S-Nitrosylation at the active site decreases the ubiquitin-conjugating activity of ubiquitin-conjugating enzyme E2 D1 (UBE2D1), an ERAD-associated protein
S-Nitrosylation at the active site decreases the ubiquitin-conjugating activity of ubiquitin-conjugating enzyme E2 D1 (UBE2D1), an ERAD-associated protein
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DOI：
10.1016/j.bbrc.2020.02.011
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发表时间：
2020-04-16
影响因子：
3.1
通讯作者：
Uehara, Takashi
中科院分区：
文献类型：
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作者：
Fujikawa, Kana;Nakahara, Kengo;Uehara, Takashi
蛋白质半胱氨酸硫醇的S亚硝化是由一氧化氮(NO)介导的翻译后修饰。NO的过量产生导致亚硝化性应激，亚硝化性应激可诱导内质网应激。我们以前曾报道，蛋白质二硫键异构酶和内质网应激感受器肌醇需求酶1的S亚硝化降低了它们的酶活性。然而，目前尚不清楚亚硝化胁迫是否影响内质网相关降解(ERAD)，ERAD是一个独立的内质网应激调节系统，负责通过泛素-蛋白酶体途径降解底物。在本研究中，我们发现已知的ERAD底物丝氨酸/苏氨酸蛋白激酶1(SGK1)的泛素化被亚硝化胁迫减弱。Hsc70相互作用蛋白的C末端(CHIP)和泛素结合酶E2D1(UBE2D1)参与了这一修饰。我们用LC-MS/MS检测到UBE2D1的活性中心为S硝基，其活性中心为Cys85。此外，体外和基于细胞的实验表明，S硝化的UBE2D1降低了泛素结合活性。我们的结果表明，亚硝化应激干扰ERAD，导致包括分子伴侣和内质网应激传感通路在内的多条途径共同中断，导致内质网应激延长。鉴于亚硝化应激和内质网应激在帕金森氏病(PD)和阿尔茨海默病(AD)患者的大脑中表达上调，我们的发现可能为进一步了解这些神经退行性疾病的发病机制提供依据。(C)2020 Elsevier Inc.保留所有权利。
S-Nitrosylation of protein cysteine thiol is a post-translational modification mediated by nitric oxide (NO). The overproduction of NO causes nitrosative stress, which is known to induce endoplasmic reticulum (ER) stress. We previously reported that S-nitrosylation of protein disulfide isomerase (PDI) and the ER stress sensor inositol-requiring enzyme 1 (IRE1) decreases their enzymatic activities. However, it remains unclear whether nitrosative stress affects ER-associated degradation (ERAD), a separate ER stress regulatory system responsible for the degradation of substrates via the ubiquitin-proteasomal pathway.In the present study, we found that the ubiquitination of a known ERAD substrate, serine/threonine-protein kinase 1 (SGK1), is attenuated by nitrosative stress. C-terminus of Hsc70-interacting protein (CHIP) together with ubiquitin-conjugating enzyme E2 D1 (UBE2D1) are involved in this modification. We detected that UBE2D1 is S-nitrosylated at its active site, Cys85 by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Furthermore, in vitro and cell-based experiments revealed that S-nitrosylated UBE2D1 has decreased ubiquitin-conjugating activity.Our results suggested that nitrosative stress interferes with ERAD, leading to prolongation of ER stress by co-disruption of various pathways, including the molecular chaperone and ER stress sensor pathways. Given that nitrosative stress and ER stress are upregulated in the brains of patient with Parkinson's disease (PD) and of those with Alzheimer's disease (AD), our findings may provide further insights into the pathogenesis of these neurodegenerative disorders. (C) 2020 Elsevier Inc. All rights reserved.