Collaborative Research: Redox Regulation of Protein Kinase Functions in Guard Cell Signaling

气孔是植物叶片表面的气孔，植物通过气孔释放氧气和水蒸气，并从大气中吸收二氧化碳。气孔孔径的大小（以及二氧化碳和水蒸气与大气的交换）是由专门的细胞控制的。这些细胞（称为保护细胞）会根据环境条件改变形状，并对植物内部的一系列信号系统做出反应。本研究的目的是了解活性氧种类和保护细胞中还原和氧化生物化学的变化如何调节酶系统（蛋白激酶），该酶系统控制气孔运动以响应环境线索。气孔的功能影响植物对环境胁迫（包括温度和干旱）的响应，该研究将为作物改良（抗逆性）植物的合理选育提供指导。该项目将使大学和高中学生（包括妇女和代表性不足群体的成员）能够在生物和化学界面的高通量科学新兴领域进行跨学科培训。该项目将采用包括分子生物学、生物化学、遗传学和分析化学在内的多种方法来解决植物生物学中的一个关键问题，即活性氧和氧化还原变化在调节激酶活性和气孔运动中的作用。核心假设是，关键蛋白激酶中半胱氨酸残基的氧化还原依赖性修饰提供了一种调节气孔信号中激酶功能的通用方法。这一假设将通过追求两个特定目标来验证：1)确定甘蓝型油菜蔗糖非发酵相关激酶的半胱氨酸修饰。该实验将能够全面分析半胱氨酸对脱落酸（一种干旱胁迫激素）和参与植物气孔入口的细菌鞭毛蛋白肽的响应。2)研究半胱氨酸修饰在气孔运动中的作用及其对激酶磷酸化的影响。该实验将确定对气孔运动至关重要的半胱氨酸修饰，并分析调节激酶活性的半胱氨酸氧化还原变化。该项目将揭示气孔运动的新调控机制，并将有助于氧化还原调节作为细胞调节重要信号成分和过程的通用机制的新兴概念。

Stomata are pores on the leaf surface through which plants release oxygen and water vapor and take up carbon dioxide from the atmosphere. The size of the stomatal aperture (and hence the exchange of carbon dioxide and water vapor with the atmosphere) is controlled by specialized cells. These cells (called guard cells) change their shape in response to environmental conditions and are responsive to a range of signaling systems within the plant. The goal of this research is to understand how reactive oxygen species and changes in the reducing and oxidizing biochemistry in guard cells regulate an enzyme system (protein kinases) that controls stomatal movements in response to environmental cues. The function of stomata impacts the response of plants to environmental stress (including temperature and drought) and this research should lead to the rational breeding of improved (stress resistant) crop plants. The project will enable cross-disciplinary training of college and high school students (including women and members of underrepresented groups), in emerging fields of high-throughput science at the interface of biology and chemistry. The project will use multiple approaches including molecular biology, biochemistry, genetics and analytical chemistry to tackle a critical problem in plant biology, i.e., the functions of reactive oxygen species and redox changes in regulating kinase activities and stomatal movements. The central hypothesis is that redox-dependent modification of cysteine residues in key protein kinases provides a versatile means of regulating kinase function in stomatal signaling. This hypothesis will be tested by pursuing two specific objectives: 1) To determine the cysteine modifications of Brassica napus sucrose non-fermenting related kinases. The experiments will enable comprehensive analysis of cysteine modifications in response to abscisic acid (a drought stress hormone) and a bacterial flagellin peptide that is involved in stomatal entrance of plant pathogens. 2) To determine cysteine modifications functional in stomatal movement and how they affect kinase phosphorylation. The experiments will identify the cysteine modifications essential for stomatal movement and analyze cysteine redox changes that regulate kinase activity. This project will reveal novel regulatory mechanisms underlying stomatal movements and will contribute to the emerging concept of redox modulation as a versatile mechanism by which cells regulate important signaling components and processes.