Time-Resolved FT-IR Microspectroscopy of Protein Aggregation Induced by Heat-Shock in Live Cells
Time-Resolved FT-IR Microspectroscopy of Protein Aggregation Induced by Heat-Shock in Live Cells
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DOI：
10.1021/ac5040659
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发表时间：
2015-04-07
影响因子：
7.4
通讯作者：
Vaccari, Lisa
中科院分区：
文献类型：
--
作者：
Mitri, Elisa;Kenig, Sasa;Vaccari, Lisa
维持细胞蛋白质的正确折叠对于维持细胞内环境的稳定至关重要。蛋白质内环境失调、蛋白质折叠异常和蛋白质聚集确实与多种疾病有关，包括癌症、衰老相关疾病和神经退行性疾病。蛋白质聚集体的积累也可以由各种应激条件诱导，如温度升高或氧化应激。在这项工作中，我们用傅里叶变换红外(FT-IR)显微光谱监测了活体乳腺癌MCF-7和乳腺癌MDA-MB 231细胞系对细胞介质温度从37+/-0.5摄氏度上升到42+/-0.5摄氏度所引起的严重热休克(HS)的反应。通过对光谱二阶导数的时间演化的研究和FT-IR吸光度数据的二维相关分析，我们能够识别这两种细胞系之间共同的突然热休克反应(HSR)。在胁迫处理的前40min内，哺乳动物细胞膜的超流性、信号脂的瞬时增加以及蛋白质组谱的改变都被监测到，而在胁迫40min至2 h后，检测到扩展的β折叠蛋白聚集体在细胞内的持续积累，进一步证明了FT-IR显微光谱在实时监测活细胞在外界刺激下发生的生化重排的能力。
Maintaining the correct folding of cellular proteins is essential for preserving cellular homeostasis. Protein dishomeostasis, aberrant protein folding, and protein aggregation are indeed involved in several diseases including cancer, aging-associated, and neurodegenerative disorders. Accumulation of protein aggregates can also be induced from a variety of stressful conditions, such as temperature increase or oxidative stress. In this work, we monitored by Fourier transform-infrared (FT-IR) microspectroscopy the response of live breast cancer MCF-7 and mammary breast adenocarcinoma MDA-MB 231 cell lines to severe heat-shock (HS), caused by the rise of the cellular medium temperature from 37 +/- 0.5 degrees C to 42 +/- 0.5 degrees C. Through the study of the time-evolution of the second derivatives of the spectra and by the 2D correlation analysis of FT-IR absorbance data, we were able to identify a common sudden heat-shock response (HSR) among the two cell lines. The hyperfluidization of mammalian cell membranes, the transient increment of the signal lipids, as well as the alteration of proteome profile were all monitored within the first 40 min of stress application, while the persistent intracellular accumulation of extended beta-folded protein aggregates was detected after 40 min up to 2 h. As a whole, this paper offers a further prove of the diagnostic capabilities of FT-IR microspectroscopy for monitoring in real-time the biochemical rearrangements undergone by live cells upon external stimulation.