Stress Hormone Signal Transduction In Arabidopsis

项目总结 脱落酸(ABA)是拟南芥中的一种中枢应激激素，它下调细胞增殖和 导致细胞周期停滞。下调细胞增殖的早期信号转导网络是关键 控制有丝分裂及其错误调控的重要性与许多人类疾病有关。长的- 这项研究的长期目标是实现对早期调节事件网络的量化理解 利用强大的拟南芥保卫细胞系统传递脱落酸信号。我们将重新定义 确定了控制早期ABA信号核心的关键细胞信号机制，包括 ABA受体、PP2C蛋白磷酸酶和SnRK2&钙(Ca~(2+))依赖性蛋白激酶。目标： 真核生物PP2C磷酸酶活性的直接调节蛋白还不是很清楚。我们会 一种新发现的介导ABA受体的早期PP2C调控环的功能 信令健壮性。这个循环包括PP2C相互作用的GDP/GTP交换因子1(GEF1)，它 激活ROP10和ROP11，进而通过将被阐明的机制稳定PP2C活性。 此外，PP2Cs保护GEF1和钙依赖的蛋白激酶(CPKs)引起ABA介导的GEF1 通过未知的机制进行降解。ABA对这个新发现的控制回路的调节将在功能上 这一环在ABA信号转导过程中调节钙专一性的作用将被描述为 生化、遗传学、生物物理学、体内时间分辨、保卫细胞信号和结构分析。 以前的研究表明，PP2C的激活下调了OST1-SnRK2蛋白激酶的表达， 它协调下游的ABA信号，需要SnRK2激酶激活环的磷酸化。 然而，无论OST1-SnRK2激酶的激活是通过自身磷酸化还是通过其他方式完成的 蛋白激酶仍不清楚。我们最近的结果表明，去磷酸化的OST1不能重新激活 它本身。通过遗传冗余筛选，我们已经确定了激活OST1的MAPKKK，in 与其他几个测试的ABA信号通路形成对比。我们将调查假设这些MAPKKK 代表了人们长期寻求的ABA信号核心中OST1-SnRK2蛋白激酶的激活机制。 通过我们最近开发的创新的全基因组人工microRNA筛选平台，a 将采用系统生物学方法，筛选关键的(部分)冗余基因， 在ABA信令网络中发挥作用。新分离和确认的脱落酸突变株 我们将结合系统生物学、遗传学和功能分析来描述这些反应。 描述它们在动态ABA信号网络中的直接作用。 这项研究将对PP2C磷酸酶和SnRK2激酶的调控有新的理解 钙离子特异性信号机制，这是许多细胞信号过程和 疾病状态，并将揭示早期ABA信号核心的新机制。

PROJECT SUMMARY Abscisic acid (ABA) is a central stress hormone in Arabidopsis that down-regulates cell proliferation and causes cell cycle arrest. Early signal transduction networks that down-regulate cell proliferation are of key importance for controlling mitogenesis and their mis-regulation is linked to many human diseases. The long- term goal of this research is to achieve a quantitative understanding of the network of events that mediate early abscisic acid signaling making use of the potent Arabidopsis guard cell system. We will characterize newly identified key cellular signaling mechanisms hypothesized to control the early ABA signaling core consisting of ABA receptors, PP2C protein phosphatases and SnRK2 & calcium (Ca2+)-dependent protein kinases. Aims: I. Direct regulatory proteins of eukaryotic PP2C phosphatase activity are not well-understood. We will characterize the functions of a newly identified early PP2C control loop that can mediate ABA receptor-PP2C signaling robustness. This loop includes the PP2C interacting GDP/GTP exchange factor 1 (GEF1), which activates ROP10 and ROP11, which in turn stabilize PP2C activity via mechanisms that will be elucidated. Moreover, PP2Cs protect GEF1 and Ca2+-dependent protein kinases (CPKs) cause ABA-mediated GEF1 degradation via unknown mechanisms. ABA regulation of this newly identified control loop will be functionally dissected and the role of this loop in mediating Ca2+ specificity during ABA signaling will be characterized by biochemical, genetic, biophysical, in vivo time-resolved, guard cell signaling and structural analyses. II. Previous studies have shown that activation of the PP2C down-regulated OST1-SnRK2 protein kinases, which orchestrate downstream ABA signaling, requires phosphorylation of the SnRK2 kinase activation loop. However, whether this activation of OST1-SnRK2 kinases is accomplished by auto-phosphorylation or by other protein kinases has remained unclear. Our recent results show that dephosphorylated OST1 cannot re-activate itself. Via a genetic redundancy screen, we have identified MAP-kinases (MAPKKKs) that activate OST1, in contrast to several other ABA signaling kinases tested. We will investigate the hypothesis that these MAPKKKs represent the long-sought activation mechanism of OST1-SnRK2 protein kinases in the ABA signaling core. III. Through our recent development of an innovative genome-wide artificial microRNA screening platform, a systems biological approach will be pursued that enables screening for key (partially) redundant genes that function in the ABA signaling network. Newly isolated and confirmed mutants with altered abscisic acid responses will be characterized through a combination of systems biology, genetics, and functional analyses towards characterizing their direct roles in the dynamic ABA signaling network. This research will result in a new understanding of PP2C phosphatase and SnRK2 kinase regulation and Ca2+ specificity signaling mechanisms, which are fundamental to numerous cell signaling processes and disease states, and will reveal novel mechanisms in the early ABA signaling core.