Ion Channel Regulation in Higher Plants

高等植物中的离子通道调节

• 批准号: 0077791
• 负责人: Julian Schroeder
• 金额: $ 52万
• 依托单位: University of California-San Diego
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-10-01 至 2005-09-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0077791&HistoricalAwards=false
• 关键词: Ion; Channel; Regulation; Higher; Plants

Stomatal pores in the epidermis of leaves regulate the diffusion of CO2 into leaves for photosynthetic carbon fixation and control water loss of plants via transpiration to the atmosphere. A network of signal transduction mechanisms in guard cells integrate environmental stimuli to regulate stomatal apertures for optimization of plant growth under diverse conditions. Control of stomatal movements in response to environmental stress conditions is an important factor in determining crop productivity and plant water loss during drought. The long-term goal of this project is to achieve a detailed quantitative understanding of the network of signal transduction events that regulate stomatal movements using the easily manipulated plant, Arabidopsis thaliana. Guard cells provide a powerful system for quantitative and time-resolved dissection of the complex molecular machinery mediating specificity in early plant signaling cascades. During the preceding funding period a signal transduction model has been developed, suggesting that several types of both positively- and negatively-regulating protein kinases and PP1- or PP2A-type protein phosphatases play central roles in mediating stomatal closing by the phytohormone, abscisic acid (ABA). However, the genes encoding these protein kinases and PP1/2A-type phosphatases, as well as their locations and functions within cascades and their downstream targets remain largely unknown. From an Arabidopsis guard cell cDNA libraries, the cDNAs that encoded for the receptor-like kinases (RLKs), the Ca2+-dependent protein kinases (CDPKs), and the catalytic PP2A subunit have been isolated.. The large gene families and redundancies of these signal transducers in plants have limited direct genetic characterization of individual kinases and PP2As. Direct functional analyses of guard cell-expressed members will now allow determination of their redundancies and of their precise functions in signal transduction. Preliminary studies show that an insertional mutation in a regulatory PP2A subunit causes recessive ABA insensitivity in guard cells. Furthermore, disruption of a receptor kinase abolishes light-induced stomatal opening, illustrating the importance and feasibility of the proposed research. The following hypotheses will be tested in this project: That both positively- and negatively-regulating protein kinases and PP2As function in ABA-induced stomatal closing. That CDPKs control ABA-induced stomatal closing. That specific PP2As may function either as positive or negative regulators of ABA signaling. To test these, the following specific aims will be done: Insertional disruption mutant alleles will be identified and transgenic repression lines will be generated in the isolated guard cell RLK, CDPK and catalytic PP2A genes. Stomatal movement analyses will be pursued to characterize phenotypic responses to ABA and light stimuli in disruption mutants or double/multi mutants of redundant homologous CDPKs and PP2As. Effects of two to three kinase and PP2A mutations with the strongest stomatal phenotypes will be characterized on stimulus-induced regulation of guard cell anion or K+ channels and signal-induced cytosolic Ca2+ changes. Epistasis analyses with known guard cell signaling mutants will be pursued to determine the relative sequence of events in the guard cell signaling network. Selected CDPKs/PP2As will be analyzed for modulation of cloned guard cell ion channels. Furthermore screens for guard cell-expressed interacting proteins will be pursued for one of the regulators. The knowledge gained from these studies will reveal novel fundamental mechanisms by which a network of kinases and PP2As control signal transduction and mediate signaling specificity in a plant cell. Furthermore, these studies will lead to elucidation of key signal transduction mechanisms by which plants balance CO2 influx into leaves and transpirational water loss and may contribute to future strategies for manipulating gas exchange in plants.

叶片表皮中的气孔孔调节二氧化碳扩散到叶片中，以通过蒸腾向大气中的蒸腾作用，以进行光合碳固定和控制植物的水损失。警卫细胞中信号转导机制的网络整合了环境刺激，以调节气孔孔，以优化不同条件下植物生长。控制气孔运动对环境压力条件的控制是确定干旱期间作物生产力和植物水分流失的重要因素。该项目的长期目标是实现对信号转导事件网络的详细定量理解，该网络使用易于操纵的植物拟南芥（Thaliana）来调节气孔运动。护罩细胞为复杂分子机械的定量和时间分辨的解剖提供了强大的系统，该系统介导了早期植物信号级联的特异性。 在前面的资金期间，已经开发了信号转导模型，这表明几种类型的阳性和负调节的蛋白激酶以及PP1-或PP2A型蛋白磷酸酶在介导植物符号，aba-abcisic Acid（ABA）中介导的气孔闭合中起着核心作用。然而，编码这些蛋白激酶和PP1/2A型磷酸酶的基因及其在级联反应中的位置和功能仍然很大程度上尚不清楚。从拟南芥防护细胞cDNA库中，编码为受体样激酶（RLK）编码的cDNA，Ca2+依赖性蛋白激酶（CDPK）和催化PP2A亚基已隔离。现在，对后卫细胞表达成员的直接功能分析将允许确定其冗余及其在信号转导中的精确功能。初步研究表明，调节性PP2A亚基中的插入突变会导致警卫细胞中隐性的ABA不敏感性。此外，受体激酶的破坏消除了光引起的气孔开口，说明了拟议研究的重要性和可行性。 在该项目中将测试以下假设：在ABA诱导的气孔闭合中，阳性和负调节的蛋白激酶和PP2AS功能。 该CDPK控制ABA诱导的气孔闭合。该特定的PP2A可以作为ABA信号的正调节剂或负调节剂的作用。 为了测试这些这些，将完成以下特定目标：将识别插入式破坏突变等位基因，并在分离的防护细胞RLK，CDPK和催化PP2A基因中生成转基因抑制线。将进行气孔移动分析，以表征对ABA的表型反应和冗余同源CDPK和PP2A的双/多突变体中对ABA和光刺激的反应。具有最强气孔表型的两到三个激酶和PP2A突变的影响将在刺激诱导的防护细胞阴离子或K+通道的调节以及信号诱导的胞质CA2+变化方面进行表征。将使用已知的后卫细胞信号突变体进行上毒分析，以确定后卫细胞信号网络中事件的相对顺序。将分析选定的CDPK/PP2AS，以调制克隆的防护细胞离子通道。此外，将为其中一个调节剂推出用于防护细胞表达的相互作用蛋白的筛选。从这些研究中获得的知识将揭示新的基本机制，通过这些机制，激酶和PP2AS控制信号转导网络并介导植物细胞中的信号特异性。此外，这些研究将导致关键信号转导机制阐明植物平衡二氧化碳流入叶片和蒸发水分流失的机制，并可能有助于未来操纵植物中气体交换的策略。