SCFFbxw7 ubiquitylates KLF7 for degradation in a manner dependent on GSK-3-mediated phosphorylation.
SCFFbxw7 ubiquitylates KLF7 for degradation in a manner dependent on GSK-3-mediated phosphorylation.
复制标题
SCFFbxw7 泛素化 KLF7，以依赖于 GSK-3 介导的磷酸化的方式降解。
DOI：
10.1111/gtc.12680
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
Nakayama KI
中科院分区：
文献类型：
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作者：
Sugiyama S;Yumimoto K;Inoue I;Nakayama KI
泛素连接酶及其底物之间的生物学关系在很大程度上还不清楚。我们之前开发了一种方法-基于差异蛋白质组学的泛素化底物鉴定(DiPIUS)-用于对给定泛素连接酶的底物进行全面鉴定。我们现在已经将DiPIUS应用于三个细胞系(mHepa，Neuro2A和C2C12)的F-box蛋白Fbxw7，从而确定Krüppel-like factor7(KLF7)是SCFFbxw7泛素连接酶复合体的候选底物。KLF7与Fbxw7相互作用，并经历Fbxw7介导的多泛素化。KLF7的稳定性可通过Fbxw7的缺失、KLF7的可能的CDC4磷酸降解(CPD)突变或暴露于糖原合成酶激酶-3(GSK-3)抑制剂而增加。Fbxw7在Neuro2A细胞中的过表达下调了p21Cip1基因的表达，而p21Cip1基因是KLF7在神经元分化和维持中的转录靶点。尽管在KLF7最接近的平行对数KLF6中存在几乎相同的CPD序列，但该序列的突变既不影响KLF6与Fbxw7的相互作用，也不影响其半衰期。我们的结果表明，KLF7而不是KLF6是SCFFbxw7的真正底物，SCFFbxw7对KLF7丰度的控制可能有助于调节神经元的分化和维持。
The biological relation between ubiquitin ligases and their substrates has been largely unclear. We previously developed a method—differential proteomics‐based identification of ubiquitylation substrates (DiPIUS)—for the comprehensive identification of substrates for a given ubiquitin ligase. We have now applied DiPIUS to the F‐box protein Fbxw7 in three cell lines (mHepa, Neuro2A and C2C12) and thereby identified Krüppel‐like factor 7 (KLF7) as a candidate substrate of the SCFFbxw7ubiquitin ligase complex. KLF7 was shown to interact with Fbxw7 and to undergo Fbxw7‐mediated polyubiquitylation. The stability of KLF7 was increased by depletion of Fbxw7, mutation of a putative Cdc4 phosphodegron (CPD) of KLF7 or exposure to inhibitors of glycogen synthase kinase‐3 (GSK‐3). Over‐expression of Fbxw7 in Neuro2A cells down‐regulated expression of the p21Cip1gene, which is a transcriptional target of KLF7 in neuronal differentiation and maintenance. Despite the presence of an almost identical CPD sequence in KLF6, the closest paralog of KLF7, mutation of this sequence affected neither the interaction of KLF6 with Fbxw7 nor its half‐life. Our results suggest that KLF7, but not KLF6, is a bona fide substrate of SCFFbxw7, and that control of KLF7 abundance by SCFFbxw7might contribute to the regulation of neuronal differentiation and maintenance.