The protease activity of human ATG4B is regulated by reversible oxidative modification.
The protease activity of human ATG4B is regulated by reversible oxidative modification.
复制标题
人 ATG4B 的蛋白酶活性受到可逆氧化修饰的调节。
DOI：
10.1080/15548627.2019.1709763
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发表时间：
2020
期刊：
影响因子：
13.3
通讯作者：
Li Min
中科院分区：
文献类型：
--
作者：
Zheng Xueping;Yang Zuolong;Gu Qianqian;Xia Fan;Fu Yuanyuan;Liu Peiqing;Yin Xiao-Ming;Li Min
巨细胞自噬/自噬在细胞质物质循环和代谢代谢中起着关键作用，其中ATG4B作为“剪刀”处理原Lc3和脂化Lc3来驱动自噬进程。越来越多的证据表明，在各种病理情况下，ROS与自噬之间存在着密切的联系。巧合的是，一些研究表明ATG4B可能受到氧化还原修饰的调节，但其潜在的分子机制及其与自噬的关系尚不明确。在这项研究中，我们验证了ATG4B活性肯定是以可逆的氧化还原方式调节的。我们还确定Cys292和Cys361是ATG4B形成可逆分子内二硫键的必需位点，这些二硫键对氧化应激做出反应。有趣的是，我们揭示了一个新的现象，即ATG4B同时在Cys292和Cys361形成二硫键连接的低聚物，并且这两个位置都经历了氧化还原修饰，从而调节了ATG4B的活性。在正常生长条件下，Cys292和Cys361双点突变细胞的自噬通量增加，氧化敏感性降低。总之，我们的研究揭示了一个新的分子机制，即Cys292和Cys361位的氧化修饰调节ATG4B的功能，从而调节自噬。缩写：AIR-OX：空气氧化；ATG4B：自噬相关的4B半胱氨酸肽酶；BCNU：1，3-二(2-氯乙基)-1-亚硝酸脲；CBB：考马斯亮蓝；CM：完全介质；CQ：氯喹；DTT：二硫苏糖醇；GSH：还原型谷胱甘肽；GSNO：S-亚硝基谷胱甘肽；GSSG：氧化的谷胱甘肽；HMW：高分子量；过氧化氢：过氧化氢；NAC：乙酰基-L-半胱氨酸；NEM：N-乙基马来酰亚胺；PE：磷脂酰乙胺；PTM：翻译后修饰；PTM：翻译后修饰；ROS，活性氧物种；WT：野生型
Macroautophagy/autophagy plays a pivotal role in cytoplasmic material recycling and metabolic turnover, in which ATG4B functions as a “scissor” for processing pro-LC3 and lipidated LC3 to drive the autophagy progress. Mounting evidence has demonstrated the tight connection between ROS and autophagy during various pathological situations. Coincidentally, several studies have shown that ATG4B is potentially regulated by redox modification, but the underlying molecular mechanism and its relationship with autophagy is ambiguous. In this study, we verified that ATG4B activity was definitely regulated in a reversible redox manner. We also determined that Cys292 and Cys361 are essential sites of ATG4B to form reversible intramolecular disulfide bonds that respond to oxidative stress. Interestingly, we unraveled a new phenomenon that ATG4B concurrently formed disulfide-linked oligomers at Cys292 and Cys361, and that both sites underwent redox modifications thereby modulating ATG4B activity. Finally, increased autophagic flux and decreased oxidation sensitivity were observed in Cys292 and Cys361 double site-mutated cells under normal growth conditions. In conclusion, our research reveals a novel molecular mechanism that oxidative modification at Cys292 and Cys361 sites regulates ATG4B function, which modulates autophagy.Abbreviations: Air-ox: air-oxidation; ATG4B: autophagy related 4B cysteine peptidase; BCNU: 1,3-bis(2-chloroethyl)-1-nitrosourea; CBB: Coomassie Brilliant Blue; CM: complete medium; CQ: chloroquine; DTT: dithiothreitol; GSH: reduced glutathione; GSNO: S-nitrosoglutathione; GSSG: oxidized glutathione; HMW: high molecular weight; H2O2: hydrogen peroxide; NAC: N-acetyl-L-cysteine; NEM: N-ethylmaleimide; PE: phosphatidylethanolamine; PTM: post-translational modification; ROS, reactive oxygen species; WT: wild type