Julius-von-Sachs Institute of Biosciences, Molecular Plant Physiology and Biophysics, University of Würzburg, Julius-von-Sachs-Platz 2, 97082 Würzburg, Germany.
Plant J. 2010 May;62(3):367-78. doi: 10.1111/j.1365-313X.2010.04155.x. Epub 2010 Jan 25.
The perception of microbes by plants involves highly conserved molecular signatures that are absent from the host and that are collectively referred to as microbe-associated molecular patterns (MAMPs). The Arabidopsis pattern recognition receptors FLAGELLIN-SENSING 2 (FLS2) and EF-Tu receptor (EFR) represent genetically well studied paradigms that mediate defense against bacterial pathogens. Stimulation of these receptors through their cognate ligands, bacterial flagellin or bacterial elongation factor Tu, leads to a defense response and ultimately to increased resistance. However, little is known about the early signaling pathway of these receptors. Here, we characterize this early response in situ, using an electrophysiological approach. In line with a release of negatively charged molecules, voltage recordings of microelectrode-impaled mesophyll cells and root hairs of Col-0 Arabidopsis plants revealed rapid, dose-dependent membrane potential depolarizations in response to either flg22 or elf18. Using ion-selective microelectrodes, pronounced anion currents were recorded upon application of flg22 and elf18, indicating that the signaling cascades initiated by each of the two receptors converge on the same plasma membrane ion channels. Combined calcium imaging and electrophysiological measurements revealed that the depolarization was superimposed by an increase in cytosolic calcium that was indispensable for depolarization. NADPH oxidase mutants were still depolarized upon elicitor stimulation, suggesting a reactive oxygen species-independent membrane potential response. Furthermore, electrical signaling in response to either flg22 or elf 18 critically depends on the activity of the FLS2-associated receptor-like kinase BAK1, suggesting that activation of FLS2 and EFR lead to BAK1-dependent, calcium-associated plasma membrane anion channel opening as an initial step in the pathogen defense pathway.
植物对微生物的感知涉及高度保守的分子特征，这些特征在宿主中不存在，通常被称为微生物相关分子模式（MAMPs）。拟南芥模式识别受体 FLAGELLIN-SENSING 2（FLS2）和 EF-Tu 受体（EFR）是遗传上研究充分的范例，它们介导对细菌病原体的防御。通过其同源配体（细菌鞭毛蛋白或细菌延伸因子 Tu）刺激这些受体，会导致防御反应，并最终增加抗性。然而，对于这些受体的早期信号通路知之甚少。在这里，我们使用电生理学方法对其进行原位表征。与释放带负电荷的分子一致，用微电极刺穿的叶片细胞和 Col-0 拟南芥根毛的电压记录显示，对 flg22 或 elf18 的反应均会迅速且剂量依赖性地引起质膜电位去极化。使用离子选择性微电极，在施加 flg22 和 elf18 时会记录到明显的阴离子电流，表明两种受体中的每一种启动的信号级联都集中在相同的质膜离子通道上。钙成像和电生理测量的组合表明，去极化被细胞质钙增加所叠加，该增加对于去极化是必不可少的。在激发剂刺激下，NADPH 氧化酶突变体仍然去极化，表明这是一种与活性氧无关的膜电位反应。此外，对 flg22 或 elf18 的电信号响应都严重依赖于 FLS2 相关受体样激酶 BAK1 的活性，这表明 FLS2 和 EFR 的激活导致 BAK1 依赖性、钙相关质膜阴离子通道打开，作为病原体防御途径的初始步骤。
