Transformations of the macromolecular landscape at mitochondria during DNA-damage-induced apoptotic cell death.
Transformations of the macromolecular landscape at mitochondria during DNA-damage-induced apoptotic cell death.
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DOI：
10.1038/cddis.2014.405
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发表时间：
2014-10-09
影响因子：
9
通讯作者：
Chandra D
中科院分区：
文献类型：
--
作者：
Yadav N;Pliss A;Kuzmin A;Rapali P;Sun L;Prasad P;Chandra D
细胞凋亡是一个由线粒体调控的动态过程，对细胞呼吸和生存至关重要。细胞凋亡的执行是由线粒体上的多个蛋白质信号事件介导的。细胞凋亡的发生和发展需要大量的促凋亡因子，这些因子要么从线粒体释放出来，要么被隔离在线粒体中，这可能会改变细胞器的生物分子组成。在这一通讯中，利用拉曼光谱，我们证明了线粒体生物分子组成的转变可以作为单个细胞的凋亡标记。我们首次发现，在细胞凋亡过程中，线粒体的RNA、DNA、蛋白质和脂肪成分的浓度发生了显著的变化。线粒体蛋白质的结构分析表明，线粒体上的α-螺旋二级结构含量减少，无规卷曲和β-片层水平增加。这可能代表了细胞凋亡的另一个标志。令人惊讶的是，我们观察到在存在和不存在关键的促凋亡蛋白Bax(Bcl2相关X蛋白)的情况下，线粒体大分子含量的变化几乎相同。线粒体DNA水平的升高与线粒体DNA(MtDNA)、细胞内活性氧(ROS)和线粒体ROS的生成增加相对应。聚合酶γ(Polg)、线粒体解旋酶Twinkle和线粒体转录因子A(TFAM)的表达上调与线粒体DNA和核糖核酸合成增加相关。氧化磷酸化复合体活性的提高支持了线粒体在细胞凋亡过程中的功能性呼吸。因此，我们定义了在存在和不存在Bax蛋白的情况下，线粒体大分子结构与细胞凋亡进程之间未知的动态关联。这些发现为用非侵入性单细胞方法监测细胞的生理状态开辟了新的途径。
Apoptosis is a dynamic process regulated by mitochondrion critical for cellular respiration and survival. Execution of apoptosis is mediated by multiple protein signaling events at mitochondria. Initiation and progression of apoptosis require numerous apoptogenic factors that are either released from or sequestered in mitochondria, which may transform the biomolecular makeup of the organelle. In this communication, using Raman microspectroscopy, we demonstrate that transformation in biomolecular composition of mitochondrion may be used as apoptosis marker in an individual cell. For the first time, we show that significant changes occur in the concentrations of RNA, DNA, protein, and lipid constituents of mitochondria during apoptosis. The structural analysis of proteins on mitochondria demonstrated a decrease in α-helix secondary structure content, and an increase in the levels of random coils and β-sheets on mitochondria. This may represent an additional hallmark of apoptosis. Strikingly, we observed nearly identical changes in macromolecular content of mitochondria both in the presence and absence of a key proapoptotic protein, Bax (Bcl-2-associated X protein). Increased DNA level in mitochondria corresponded with higher mitochondrial DNA (mtDNA), cellular reactive oxygen species (ROS), and mitochondrial ROS production. Upregulation of polymerase-γ (POLG), mitochondrial helicase Twinkle, and mitochondrial transcription factor A (Tfam) in response to DNA damage correlated with increased mtDNA and RNA synthesis. Elevated activity of oxidative phosphorylation complexes supports functional mitochondrial respiration during apoptosis. Thus, we define previously unknown dynamic correlation of macromolecular structure of mitochondria and apoptosis progression in the presence and absence of Bax protein. These findings open up a new approach for monitoring physiological status of cells by non invasive single-cell method.