Structural and functional principles of low potassium signaling and the integration of nutrient sensing and adaptation in Arabidopsis

有效地感知和吸收各种营养物质，如钾、硝酸盐和铵，对植物来说是必不可少的。植物产生了一系列的适应性反应，这些反应是由感知外部养分供应的传感系统触发的。然而，植物如何感知和适应波动的营养条件的分子机制才刚刚开始出现。K+吸收的关键是K+通道AKT1和它的钙(Ca+)依赖的激活CBL1/CBL9-CIPK23复合体。然而，尽管K+的感知和吸收具有基本的重要性，但植物感知低K+条件的机制以及钙离子信号在这一过程中的潜在贡献在很大程度上仍然是个谜。与AKT1类似，硝酸盐转运蛋白NPF6.3(也称为NRT1.1/CHL1.5)受CBL1/9-CIPK23调控，已被报道起硝酸盐转运蛋白的作用。此外，CIPK23还调节着其他几个离子通道和转运体，使CIPK23成为拟南芥中的营养感知蛋白，可能在协调和整合植物的一般营养动态平衡方面发挥核心作用。然而，允许CIPK23发挥这种核心和复杂作用的机制基础是完全未知的。我们的初步工作和数据表明：(A)K+通道AKT1本身形成或是拟南芥中主要的低K+感受器-受体的一部分，(B)鉴定了CBL1/9-CIPK23复合体对AKT1的磷酸化对其传感和运输功能的需求，(C)分离和鉴定了具有解偶联传感和运输缺陷的新的AKT1突变等位基因，以及CIPK23中多个顺式和反式磷酸化位点，以及(D)首次鉴定和表征了植物根中低K+诱导的钙信号的发生。根据我们的初步数据，该项目的具体目标是：(I)识别和表征与低K+感知有关的成分和信号事件，(Ii)阐明这些过程背后的结构和机制原理，(Iii)研究调节CIPK23功能和活性的结构和调控机制，以实现营养感知整合，以及(Iv)了解这些适应过程如何转化为对根系生长和养分分配的适应性控制和调节。通过结合我们现有的突变体，新分离的将K+感知与K+运输分离的抑制系，并结合表型植物分析、生化/结构方法和细胞生物学分析，我们打算识别和表征初级低K+感知的基本分子机制和结构原理，从而实现对植物生长适应的理解，并了解植物如何整合以及允许协调和严格调控多种必需营养的吸收和动态平衡的多种机制。

Efficient sensing and uptake of various nutrients like potassium, nitrate and ammonium is essential for plants. Plants developed a wide range of adaptive responses triggered by sensing systems that perceive the external nutrient availability. However, the molecular mechanisms how plants sense and adapt to fluctuating nutrient conditions are only beginning to emerge.Crucial for K+ uptake is the K+ channel AKT1 and its calcium (Ca2+) dependent activation by CBL1/CBL9-CIPK23 complexes. However, despite the fundamental importance of K+ sensing and uptake, the mechanisms how plants sense low K+ conditions and the potential contribution of Ca2+ signaling to this process have remained largely enigmatic.Similar to AKT1, the nitrate transporter NPF6.3 (also known as NRT1.1/CHL1.5) is regulated by CBL1/9-CIPK23 and has been reported to function as nitrate transceptor. In addition, the kinase CIPK23 regulates several other ion channels and transporters, establishing CIPK23 as nutrient sensing kinase in Arabidopsis that likely fulfills a central role in coordinating and integrating the general nutritional homeostasis in plants. However, the mechanistic basis that would allow CIPK23 to fulfill such central and complex role is completely unknown. Our preliminary work and data, indicated (a) that the K+ channel AKT1 itself forms or is part of the primary low K+ sensor-receptor in Arabidopsis, (b) identified the requirement of AKT1 phosphorylation by CBL1/9-CIPK23 complexes for its sensing and transport function, (c) isolated and identified novel akt1 mutant alleles with uncoupled sensing- and transport defects as well as multiple cis- and trans-phosphorylation sites in the regulatory kinase CIPK23 and (d) for the first time identified and characterized the occurrence of low K+ induced Ca2+ signals in plants roots. Based on our preliminary data the proposed project specifically aims to: (i) identify and characterize the components and signaling events that are involved in low K+ sensing, (ii) elucidate the structural and mechanistic principles underlying these processes, (iii) investigate the structural and regulatory mechanisms that modulate CIPK23 function and activity to allow nutrient sensing integration and (iv) to understand how these adaptation processes are translated into the adaptive control and regulation of root growth and nutrient distribution. By combining our available mutants, newly isolated suppressor lines that uncouple K+ sensing from K+ transport and by using a combination of phenotypic plant analyses, biochemical / structural approaches, and cell biological assays, we intend to identify and characterize the fundamental molecular mechanisms and structural principles that underlie primary low K+ sensing, the resulting growth adaptation of plants and to achieve an understanding of how plants integrate and the multiple mechanisms that allow for coordinated and tightly regulated uptake and homeostasis of multiple essential nutrients.