Nitrosothiol-Trapping-Based Proteomic Analysis of S-Nitrosylation in Human Lung Carcinoma Cells.
Nitrosothiol-Trapping-Based Proteomic Analysis of S-Nitrosylation in Human Lung Carcinoma Cells.
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DOI：
10.1371/journal.pone.0169862
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发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Benhar M
中科院分区：
文献类型：
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作者：
Ben-Lulu S;Ziv T;Weisman-Shomer P;Benhar M
半胱氨酸残基的亚硝基化（S-亚硝基化）介导一氧化氮在正常和病变细胞中的许多细胞效应。最近的研究表明，某些蛋白质的S-亚硝基化可能在肿瘤进展和对治疗的反应性中发挥作用。然而，在癌细胞中S-亚硝基化的蛋白质靶点在很大程度上仍然是未知的。在这项研究中，我们使用我们最近开发的亚硝基巯基捕获方法来探索人A549肺癌细胞经S-亚硝基半胱氨酸或促炎细胞因子处理后的亚硝基蛋白质组。使用这种方法，我们确定了约300个假定的亚硝基化目标在S-亚硝基半胱氨酸处理的A549细胞和大约400个目标在精氨酸刺激的细胞。在这两个屏幕中确定的500多个蛋白质中，大多数代表S-亚硝基化的新靶点，通过与公开的亚硝基蛋白质组学数据进行比较发现。通过耦合的捕获程序与差异巯基标记，我们确定了近300个潜在的亚硝基化位点在约150个蛋白质。通过独立的方法对几种蛋白质的蛋白质组学结果进行了验证。生物信息学分析突出了S-亚硝基化靶向的重要细胞途径，特别是细胞周期和炎症信号传导。总之，我们的研究结果确定了肺癌细胞中一氧化氮的新分子靶点，并表明S-亚硝基化可能调节与肺癌进展密切相关的信号通路。
Nitrosylation of cysteines residues (S-nitrosylation) mediates many of the cellular effects of nitric oxide in normal and diseased cells. Recent research indicates that S-nitrosylation of certain proteins could play a role in tumor progression and responsiveness to therapy. However, the protein targets of S-nitrosylation in cancer cells remain largely unidentified. In this study, we used our recently developed nitrosothiol trapping approach to explore the nitrosoproteome of human A549 lung carcinoma cells treated with S-nitrosocysteine or pro-inflammatory cytokines. Using this approach, we identified about 300 putative nitrosylation targets in S-nitrosocysteine-treated A549 cells and approximately 400 targets in cytokine-stimulated cells. Among the more than 500 proteins identified in the two screens, the majority represent novel targets of S-nitrosylation, as revealed by comparison with publicly available nitrosoproteomic data. By coupling the trapping procedure with differential thiol labeling, we identified nearly 300 potential nitrosylation sites in about 150 proteins. The proteomic results were validated for several proteins by an independent approach. Bioinformatic analysis highlighted important cellular pathways that are targeted by S-nitrosylation, notably, cell cycle and inflammatory signaling. Taken together, our results identify new molecular targets of nitric oxide in lung cancer cells and suggest that S-nitrosylation may regulate signaling pathways that are critically involved in lung cancer progression.