Department of Biotechnology, University of Verona, Verona, 37134, Italy.
Department of Biology and Biotechnology, University of Pavia, Pavia, 27100, Italy.
Photosynth Res. 2019 Dec;142(3):249-264. doi: 10.1007/s11120-019-00656-3. Epub 2019 Jul 3.
Non-photochemical quenching, NPQ, of chlorophyll fluorescence regulates the heat dissipation of chlorophyll excited states and determines the efficiency of the oxygenic photosynthetic systems. NPQ is regulated by a pH-sensing protein, responding to the chloroplast lumen acidification induced by excess light, coupled to an actuator, a chlorophyll/xanthophyll subunit where quenching reactions are catalyzed. In plants, the sensor is PSBS, while the two pigment-binding proteins Lhcb4 (also known as CP29) and LHCII are the actuators. In algae and mosses, stress-related light-harvesting proteins (LHCSR) comprise both functions of sensor and actuator within a single subunit. Here, we report on expressing the lhcsr1 gene from the moss Physcomitrella patens into several Arabidopsis thaliana npq4 mutants lacking the pH sensing PSBS protein essential for NPQ activity. The heterologous protein LHCSR1 accumulates in thylakoids of A. thaliana and NPQ activity can be partially restored. Complementation of double mutants lacking, besides PSBS, specific xanthophylls, allowed analyzing chromophore requirement for LHCSR-dependent quenching activity. We show that the partial recovery of NPQ is mostly due to the lower levels of Zeaxanthin in A. thaliana in comparison to P. patens. Complemented npq2npq4 mutants, lacking besides PSBS, Zeaxanthin Epoxidase, showed an NPQ recovery of up to 70% in comparison to A. thaliana wild type. Furthermore, we show that Lutein is not essential for the folding nor for the quenching activity of LHCSR1. In short, we have developed a system to study the function of LHCSR proteins using heterologous expression in a variety of A. thaliana mutants.
非光化学猝灭，NPQ，调节叶绿素激发态的热耗散，并决定了放氧光合作用系统的效率。NPQ 由一种 pH 感应蛋白调节，对过量光诱导的叶绿体腔酸化做出响应，与一个执行器相耦合，该执行器是一个叶绿素/叶黄素亚基，其中猝灭反应被催化。在植物中，传感器是 PSBS，而两个色素结合蛋白 Lhcb4（也称为 CP29）和 LHCII 是执行器。在藻类和苔藓中，与应激相关的光捕获蛋白（LHCSR）在单个亚基中包含传感器和执行器的双重功能。在这里，我们报告了从苔藓 Physcomitrella patens 中表达 lhcsr1 基因到几个拟南芥 npq4 突变体中，这些突变体缺乏 NPQ 活性所必需的 pH 感应 PSBS 蛋白。异源蛋白 LHCSR1 在拟南芥的类囊体中积累，并且可以部分恢复 NPQ 活性。除 PSBS 外，还缺乏特定叶黄素的双突变体的互补作用，使得可以分析发色团对 LHCSR 依赖性猝灭活性的要求。我们表明，NPQ 的部分恢复主要是由于与 P. patens 相比，拟南芥中 Zeaxanthin 的水平较低。除 PSBS 外，还缺乏 Zeaxanthin Epoxidase 的 npq2npq4 突变体，与拟南芥野生型相比，NPQ 恢复率高达 70%。此外，我们表明，叶黄素对于 LHCSR1 的折叠和猝灭活性不是必需的。简而言之，我们已经开发了一种使用各种拟南芥突变体进行异源表达来研究 LHCSR 蛋白功能的系统。
