to activate SCF ligases following modification of their cullin component. Here we show that NEDDylation is required for conjugation and processing of p105 by SCF(beta-TrCP) following phosphorylation of the molecule. In a crude extract, a dominant negative E2 enzyme, UBC12, inhibits both conjugation and processing of p105, and inhibition is alleviated by an excess of WT- UBC12. In a reconstituted cell-free system, ubiquitination of p105 was stimulated only in the presence of all three components of the NEDD8 pathway, E1, E2, and NEDD8. A Cul-1 mutant that cannot be NEDDylated could not stimulate ubiquitination and processing of p105. Similar findings were observed also in cells. It should be noted that NEDDylation is required only for the stimulated but not for basal processing of p105. Although the mechanisms that underlie processing of p105 are largely obscure, it is clear that NEDDylation and the coordinated activity of SCF(beta-TrCP) on both p105 and IkappaBalpha serve as an important regulatory mechanism controlling NF-kappaB activity.