Regulation of xanthophyll conversion and the function of zeaxanthin in land plants

在陆地植物中，叶黄素玉米黄质（Zx）作为光系统II （PSII）中多余光能作为热量（非光化学猝灭，NPQ）消散的调节器和类囊体膜中的抗氧化剂具有中心的光保护功能。在黄叶素循环的去环氧化反应（由腔内定位的Vx去环氧化酶催化）中，在强光（HL）下紫黄质（Vx）在类囊体膜中形成Zx，在弱光或黑暗的环氧化反应中，由基质定位的Zx环氧化酶（ZEP）催化，再转化回Vx。ZEP活性决定了HL胁迫后叶绿体中高Zx量维持的时间，从而决定了Zx依赖性NPQ的维持时间。我们已经证明，随着HL胁迫和PSII光抑制的增加，ZEP活性逐渐下调，并且在PSII光抑制时，ZEP蛋白与D1蛋白一起被降解。HL诱导的ZEP活性下调确保了高水平的Zx在长时间HL期的反应中被保留，从而允许NPQ的有效再激活（或保留），从而在波动光条件下的短暂弱光期后（或期间）进行光保护。因此，Zx被认为是叶绿体对先前光氧化应激“记忆”的关键成分。因此，详细了解ZEP活性的功能和（光）调节对于理解植物对HL或波动光的光保护NPQ机制的适应至关重要。在前期工作中，我们提供了证据，证明HL诱导的ZEP失活可能是由过氧化氢引起的，过氧化氢是HL胁迫下叶绿体中经常形成的活性氧（ROS）。对ZEP体内和体外活性的分析进一步表明，ZEP活性可能需要在类囊体膜和未知的辅助因子中进行特定的蛋白质相互作用。为了在分子水平上详细描述ZEP的特性和调控，我们将在计划的项目中解决三个关键问题：(i) ZEP在类囊体膜中的确切定位和相互作用伙伴，（ii） hl诱导ZEP失活的分子基础，以及（iii） ZEP的详细生化特性，重点是必需的辅因子。这种方法可以在分子水平上理解ZEP在光保护中的作用。

The xanthophyll zeaxanthin (Zx) serves central photoprotective functions in land plants as regulator of the dissipation of excess light energy as heat (non-photochemical quenching, NPQ) in photosystem II (PSII) and as antioxidant in the thylakoid membrane. Zx is formed in the thylakoid membrane from violaxanthin (Vx) under high light (HL) in the de-epoxidation reactions (catalyzed by the lumen-localized Vx de-epoxidase) of the xanthophyll cycle and is reconverted back to Vx under low light or in darkness in the epoxidation reactions, catalyzed by the stroma-localized Zx epoxidase (ZEP). ZEP activity determines the period of the maintenance of a high Zx amount, and thus of Zx-dependent NPQ, in the chloroplast after HL stress. We have shown that ZEP activity is gradually down-regulated in response to increasing HL stress and photoinhibition of PSII, and that ZEP protein is degraded concomitant with the D1 protein upon PSII photoinhibition. The HL-induced down-regulation of ZEP activity ensures that high levels of Zx are retained in response to prolonged HL periods to allow for efficient reactivation (or retention) of NPQ and thus photoprotection after (or during) transient low light periods under fluctuating light conditions. Therefore, Zx is supposed to be a key component in the “memory” of the chloroplast with respect to preceding photo-oxidative stress. Detailed knowledge of the functioning and (light) regulation of ZEP activity is thus central for the understanding of the acclimation of photoprotective NPQ mechanisms in plants to HL or to fluctuating light. In preliminary work, we provided evidence that the HL-induced inactivation of ZEP may be caused by hydrogen peroxide, a reactive oxygen species (ROS) that is frequently formed in chloroplasts under HL stress. Analyses of the in vivo and in vitro activity of ZEP further showed that ZEP activity likely requires specific protein interactions in the thylakoid membrane and yet unknown cofactors. To characterize the properties and regulation of ZEP in detail at the molecular level, we will address three key issues in the planned project: (i) the exact localization and interaction partners of ZEP in the thylakoid membrane, (ii) the molecular basis of HL-induced ZEP inactivation, and (iii) the detailed biochemical properties of ZEP with focus on essential cofactors. This approach should allow the understanding of the functioning of ZEP in photoprotection at the molecular level.