PYR/PYL/RCAR abscisic acid receptors regulate K+ and Cl- channels through reactive oxygen species-mediated activation of Ca2+ channels at the plasma membrane of intact Arabidopsis guard cells

• Published in: Plant physiology (Bethesda) 2013-10, Vol.163 (2), p.566-577
• Main Authors: Wang, Yizhou, Chen, Zhong-Hua, Zhang, Ben, Hills, Adrian, Blatt, Michael R
• Format: Article
• Language: English
• Subjects: Abscisic Acid - metabolism; Abscisic Acid - pharmacology; Anions - metabolism; Arabidopsis - cytology; Arabidopsis - drug effects; Arabidopsis - metabolism; Arabidopsis Proteins - metabolism; Calcium - pharmacology; Calcium Channels - metabolism; Cell Membrane - drug effects; Cell Membrane - metabolism; Chloride Channels - metabolism; Ion Channel Gating - drug effects; Ion Channels - metabolism; Mutation - genetics; Plant Stomata - cytology; Potassium Channels - metabolism; Reactive Oxygen Species - metabolism; Receptors, Cell Surface - metabolism
• ISSN: 1532-2548; 1532-2548
• DOI: 10.1104/pp.113.219758

The discovery of the START family of abscisic acid (ABA) receptors places these proteins at the front of a protein kinase/phosphatase signal cascade that promotes stomatal closure. The connection of these receptors to Ca(2+) signals evoked by ABA has proven more difficult to resolve, although it has been implicated by studies of the pyrbactin-insensitive pyr1/pyl1/pyl2/pyl4 quadruple mutant. One difficulty is that flux through plasma membrane Ca(2+) channels and Ca(2+) release from endomembrane stores coordinately elevate cytosolic free Ca(2+) concentration ([Ca(2+)]i) in guard cells, and both processes are facilitated by ABA. Here, we describe a method for recording Ca(2+) channels at the plasma membrane of intact guard cells of Arabidopsis (Arabidopsis thaliana). We have used this method to resolve the loss of ABA-evoked Ca(2+) channel activity at the plasma membrane in the pyr1/pyl1/pyl2/pyl4 mutant and show the consequent suppression of [Ca(2+)]i increases in vivo. The basal activity of Ca(2+) channels was not affected in the mutant; raising the concentration of Ca(2+) outside was sufficient to promote Ca(2+) entry, to inactivate current carried by inward-rectifying K(+) channels and to activate current carried by the anion channels, both of which are sensitive to [Ca(2+)]i elevations. However, the ABA-dependent increase in reactive oxygen species (ROS) was impaired. Adding the ROS hydrogen peroxide was sufficient to activate the Ca(2+) channels and trigger stomatal closure in the mutant. These results offer direct evidence of PYR/PYL/RCAR receptor coupling to the activation by ABA of plasma membrane Ca(2+) channels through ROS, thus affecting [Ca(2+)]i and its regulation of stomatal closure.