THE CYCLOSOME, A LARGE COMPLEX CONTAINING CYCLIN-SELECTIVE UBIQUITIN LIGASE ACTIVITY, TARGETS CYCLINS FOR DESTRUCTION AT THE END OF MITOSIS
THE CYCLOSOME, A LARGE COMPLEX CONTAINING CYCLIN-SELECTIVE UBIQUITIN LIGASE ACTIVITY, TARGETS CYCLINS FOR DESTRUCTION AT THE END OF MITOSIS
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DOI：
10.1091/mbc.6.2.185
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发表时间：
1995-02-01
影响因子：
3.3
通讯作者：
HERSHKO, A
中科院分区：
文献类型：
--
作者：
SUDAKIN, V;GANOTH, D;HERSHKO, A
泛素介导的有丝分裂周期蛋白的降解是细胞退出有丝分裂所必需的。以前对无细胞系统的研究已经揭示了细胞周期蛋白-泛素连接和蛋白降解所需的四种成分：非特异性泛素激活酶E(1)，含有泛素载体蛋白活性的可溶性部分E(2)-C，含有泛素连接酶(E(3))活性的粗颗粒部分(E(3))，以及降解泛素化蛋白的固有活性26S蛋白酶体。在这里，我们鉴定了一种新的类似于1500 kDa的复合体，称为环体，它含有对细胞周期蛋白具有选择性的泛素连接酶活性E(3)-C。E(3)-C存在，但在间期没有活性；它可以通过添加cdc2在体外被激活，使泛素从E(2)-C转移到细胞周期蛋白。E(3)-C的活化动力学表明，在Cdc2和E(3)-C之间存在一个或多个中间体。环体相关的E(3)-C作用于细胞周期蛋白A和B，并且需要在每个细胞周期蛋白中存在野生型N-末端破坏框基序。泛素化的细胞周期蛋白随后被蛋白酶体迅速识别和降解。这些结果表明，环体相关的E(3)-C是周期蛋白破坏机制的组成部分，其活性最终由cdc2调节，因此，它是在早期胚胎细胞周期中直接负责设定有丝分裂周期蛋白水平的元件。
The ubiquitin-mediated degradation of mitotic cyclins is required for cells to exit from mitosis. Previous work with cell-free systems has revealed four components required for cyclin-ubiquitin ligation and proteolysis: a nonspecific ubiquitin-activating enzyme E(1), a soluble fraction containing a ubiquitin carrier protein activity called E(2)-C, a crude particulate fraction containing a ubiquitin ligase (E(3)) activity that is activated during M-phase, and a constitutively active 26S proteasome that degrades ubiquitinated proteins. Here, we identify a novel similar to 1500-kDa complex, termed the cyclosome, which contains a cyclin-selective ubiquitin Ligase activity, E(3)-C. E(3)-C is present but inactive during interphase; it can be activated in vitro by the addition of cdc2, enabling the transfer of ubiquitin from E(2)-C to cyclin. The kinetics of E(3)-C activation suggest the existence of one or more intermediates between cdc2 and E(3)-C. Cyclosome-associated E(3)-C acts on both cyclin A and B, and requires the presence of wild-type N-terminal destruction box motifs in each cyclin. Ubiquitinated cyclins are then rapidly recognized and degraded by the proteasome. These results identify the cyclosome-associated E(3)-C as the component of the cyclin destruction machinery whose activity is ultimately regulated by cdc2 and, as such, the element directly responsible for setting mitotic cyclin levels during early embryonic cell cycles.