关键词: 氧爆发, 单细胞, 细胞壁, 微电极, 实时监测

Abstract: Oxidative burst by rapid, transient generation of reactive oxygen species (ROS) is a fundamental process for plant disease defense and signal transduction. ROS can be protective or toxic depending on their concentrations, and critical balance between ROS production and scavenging is regulated by complex and perfect mechanisms in plant cells. However, the process of rapid and transient ROS burst from plant cells and their regulatory mechanisms are far from being completely understood, owing to the methodological obstacles in real-time monitoring such a fast process. In this work, we showed the application of microelectrode electrochemistry with high spatiotemporal resolution to monitor the kinetics of ROS burst from single plant cell and to investigate the regulatory functions of cell wall. An agarose chip with numerous microwell arrays was fabricated to efficiently trap individual plant cells for electrochemical detection. The results revealed that the oxidative burst from single Arabidopsis thaliana cells is characterized by numerous transient spikes with fast kinetics. Further experiments indicated that intact plant cells released much less H₂O₂ molecules with faster kinetics to the extracellular environment compared with single protoplasts, while the ROS level could be recovered when the activity of cell wall peroxidase (POD) was inhibited. The results provide evidence that cell wall participates in regulating transient oxidative burst transient by a mechanism involving cell wall POD machinery at single cell level. We think that the results presented here would facilitate better understanding on ROS burst and their regulatory mechanisms.

Key words: oxidative burst, single plant cell, cell wall, microelectrochemical sensor, real-time detection