Analysis and reconstitution of regulatory networks governing epidermal differentiation and polar root hair growth

表皮分化和极根毛生长调控网络的分析和重建

• 批准号: 264387517
• 负责人: Professor Dr. Jörg Kudla
• 金额: --
• 依托单位: Arbeitsgruppe Molecular Genetics and Cell Biology of Plants
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/264387517?language=en
• 关键词: Analysis; reconstitution; regulatory; networks; governing

Tip-focused growth represents an important mechanism to establish cellular polarity and is a fundamental process that underlies the growth of neuronal axon, fungal hyphae and plant root hairs. Root hairs are cylindrical shaped cellular protuberances of specialized epidermal cells. Polar tip growth of root hairs requires tight coordination and regulation of cytoskeleton dynamics, vesicle transport and ion fluxes. In root hairs a high-tip Ca2+ gradient has been shown to be essential for polar growth. Oscillatory Ca2+ increases are superimposed on this Ca2+ gradient and are likely to coordinate oscillatory tip growth and oscillatory fluctuations of reactive oxygen species (ROS) concentrations and pH values. The long-term goal of this project is to understand the molecular components and mechanisms underlying polar RH tip growth. Moreover, we intend to reconstitute such an oscillating network. Our preliminary data as well as published findings indicate that Ca2+ activated CBL/CIPK complexes and CDPKs simultaneously activate NADPH oxidases. Moreover, accumulating evidence also suggests Ca2+ independent phosphorylation as an alternative means of NADPH oxidase activation and a role of specific phosphatases in counteracting such activation. In this project we want to elucidate how exactly the different components of signaling network function and how they are dynamically interconnected and how quantitative changes in Ca2+ parameters govern information processing in root hairs. Considering our new findings and results obtained during the last years, we now specifically want to focus on the following research aims: (i) Explore the mechanisms and components contributing to the generation of the tip-focused Ca2+ gradient and Ca2+ oscillations in RHs. (ii) Elucidate the role of Ca2+ dependent phosphorylation in modulating RbohC activity. (iii) Investigate a potential contribution of Ca2+-independent phosphorylation to the regulation of RbohC. (iv) Explore the currently elusive components and mechanisms that bring about the required deactivation of RbohC in RHs. By combining the insights gained from project areas (i) – (iv), we subsequently intend to (v) characterize the complex interplay of these kinases and phosphatases in regulating RbohC and (vi) initiate the reconstitution of a regulatory network underlying the formation of Ca2+ oscillations and ROS signals heterologously in HEK293T cells. Complementary to these approaches, we intend to further advance our investigations of second messenger dynamics in RHs and continue to identify novel components that contribute to these processes. Collectively, our approaches should reveal detailed and fundamental insights of the complex interplay of these signaling components in growing RH.

尖端聚焦生长是细胞极性形成的重要机制，是神经元轴突、真菌菌丝和植物根毛生长的基础过程。根毛是特化表皮细胞的圆柱形细胞突起。根毛极尖的生长需要细胞骨架动力学、囊泡运输和离子通量的紧密协调和调节。在根毛中，高尖端Ca2+梯度已被证明是极性生长所必需的。振荡Ca2+增加叠加在Ca2+梯度上，并可能协调振荡尖端生长和活性氧（ROS）浓度和pH值的振荡波动。该项目的长期目标是了解极性RH尖端生长的分子成分和机制。此外，我们打算重建这样一个振荡网络。我们的初步数据和已发表的研究结果表明，Ca2+激活CBL/CIPK复合物和CDPKs同时激活NADPH氧化酶。此外，越来越多的证据还表明，Ca2+独立磷酸化是NADPH氧化酶激活的另一种方式，以及特定磷酸酶在抵消这种激活中的作用。在这个项目中，我们想要阐明信号网络的不同组成部分是如何准确地起作用的，它们是如何动态地相互连接的，以及Ca2+参数的定量变化是如何控制根毛中的信息处理的。考虑到我们在过去几年中获得的新发现和结果，我们现在特别想关注以下研究目标：(i)探索有助于RHs中尖端聚焦Ca2+梯度和Ca2+振荡产生的机制和成分。（ii）阐明Ca2+依赖性磷酸化在调节RbohC活性中的作用。（iii）研究Ca2+非依赖性磷酸化对RbohC调控的潜在贡献。（iv）探索目前难以捉摸的导致RHs中RbohC失活的成分和机制。通过结合从项目领域(i) - (iv)获得的见解，我们随后打算(v)表征这些激酶和磷酸酶在调节RbohC中的复杂相互作用，（vi）在HEK293T细胞中启动Ca2+振荡和ROS信号异种形成的调控网络的重建。作为这些方法的补充，我们打算进一步推进RHs中第二信使动力学的研究，并继续确定有助于这些过程的新成分。总的来说，我们的方法应该揭示这些信号成分在RH生长过程中复杂相互作用的详细和基本见解。