Live-cell visualization of excitation energy dynamics in chloroplast thylakoid structures.
Live-cell visualization of excitation energy dynamics in chloroplast thylakoid structures.
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DOI：
10.1038/srep29940
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发表时间：
2016-07-15
影响因子：
4.6
通讯作者：
Nakano A
中科院分区：
文献类型：
--
作者：
Iwai M;Yokono M;Kurokawa K;Ichihara A;Nakano A
长期以来，人们一直使用各种分析方法研究光合作用背后复杂的分子过程。然而，这种光合作用过程的三维（3D）动力学仍然未被探索，由于技术上的限制，在体内的细胞器内机制的调查。通过开发一个系统，高速三维激光扫描共聚焦显微镜结合高灵敏度的多通道检测，我们可视化的激发能量动力学活小立碗藓细胞叶绿体内的类囊体结构。两个不同的类囊体结构的叶绿体，即基粒和间质片层，可视化三维活细胞。同时检测叶绿素（Chl）荧光的短波长区（~670 nm）和长波长区（~680 nm）揭示了不同的空间特征--不规则结构和垂直结构。光谱分析表明，叶绿素荧光的较短和较长的波长区域的影响更多的自由捕光天线蛋白和光系统II超复合物，分别。较短和较长波长区域的高速3D延时成像也揭示了不同的结构动态-分别在1.5秒内快速和缓慢移动。叶绿素荧光的两个波长区域的这种结构动力学将指示捕光天线蛋白和光系统之间的激发能量动力学，反映了类囊体膜中光合作用蛋白的能量活性性质。
The intricate molecular processes underlying photosynthesis have long been studied using various analytic approaches. However, the three-dimensional (3D) dynamics of such photosynthetic processes remain unexplored due to technological limitations related to investigating intraorganellar mechanisms in vivo. By developing a system for high-speed 3D laser scanning confocal microscopy combined with high-sensitivity multiple-channel detection, we visualized excitation energy dynamics in thylakoid structures within chloroplasts of live Physcomitrella patens cells. Two distinct thylakoid structures in the chloroplast, namely the grana and stroma lamellae, were visualized three-dimensionally in live cells. The simultaneous detection of the shorter (than ~670 nm) and longer (than ~680 nm) wavelength regions of chlorophyll (Chl) fluorescence reveals different spatial characteristics—irregular and vertical structures, respectively. Spectroscopic analyses showed that the shorter and longer wavelength regions of Chl fluorescence are affected more by free light-harvesting antenna proteins and photosystem II supercomplexes, respectively. The high-speed 3D time-lapse imaging of the shorter and longer wavelength regions also reveals different structural dynamics—rapid and slow movements within 1.5 seconds, respectively. Such structural dynamics of the two wavelength regions of Chl fluorescence would indicate excitation energy dynamics between light-harvesting antenna proteins and photosystems, reflecting the energetically active nature of photosynthetic proteins in thylakoid membranes.