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DNA Walker-Driven Mass Nanotag Assembly System for Simultaneously Profiling Dual Markers of Oxidative Stress at Different Cellular Locations
Analytical Chemistry ( IF 7.4 ) Pub Date : 2024-05-13 , DOI: 10.1021/acs.analchem.4c01115
Yinyin Fan 1 , Zhenzhen Zhang 1 , Xue Zhang 1 , Aobo Xu 2 , Jun-Jie Zhu 1 , Qianhao Min 1
Affiliation  

Simultaneous profiling of redox-regulated markers at different cellular sublocations is of great significance for unraveling the upstream and downstream molecular mechanisms of oxidative stress in living cells. Herein, by synchronizing dual target-triggered DNA machineries in one nanoentity, we engineered a DNA walker-driven mass nanotag (MNT) assembly system (w-MNT-AS) that can be sequentially activated by oxidative stress-associated mucin 1 (MUC1) and apurinic/apyrimidinic endonuclease 1 (APE1) from plasma membrane to cytoplasm and induce recycled assembly of MNTs for multiplex detection of the two markers by matrix-assisted laser desorption ionization mass spectrometry (MALDI MS). In the working cascade, the sensing process governs the separate activation of w-MNT-AS by MUC1 and APE1 in diverse locations, while the assembly process contributes to the parallel amplification of the ion signal of the characteristic mass tags. In this manner, the differences between MCF-7, HeLa, HepG2, and L02 cells in membrane MUC1 expression and cytoplasmic APE1 activation were fully characterized. Furthermore, the oxidative stress level and dynamics caused by exogenous H2O2, doxorubicin, and simvastatin were comprehensively demonstrated by tracking the fate of the two markers across different cellular locations. The proposed w-MNT-AS coupled MS method provides an effective route to probe multiple functional molecules that lie at different locations while participating in the same cellular event, facilitating the mechanistic studies on cellular response to oxidative stress and other disease-related cellular processes.

中文翻译:


DNA Walker 驱动的质量纳米标签组装系统可同时分析不同细胞位置氧化应激的双标记



同时分析不同细胞亚位的氧化还原调节标记对于揭示活细胞氧化应激的上下游分子机制具有重要意义。在此,通过在一个纳米实体中同步双靶标触发的DNA机器,我们设计了一种DNA步行者驱动的质量纳米标签(MNT)组装系统(w-MNT-AS),该系统可以被氧化应激相关粘蛋白1(MUC1)依次激活和无嘌呤/无嘧啶核酸内切酶 1 (APE1) 从质膜到细胞质,并诱导 MNT 的循环组装,以便通过基质辅助激光解吸电离质谱 (MALDI MS) 多重检测这两种标记物。在工作级联中,传感过程控制MUC1和APE1在不同位置对w-MNT-AS的单独激活,而组装过程有助于并行放大特征质量标签的离子信号。通过这种方式,充分表征了 MCF-7、HeLa、HepG2 和 L02 细胞在膜 MUC1 表达和细胞质 APE1 激活方面的差异。此外,通过跟踪两种标记物在不同细胞位置的命运,全面证明了外源性 H 2 O 2 、阿霉素和辛伐他汀引起的氧化应激水平和动态。所提出的w-MNT-AS耦合MS方法提供了一种有效的途径来探测参与同一细胞事件时位于不同位置的多个功能分子,促进细胞对氧化应激和其他疾病相关细胞过程的反应的机制研究。
更新日期:2024-05-13
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