Subquinocin, a small molecule inhibitor of CYLD and USP-family deubiquitinating enzymes, promotes NF-κB signaling
Subquinocin, a small molecule inhibitor of CYLD and USP-family deubiquitinating enzymes, promotes NF-κB signaling
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Subquinocin 是 CYLD 和 USP 家族去泛素化酶的小分子抑制剂，可促进 NF-κB 信号转导
DOI：
10.1016/j.bbrc.2019.12.049
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发表时间：
2020
影响因子：
3.1
通讯作者：
Sawasaki Tatsuya
中科院分区：
文献类型：
--
作者：
Yamanaka Satoshi;Sato Yusuke;Oikawa Daisuke;Goto Eiji;Fukai Shuya;Tokunaga Fuminori;Takahashi Hirotaka;Sawasaki Tatsuya
肿瘤抑制因子CYLD负调控多泛素化依赖的细胞信号，如核因子-κB信号。除CYLD外，多种去泛素化酶（deubiquitination enzyme, DUBs）也参与了该信号通路的调控，CYLD的具体作用尚不清楚。在这里，我们发现了一种名为Subquinocin的小化学物质，使用小麦无细胞蛋白合成和alphasgreen技术抑制重组CYLD的DUB活性。在细胞中，亚喹啉增加了NEMO和RIP1的多泛素化，增强了NF-κB的活化。建模和突变分析表明，Subquinocin与CYLD中的Y940相互作用，该基因位于CYLD的催化中心附近，在usp家族dub中保守。进一步的生化评价显示，亚喹诺肽抑制usp家族DUBs，但不抑制OTU等其他家族DUBs。尽管Subquinocin对usp家族DUBs具有广泛的特异性，但在CYLD- ko细胞中，Subquinocin对NF-κB信号的抑制作用可以忽略不计，这表明CYLD是Subquinocin抑制NF-κB信号的主要靶点。总之，本文鉴定的亚喹啉是分析usp家族DUBs介导的信号转导的有用工具。
The tumor suppressor CYLD negatively regulates polyubiquitination-dependent cellular signaling such as nuclear factor (NF)-κB signaling. In addition to CYLD, multiple deubiquitinating enzymes (DUBs) are also involved in the regulation of this signaling pathway, and distinct role of CYLD is yet to be clarified. Here, we identified a small chemical named Subquinocin that inhibited the DUB activity of recombinant CYLD using a wheat cell-free protein synthesis and an AlphaScreen technology. In cells, Subquinocin increased the polyubiquitination of NEMO and RIP1 and enhanced NF-κB activation. Modeling and mutation analyses indicated that Subquinocin interacted with Y940 in CYLD, which locates close to catalytic center of CYLD, and is conserved among the USP-family DUBs. Further biochemical evaluation revealed that Subquinocin inhibited USP-family DUBs, but not other family DUBs including OTU. Although Subquinocin showed a broad specificity toward USP-family DUBs, the inhibitory effect of Subquinocin on NF-κB signaling was negligible inCYLD-KO cells, indicating that CYLD is a major target of Subquinocin on the suppression of NF-κB signaling. In conclusion, Subquinocin identified here is a useful tool to analyze the signal transduction mediated by USP-family DUBs.