P05 –Comparative exploration of KEA3 function in PMF partitioning and photoprotection

叶绿体PMF是通过光驱动质子耦合电子在类囊体膜上传递而产生的。这产生了膜电位（∆ψ）和质子浓度梯度（∆pH）。它们对PMF的贡献部分受类囊体离子转运过程的支配。PMF的∆pH分量，或者更具体地说，管腔质子浓度决定了用于光化学的光能的比例。来自拟南芥（Arabidopsis thaliana）的数据表明，类囊体K+交换反转运蛋白3 （KEA3）在光照波动下对维管植物光合作用的灵活性起着重要作用。KEA3可以感知叶绿体基质的能量状态，并相应地调节光保护能量依赖猝灭（qE）机制的水平。如果能态较低，KEA3被激活以降低管腔的质子浓度并关闭qE。虽然KEA3活性会影响PMF的组成并降低pH值，但对PMF的大小及其能量功能影响不大。先前，我们可以证明去调控的KEA3活性可以导致短暂的更高的CO2固定率。这表明在自然界中发现的动态光照条件下，KEA3调控可能是增强光合作用的一个目标。深入了解KEA3调控及其与复杂光合反应网络的相互作用将是进一步探索这一途径的关键。在GoPMF的框架内，我们试图通过比较的方法来扩展我们对KEA3在PMF分配及其调控中的功能的了解：(1)通过测定单细胞藻莱茵衣藻（Chlamydomonas reinhardtii, Chlamydomonas）中KEA3的活性，旨在了解KEA3功能和调控的进化影响；(2)通过比较拟南芥和衣藻间质能量动态，旨在表征环境变化如何影响叶绿体能量状态，并将其与KEA3活性联系起来；(3)通过确定KEA3调节因子。我们将把基质能量状态的变化与KEA3活性联系起来，最后（iv）通过在植物中表达修饰的KEA3版本，我们将通过实验证实这些联系，并利用我们关于KEA3调节PMF的知识来尝试改善光合性能和生长（参见图形摘要P05）。我们的工作将是目标GoPMF 2的组成部分，同时也有助于目标1、3、4和5。

The chloroplast PMF is produced via light-driven proton-coupled electron transport across the thylakoid membrane. This generates both a membrane potential (∆ψ) and a proton concentration gradient (∆pH). Their contribution to PMF is governed in part by thylakoid ion transport processes. The ∆pH component of the PMF or more specifically, the lumenal proton concentration determines the fraction of light energy that is used for photochemistry. Our data from Arabidopsis thaliana (Arabidopsis) show that the thylakoid K+-exchange antiporter 3 (KEA3) plays a fundamental role for the flexibility of vascular plant photosynthesis during light fluctuations. KEA3 can sense the energy state of the chloroplast stroma and adjusts levels of the photoprotective energy dependent quenching (qE) mechanism in response. If the energy state is low, KEA3 gets activated to decrease the proton concentration of the lumen and shuts off qE. While KEA3 activity affects PMF composition and decreases the ∆pH component, it appears to have little effect on PMF size and thus its energetic function. Previously, we could demonstrate that de-regulated KEA3 activity can result in transiently higher CO2 fixation rates. This proposes KEA3 regulation as a putative target for enhancing photosynthesis under the dynamic light conditions found in nature. An in-depth understanding of KEA3 regulation and its interaction with the complex network of photosynthetic reactions will be pivotal to further explore this avenue. Within the framework of GoPMF, we seek to extent our knowledge of KEA3 function in PMF partitioning and its regulation via a comparative approach: (i) By measuring KEA3 activity in the unicellular alga Chlamydomonas reinhardtii (Chlamydomonas), we aim to understand evolutionary effects on KEA3 function and regulation, (ii) by comparing stromal energy dynamics between Arabidopsis and Chlamydomonas, we aim to characterize how environmental changes affect the chloroplast energy state and relate this to KEA3 activity , (iii) by determining KEA3 regulating factors, we will link changes in the stromal energy state with KEA3 activity and finally (iv) by expressing modified KEA3 versions in planta, we will confirm these links experimentally and use our knowledge on PMF modulation by KEA3 for attempts to improve photosynthetic performance and growth (see also Graphical Abstract P05). Our work will be integral for the aim GoPMF 2 and additionally contribute to aims 1, 3, 4 and 5.