The APC subunit Doc1 promotes recognition of the substrate destruction box
The APC subunit Doc1 promotes recognition of the substrate destruction box
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DOI：
10.1016/j.cub.2004.12.066
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发表时间：
2005-01-11
期刊：
影响因子：
9.2
通讯作者：
Morgan, DO
中科院分区：
文献类型：
--
作者：
Carroll, CW;Enquist-Newman, M;Morgan, DO
背景：有丝分裂过程中准确的染色体分离需要协同破坏有丝分裂调节因子Securin和Cyclins。后期促进复合体(APC)是一种多亚基泛素-蛋白质连接酶，催化这些蛋白质和其他蛋白质的多泛素化，从而促进它们的破坏。APC如何识别其底物还不是很清楚。在有丝分裂中，APC激活子CDC20与APC结合，并被认为通过与称为破坏框的保守靶蛋白基序相互作用来招募底物。一种名为CDH1的相关蛋白质在G1期发挥着类似的功能。结果：为了更好地理解DOC1促进底物与APC结合的机制，我们在DOC1中产生了一系列点突变，并分析了它们对底物泛素化过程的影响。降低doc1功能的突变分为两类，它们定义了蛋白质的空间和功能不同的区域。一个区域，包括羧基末端，将DOC1锚定到APC上，但不影响底物识别。另一个区域位于DOG1的相反表面，DOG1需要该区域来增强底物与APC的结合。重要的是，刺激DOC1结合还需要底物包含完整的破坏框。携带doc1突变的细胞在有丝分裂过程中消除了高水平APC底物的底物识别延迟。结论：doc1参与了APC对底物破坏框的识别。DOC1的这一功能是体内有效底物蛋白分解所必需的。
Background: Accurate chromosome segregation during mitosis requires the coordinated destruction of the mitotic regulators securin and cyclins. The anaphase-promoting complex (APC) is a multisubunit ubiquitin-protein ligase that catalyzes the polyubiquitination of these and other proteins and thereby promotes their destruction. How the APC recognizes its substrates is not well understood. In mitosis, the APC activator Cdc20 binds to the APC and is thought to recruit substrates by interacting with a conserved target protein motif called the destruction box. A related protein, called Cdh1, performs a similar function during G1. Recent evidence, however, suggests that the core APC subunit Doc1 also contributes to substrate recognition.Results: To better understand the mechanism by which Doc1 promotes substrate binding to the APC, we generated a series of point mutations in Doc1 and analyzed their effects on the processivity of substrate ubiquitination. Mutations that reduce Doc1 function fall into two classes that define spatially and functionally distinct regions of the protein. One region, which includes the carboxy terminus, anchors Doc1 to the APC but does not influence substrate recognition. The other region, located on the opposite face of Doc1, is required for Doc1 to enhance substrate binding to the APC. Importantly, stimulation of binding by Doc1 also requires that the substrate contain an intact destruction box. Cells carrying DOC1 mutations that eliminate substrate recognition delay in mitosis with high levels of APC substrates.Conclusions: Doc1 contributes to recognition of the substrate destruction box by the APC. This function of Doc1 is necessary for efficient substrate proteolysis in vivo.