The Role of ROS Homeostasis in ABA-Induced Guard Cell Signaling

Abstract

The hormonal and environmental regulation of stomatal aperture is mediated by a complex signaling pathway found within the guard cells that surround stomata. Abscisic acid (ABA) induces stomatal closure in response to drought stress by binding to its guard cell localized receptor, initiating a signaling cascade that includes synthesis of reactive oxygen species (ROS). Genetic evidence in Arabidopsis indicates that ROS produced by plasma membrane respiratory burst oxidase homolog (RBOH) enzymes RBOHD and RBOHF modulate guard cell signaling and stomatal closure. However, ABA-induced ROS accumulates in many locations such as the cytoplasm, chloroplasts, nucleus, and endomembranes, some of which do not coincide with plasma membrane localized RBOHs. ABA-induced guard cell ROS accumulation has distinct spatial and temporal patterns that drive stomatal closure. Productive ROS signaling requires both rapid increases in ROS, as well as the ability of cells to prevent ROS from reaching damaging levels through synthesis of antioxidants, including flavonols. The relationship between locations of ROS accumulation and ABA signaling and the role of enzymatic and small molecule ROS scavengers in maintaining ROS homeostasis in guard cells are summarized in this review. Understanding the mechanisms of ROS production and homeostasis and the role of ROS in guard cell signaling can provide a better understanding of plant response to stress and could provide an avenue for the development of crop plants with increased stress tolerance.

Keywords: abscisic acid; flavonols; guard cell; reactive oxygen species; respiratory burst oxidase homolog; stomata.

Figure 1

ABA increases ROS levels in guard cells in multiple subcellular locations. (A) A schematic model of the ABA signaling pathway during stomatal closure. Blue fonts represent proteins, while green fonts represent molecules. (B) Treatment with ABA for 45 min increases DCF fluorescence and decreases stomatal aperture in tomato guard cells. (C) The signal of the generic ROS sensor, DCF (green), is detected in the cytosol, nucleus, chloroplasts (pink), and endomembranes, with rapid and dramatic increases in all these locations in response to ABA treatment. RBOH enzymes (purple) produce ROS at the plasma membrane but can be internalized into endosomes. ROS signal also overlays peroxisome (orange), but whether this signal increases with ABA has not yet been reported. Central vacuole is not shown in illustration.

Figure 2

Arabidopsis and tomato mutants with decreased flavonoid antioxidants have increased ROS accumulation. (A) Confocal micrographs of DCF-stained guard cells of 4-week-old VF36 (wild-type) and are plants show that flavonol deficient mutants have increased ROS levels both in the absence or presence of 20 µM ABA. (B) Confocal micrographs of DCF and PO1 fluorescence in guard cells of 4-week-old tomato and Arabidopsis leaves show tomato and Arabidopsis with decreased flavonol levels have increased total ROS and H₂O₂ levels. Scale bars = 5 µm. DCF signal is shown in green, PO1 signal in blue, and chlorophyll autofluorescence in magenta. Images obtained from experiments completed in (Watkins et al., 2014; Watkins et al., 2017).