Department of Plant Physiology and Biochemistry, University of Hohenheim, Stuttgart, Germany.
Methods Mol Biol. 2024;2731:169-178. doi: 10.1007/978-1-0716-3511-7_13.
Extracellular alkalinization mediated by the inhibition of plasma membrane-located proton pumping ATPases hallmarks the initiation of defense signaling in plant cells. Early defense responses also include depolarization of the plasma membrane, increase in cytosolic Ca concentration, and an oxidative burst. Together these early signaling events lead to the activation of plant immunity. The transient alkalinization response is triggered by well-studied pathogen-derived and plant endogenous elicitors, including, for example, bacterial flagellin, fungal chitin, and tomato systemin in both model and agronomic species. Employing cell suspension cultures, extracellular alkalinization can be easily assessed by measuring the elicitor-induced pH changes of the cultivating medium. Here, we provide a protocol for an improved alkalinization assay in a system which is able to simultaneously monitor multiple samples, and fully automatically transfer customizable real-time pH records. In this system flagellin, chitin and systemin elicit robust time- and dose-dependent responses, proving a powerful tool for assessing plant early defense signaling.
细胞外碱化由质膜定位质子泵 ATP 酶的抑制介导，是植物细胞防御信号起始的标志。早期防御反应还包括质膜去极化、胞质 Ca 浓度增加和氧化爆发。这些早期信号事件共同导致植物免疫的激活。瞬态碱化反应是由研究充分的病原体衍生和植物内源性激发子触发的，包括细菌鞭毛蛋白、真菌几丁质和番茄系统素等，在模式和农业物种中均有研究。在细胞悬浮培养中，可以通过测量培养介质中激发子诱导的 pH 值变化来轻松评估细胞外碱化。在这里，我们提供了一种在能够同时监测多个样本并自动传输可定制的实时 pH 记录的系统中进行改良碱化测定的方案。在该系统中，鞭毛蛋白、几丁质和系统素可引发强大的时间和剂量依赖性反应，是评估植物早期防御信号的有力工具。
