Functional characterization of a calcium-permeable cation channel in plants

为了对抗非生物和生物胁迫，控制植物防御和抗性反应的复杂信号网络已经进化。对刺激-反应耦合的核心要素的理解对于有针对性地改良作物至关重要。植物对大多数非生物和生物胁迫因子的最早反应是胞内游离钙([Ca~(2+)]Cyt)的升高，这是由钙离子通透离子通道的激活所介导的。我们对这些通道蛋白的分子同一性的了解仍然非常初级，这是我们理解植物信号和逆境反应的主要障碍。先前在液泡膜中发现了钙离子通透性的双孔通道1，初步分析表明它参与了钙离子依赖的过程。本项目旨在阐明该通道在刺激反应中的作用，并分析在什么情况下需要该通道来产生[Ca~(2+)]Cyt信号。此外，将结合电生理学和细胞生物学方法分析通道的负反馈调节和第二信使分子激活的模式，包括使用Aequorin和黄色Cameleon技术的[Ca2+]Cyt成像。从这些实验中获得的知识将有助于我们理解钙离子的动态平衡和信号转导，这最终可能用于设计具有更有效逆境反应的作物。

To counter abiotic and biotic stress, intricate signalling networks that govern the initation of defense and resistance responses have evolved in plants. The understanding of the central elements of stimulus-response coupling is vital for a targeted improvement of crops. A very early response of plants to most abiotic and biotic stress factors is an elevation of cytosolic free calcium ([Ca2+]cyt), which is mediated by the activation of Ca2+-permeable ion channels. Our knowledge on the molecular identity of those channel proteins is still very rudimentary, which presents a major obstacle in our understanding of signalling und stress responses in plants. The Ca2+-permeable Two Pore Channel 1 has previously been identified in the vacuolar membrane and initial analyses suggested its involvement in Ca2+-dependent processes. This project aims to elucidate the role of this channel in stimulus responses and to analyse the circumstances under which this channel is required in the generation of [Ca2+]cyt signals. Furthermore, the mode of negative feedback regulation of the channel and its activation by second messenger molecules will be analysed by a combination of electrophysiological and cell biological approaches, including [Ca2+]cyt imaging using aequorin and Yellow Cameleon techniques. The knowledge gained from those experiments will aid in our understanding of Ca2+ homeostasis and signalling, which may eventually be used in the design of crops with more efficient stress responses.