Xing Jiale, Liu Peng, Zhao Lei, Huang Fang
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Beijing, China.
University of Chinese Academy of Sciences, Beijing, China.
Front Plant Sci. 2017 Dec 15;8:2154. doi: 10.3389/fpls.2017.02154. eCollection 2017.
The green alga is a key model organism for studying photosynthesis and oxidative stress in unicellular eukaryotes. Using a forward genetics approach, we have identified and characterized a mutant , which lacks a predicted protein named CGLD1 (Conserved in Green Lineage and Diatom 1) in GreenCut2, under normal and stress conditions. We show that loss of CGLD1 resulted in minimal photoautotrophic growth and PSII activity in the organism. We observed reduced amount of PSII complex and core subunits in the mutant based on blue-native (BN)/PAGE and immunoblot analysis. Moreover, exhibited increased sensitivity to high-light stress and altered tolerance to different reactive oxygenic species (ROS) stress treatments, i.e., decreased resistance to HO/or tert-Butyl hydroperoxide (t-BOOH) and increased tolerance to neutral red (NR) and rose bengal (RB) that induce the formation of singlet oxygen, respectively. Further analysis via quantitative real-time PCR (qRT-PCR) indicated that the increased singlet-oxygen tolerance of was largely correlated with up-regulated gene expression of glutathione--transferases (GST). The phenotypical and physiological implications revealed from our experiments highlight the important roles of CGLD1 in maintaining structure and function of PSII as well as in protection of Chlamydomonas under photo-oxidative stress conditions.
绿藻是研究单细胞真核生物光合作用和氧化应激的关键模式生物。我们采用正向遗传学方法，在正常和应激条件下，鉴定并表征了一个突变体，该突变体在GreenCut2中缺乏一种预测的名为CGLD1（绿藻谱系和硅藻中保守的1）的蛋白质。我们发现，CGLD1的缺失导致该生物体的光合自养生长和PSII活性极低。基于蓝色非变性（BN）/PAGE和免疫印迹分析，我们观察到该突变体中PSII复合物和核心亚基的数量减少。此外，该突变体对高光胁迫的敏感性增加，对不同活性氧（ROS）胁迫处理的耐受性改变，即对过氧化氢（HO）/叔丁基过氧化氢（t-BOOH）的抗性降低，对分别诱导单线态氧形成的中性红（NR）和孟加拉玫瑰红（RB）的耐受性增加。通过定量实时PCR（qRT-PCR）进一步分析表明，该突变体对单线态氧耐受性的增加在很大程度上与谷胱甘肽-S-转移酶（GST）基因表达上调相关。我们实验揭示的表型和生理意义突出了CGLD1在维持PSII结构和功能以及在光氧化应激条件下保护衣藻中的重要作用。
